Date: Thu Jan 22 14:55:43 EST 2004
Name: Erik
Email:iepag@yahoo.ca URL:http:// Remote Host: yahoo.ca
Subject: Meaning
Bonjour Dr Zuker,
I'm totally novice in your field. Your web site is very user friendly and I appreciate your tutorial notes a lot. They were extremely helpfull to introduce myself to basics of such a large field.
I'm doing full-length RT-PCR and I was wondering if the difficulties I'm having with one of clones (aptNR2C) are due to the fact of stable conformation in the mRNA for that subunit. So, for fun, I came on your web site and tested my three clones (aptNR2A:/04Jan21-16-14-38/, aptNR2B:/15/04Jan21-15-40-59/ and aptNR2C:/11/04Jan21-11-27-22/).
My interest was to verify if aptNR2C has a more stable conformation than the two other clones. I checked the most stable conformational structure for the three of them. Here's are the global values for these most stable structures:
What should I understand from these? What does "initially" mean? Does dG mean "deltaG"? If so, why aptNR2C has a positive value? Shouldn't be an unstable conformation which unlikely happens at this temperature and conditions? I missed that in your tutorial notes? Actually, could I compared the data in such a way? Is it legitimate? I think I understood that the minimum energy allow to calculate the probability of occurance of a given conformational structure for a given RNA sequence. However, my interest is to compare the conformational structure stability of different RNA molecules. Again, is it legitimate?
Well,...to my understanding of the situation, I think I'm lost! Could you help me find my way out?
=====
Erik Harvey-Girard
Université d'Ottawa
Laboratoire de Leonard Maler
Département de Médecine Cellulaire et Moléculaire, Faculté de Médecine
Local #2232, 451 Smyth Road, Ottawa (Ontario), Canada, K1H 8M5
(613) 562-5800 #8257
<;)))>< <;)))>< <;)))>< <;)))><
Date: Sun Jan 25 15:42:17 EST 2004
Name: Kayvan
Email:kayvan@mit.edu URL:http:// Remote Host: tang-five-ninety-one.mit.edu
Subject: MFOLD
Hello,
I entered my sequence by don't know how to make sense of the output. Could somebody spend a few minutes with me to help make sense of this data? I'd really appreciate it. I'de like to include the mfold results (if applicable) in a paper I'm planning to submit to Oncogene in 2-3 weeks.
Thank you,
Kayvan Zainabadi
617-680-8746
kayvan@mit.edu
Date: Thu Feb 5 10:24:07 EST 2004
Name: Email:mick@cict.fr URL:http:// Remote Host: ondine.cict.fr
Subject: OligoArray
I Have a problem with OlogoArrayAux
OligoArray 2.1.1 will start to process sequences from the file chr1.fas using the following parameters :
Blast database: 'yeast_orf.fas'
Oligo data will be saved in: 'oligo.txt'
Sequence without oligo will be saved in: 'rejected.fas'
The log file will be: 'OligoArray.log'
Maximum number of oligo to design per input sequence: '2'
Size range: '45' to '47'
Maximum distance between the 5' end of the oligo and the 3' end of the input sequence: '1500'
Minimum distance between the 5' ends of two adjacent oligos: '69'
Tm range: '82' to '88'
GC range: '35.0' to '50.0'
Threshold to reject secondary structures: '65.0'
Threshold to start to consider cross-hybridizations: '65'
Sequence to avoid in the oligo: 'AAAAA;GGGGG;CCCCC;TTTTT'
Number of sequence to run in parallel: '1'
Data initialization: DONE
Is yeast_orf.fas a valid Blast database? YES
Is OligoArrayAux installed? NO
Design aborted due to a failure in the test above Date: Fri Mar 5 06:00:59 EST 2004
Name: Agung
Email:trisetyarso@yahoo.com URL:http://www.agungtri.cjb.net Remote Host: 202.155.151.71
Subject: About
Dear Professor Michael Zuker,
My name is Agung Trisetyarso, I am a PhD student candidate at Research Center Juelich Biophysics group. I have a strong background in theoretical physics and lack of biology knowledge.
I have read a lot of information from your website and it is very useful for my PhD research.
I interested in quantifying properties of biochemical network. The ambitious goal of my PhD research is to build a self-consistent theory of cell locomotion.
Sincerely yours,
Agung Trisetyarso
-----------------
http://www.agungtri.cjb.net
Date: Fri Apr 16 15:44:02 EDT 2004
Name: Paul
Email:pgottl@med.cuny.edu URL:http:// Remote Host: 134.74.54.158
Subject: salt
The server does not allow the NaCl concentration to be changed. Is there a way to do this? M. Zuker replies:
You can change the mono-valent ion concentration (Na+, Li+, K+ or NH4+) and the di-valent ion conentration (Mg++ or Ca++) for DNA only. We have no such corrections for RNA. However, the RNA folding parameters have been determined at [Na+] = 1M and [Mg++] = 0M, which actually mimic physiological conditions.
Date: Tue Apr 20 12:22:16 EDT 2004
Name: Mike
Email:portereiko@biology.utah.edu URL:http:// Remote Host: biol-81.biology.utah.edu
Subject: mfold
I used your program to fold an RNA for me, and I got a returned document entitled nph-mfold-3.1.cgi. How do I open this document? I click on it, and it opens my
endnote program!
I am on a Mac with osx.
Thanks. M. Zuker replies:
According to my log files, you submitted 4 sequences today. All were
folded successfully and the results have not yet been erased (22:07,
April 20, 2004). You may always view your results if you have a static
IP address by using the following link. Your problem has nothing to do with
using a Mac, but everything to do with using the crappy Safari
Internet browser. However, you used the Safari browser with only the
first folding. The others were made using MicroSoft Internet Explorer
version 5.22. You should not have had problems in the last three
cases. Did you?
I will ask my systems person, Alex Yu yua@rpi.edu, to look into this
problem. You are the second person who has reported such a problem. We
may have to change things at this end, or you might need to change
settings on your browser.
April 21: The mfold web-server script
will now deal with submissions from people who use the Safari web
browser.
Date: Thu Apr 29 07:18:29 EDT 2004
Name: Steen
Email:swknudsen@biostud.ku.dk URL:http:// Remote Host: pc2006.aki.ku.dk
Subject: Output
Thanks for a very nice homepage.
I have tried to fold some 16S sequences in various ways, and I'm impressed by the variety of output-formats.
But I have not been able to find an output-format in which the structure is not shown in a figure, but just presented as a simple linear text-string, with colored bases indicating loops and stems in the structure.
M. Zuker replies:
I can't easily give you colored bases, but I can easily give you a
simple linear text string showing the sequence followed by another simple
linear text string that gives base pair information. This is known as
the Vienna format.
When using 16S sequences in a phylogeny-analysis, you are often interested in the secondary structure of the sequences in order to better align the sequences, in order to make a better phylogeny.
Mfold gives me the structure of my sequence - but it's a hard job printing all structures for every taxa, and then afterwards marking regions of bases in my sequence-text-file corresponding to loops and stems.
My software is a very poor tool to compute ab initio foldings
of entire rRNAs. I suggest that you familiarize yourself with what Robin Gutell has to offer. In particular, you should
register at his Comparative RNA Web Site.
I don't know if this feature I request is allready avialable in mfold.
The Vienna format is available as of today.
If not. You should surely consider making this output-format available. The file format could be a simple RTF or word-file, in which the sequence is presented as a linear text-string with different background-colorings denoting bases which are likely/unlikely to form stems.
If this feature is allready available, please let me know how.
What you request is harder, and it will have to wait until I can raise
money specifically to support this web server. Up to now, I have used
some resources from an existing grant and my University has been very
helpful as well.
Thanks
Best Regards
Steen Knudsen (master student)
Date: Fri Apr 30 05:23:46 EDT 2004
Name: claire
Email:claire_hoede@yahoo.fr URL:http:// Remote Host: st159-10.bichat.inserm.fr
Subject: Trouble
Hello Mr M. Zucker,
I am writing a mail because I have got some problem to install mfold-3.1 for Mac OS X.
Indeed, when I install, it doesn't succeed to copy a file. I present below the display that I obtain:
M. Zuker replies:
Your problem seems to be simple. Either the directory
"/Users/apple/bin" does not exist or else you do not have permission
to write into that directory. You could do this as superuser, or else
you could copy all executable files into another directory that is on
your exec path (contained in the environment variable $PATH).
Date: Mon May 3 18:17:35 EDT 2004
Name: Jian
Email:jian@lanl.gov URL:http://www.bioinfo.rpi.edu/applications/mfold/old/rna/form5.cgi Remote Host: wwwcache.lanl.gov
Subject: Using
Dear Dr. Zuker,
Thank you for the wonderful tools you have developed for predicting the fold of RNA and DNA. I am writing a program to design probes for our microarray study. One of the tasks is to eliminate that probes that form stable hairpin with Tm > 50 degree. I found your Tm server does exactly what I want to do. But can I use your code in my program instead of using your webserver?
Thank you very much!
Jian Date: Wed May 5 07:48:11 EDT 2004
Name: Ebbe
Email:esa@mb.au.dk URL:http://www.rna.dk/esa/ Remote Host: mbio-38.mbio.au.dk
Subject: RNAML
Dear Michael,
It would be nice if you could provide RNAML as an output file for each structure. I would like to use the RNAML file to annotate the RNA structure and to import the structure in RnaViz.
Best regards,
Ebbe
Date: Tue May 25 07:24:26 EDT 2004
Name: roya
Email:roya1969@hotmail.com URL:http:// Remote Host: 81.91.144.61
Subject: rna
Hi dear sir (or as a muslim salam dear sir !) it was my good luck to find yr interesting site it was really helpful for my lecture ,i am an iranian woman and also doing my m.s in genetics. and i needed something about RNA structure,anyway i am thankful of your reply ,it was my proud, and my lecture will indebt of you! best regard-Roya
Date: Tue May 25 18:55:06 EDT 2004
Name: john
Email: URL:http://www.optimates.us/robyn-matthew.htm Remote Host: adsl-66-142-43-153.dsl.kscymo.swbell.net
Subject: nice site
Date: Wed May 26 11:36:10 EDT 2004
Name: Tony
Email:xxu@danforthcenter.org URL: Remote Host: 65.254.111.250
Subject: batch
Dear Dr. Zuker,
Very nice server!
If I run Mfold for RNA sequences (short), I can obtain "jpg" file for the prediction,
which is very nice for visualization. However, when if I run Quickfold for a batch of
short RNA sequences, I could not have any visualized result. What I could have is ct
file or text output. Is there any program I can use to convert the ct file or text output
into jpg format?
Thanks?
Tony
M. Zuker replies:
Yes. Download the ct file. You must split it into separate files for
each folding. You can produce images using my "sir_graph" program from
the "mfold_util" package which you may download from:
http://www.rpi.edu/~zukerm/export/.
Take: mfold_util-3.1.tar.gz This software currently works under:
Mac OS X: Darwin-Fink-X11, Darwin-Panther
PC: Windows-Cygwin, Windows-MingW
Linux: RedHat
SGI IRIX, 32 and 64 bit
Date: Sun Jun 6 00:09:55 EDT 2004
Name: Xu
Email:xuhy@genomics.org.cn URL:http:// Remote Host: 211.152.157.234
Subject: the
Dear Prof. Zuker:
I have downloaded the mfold-3.1.2(http://www.rpi.edu/~zukerm/mfold-3.1/all-OJb3E4D8krAAAAlp3yY/)and made it run successfully on Redhat Linux 9.0. Now through rewriting MAXN=16383 in Makefile and MAXN.inc, I know that RNA sequence length=16383bases(i think the number is based on 1024*16bit) is the upper limit that the wonderful software can do. But the problem is that I need to deal with RNA sequence length>=30000bases, and I have been trying sorts of different ways but failed. Would you give me some advices? Thanks a lot!!
M. Zuker replies:
The problem with 16bit (short) integers is not the sequence length. The sequence length is stored as a 32bit integer. It is the enengies, expressed as integers, that cause problems. Once folding energies approach 3,200 kcal/mole, problems begin. In "Makefile", you must set MAXN2 to be the maximum sequence size. This means that only "nafold2" should be used for folding very large sequences. One solution is a slight alteration in the "mfold" script. Change:
# Choose folding program
if [ $LENGTH -le 800 ]; then
PROG=nafold2
else
PROG=nafold
fi
to
# Choose folding program
PROG=nafold2
In order to fold about 30Kb (such as the SARS virus), you will need a 64bit computer with a lot of RAM. Good luck.
Date: Wed Jun 9 06:49:09 EDT 2004
Name: Daen
Email:guestbooks@daen.dk URL:http://www.daen.dk Remote Host: nuevolution.com
Subject: Linköping
Hi - great to meet you in Sweden - very much enjoyed the talk, although I'm not entirely clear on your explanation of the database search algorithm that you gave - kindest regards Daen
Date: Wed Jun 9 12:34:15 EDT 2004
Name: Odd
Email:odd_bres@hotmail.com URL:http:// Remote Host: lab12.zoo.umanitoba.ca
Subject: thanks
Dear Sirs, thank you for making mfold available on the www. It is well set up and it ran fast. I used it to identify a putative selenocysteine insertion element on a cDNA for a deiodinase gene in trout. I'll make the appropriate citation.
Odd Bres, Ph.D.
Department of Zoology
University of Manitoba
Winnipeg, Manitoba, CANADA R3T 2N2
obres@cc.umantitoba.ca
204 474 9245
Date: Sun Jun 13 06:35:46 EDT 2004
Name: Gérard
Email:gniyonizigiye@hotmail.com URL:http:// Remote Host: d51a5e345.kabel.telenet.be
Subject: Température
Bonjour Monsieur le Professeur et merci pour les informations que vous nous donnez à travers vos pages Web.
En effet, j'aimerais différentier deux séquences par la technique PCR basée sur l'ELISA en utilisant un oligonucléotide qui va reconnaître la séquence spécifique. Mais comme les deux séquences présentent une forte homologie, la sonde spécifique d'une longueur de 17 paires de bases avec seulement une différence de 2 bases a été choisie:
actggcgcagatggacc.
A quelle température d'hybridation dois-je faire l'incubation pour la différentier avec: actggcgcagatgtaca
Merci
Date: Mon Jun 14 17:00:36 EDT 2004
Name: Johnny
Email:lokecj@muohio.edu URL:http:// Remote Host: ip134-053-027-184.s27.muohio.edu
Subject: mFold
Dear Dr. Michael Zuker,
I am a grad student here at Miami University, Ohio studying Arabidopsis polyadenylation signals. I have came to you site to fold some RNA sequences, and I have about 1000 sequences, the problem with my summission using your quickfold is that it does not give jpg images output. Is there a way I can do a big summission with jpg image output?
Please advice. I appreciate for your help in this.
Thanks.
Johnny Loke
Date: Wed Jun 23 13:35:19 EDT 2004
Name: chabane
Email:tibichec@iro.umontreal.ca URL:http:// Remote Host: tweed.iro.umontreal.ca
Subject: mfod
Hye!
My comment is that when I enter a sequence where there is a mix of upper and lower case characters, the output is don't respect the case of the input sequence and give the result only in upper case characters.
Thanks
Date: Sat Jun 26 14:02:12 EDT 2004
Name: Xu
Email:xuhy@genomics.org.cn URL:http:// Remote Host: 211.152.157.234
Subject: puzzle
Dear Prof. Zuker,
Thank you very much for your replying last time. Now I am also puzzled by two questions: (1)why in /mfold-3.1.2/samples the energy value in mdv1.log is different from it in mdv1_*.gif, for example, the minimum energy (that is shown in the bottom of mdv1_1.gif)is dG=-143.45(initially -145.3),but in the mdv1.log, however, it(also mdv1_1) is dG=-139.86(initially -141.8)? In my opinion, they would have been the same value, because they are calculated for the same structure mdv1_1. (2) during installing mfold-3.1.2 under Red hat Linux 9.0 and 32bit CPU and 2.0G RAM, I found that if in Makefile£¬I set maxn(nafold)>22612 or maxn1(quickfold)>16383 or maxn2(nafold2)>16182, after ¡°make all¡±£¬the screen shown ¡°bigstuff is too large¡±£¬and installing failed. I wonder how the three different numbers are come from, and why they seem a bit strange? Thanks a lot!! Date: Wed Jun 30 09:11:36 EDT 2004
Name: Verónica
Email:mvr@agro.uba.ar URL:http:// Remote Host: ol168-193.fibertel.com.ar
Subject: thanks!
I used your program to analize cDNA secondary structure of a sorghum gene and obtained a very complicated shape; I will use this information to design new primers for real time pcr, I hope this time things will work! Thanks to all the people who make this software available on the internet.
Kind regards,
Verónica
Date: Sat Jul 10 07:01:30 EDT 2004
Name: mr
Email:aba_agu@yahoo.com URL:www.aba Remote Host: 62.56.189.138
Subject: good
i love this guest page.
Date: Wed Jul 14 11:42:53 EDT 2004
Name: Fernando
Email:fgomez318@aol.com URL:http:// Remote Host: b12-arbiter-d.net.nih.gov
Subject: Great
Dear Fellows;
I just wanted to drop you a note letting you know that I really enjoy using your RNAfolding service. I use the nucleic acid folding option when I want to see if my siRNA duplex components are going to form any energetically favorable secondary loops, and therefore should not be used for the siRNA KO I am designing. However, just for clarification, I wanted to confirm that the "dG" indicated in the thermodynamic profile you guys give when analyzing a particular sequence means "delta G". I ask this because other sites give delta E. Is "dG" delta G, and how should I view this value differently from the delta E value given in other sites?
Thanks for taking the time to answer my questions. I really appreciate it!
Sincerely,
Fernando Gomez
Summer Intern, National Institues of Aging M. Zuker replies:
I don't know what other sites do and I don't know what "delta E" is supposed to be. However, our "dG" is "delta G", which is "free energy".
Date: Thu Jul 15 10:36:18 EDT 2004
Name: Email:saskia.rutjes@rivm.nl URL:http:// Remote Host: proxy02.rivm.nl
Subject:Date: Tue Aug 3 21:05:05 EDT 2004
Name: Gabriela
Email:golmedo@ira.cinvestav.mx URL:http:// Remote Host: ciea.ira.cinvestav.mx
Subject: suggestion
I wish there was a second button to start RNA folding right next to the box where the
sequence is paste. When testing many RNA segments with the same parameters I hate to
scroll down from the box to the clickable fold RNA botton.
Date: Wed Aug 11 16:41:24 EDT 2004
Name: Dr
Email:bourdonv@mskcc.org URL:http:// Remote Host: 140.163.188.72
Subject: mfold
Hi
I need to send a batch of short sequences to be fold and to get back the vienna file of each structure obtained. I can not find how to do, is it possible?
thanks
Date: Wed Aug 18 07:20:57 EDT 2004
Name: Nikolai
Email:ns01@ic.ac.uk URL:http:// Remote Host: ic.ac.uk
Subject: Folding
Dear Prof. Zuker,
We are currently working on a novel nucleic acid-based diagnostic assay involving the formation of DNA/RNA 3-way junctions. I was wondering if you are aware of any software capable of modelling such hybridisations between 3 or more single stranded nucleic acid sequences.
Many thanks,
Nikolai Szep
Date: Fri Aug 27 13:30:16 EDT 2004
Name: Matthew
Email:mrounseville@htgenomics.com URL:http:// Remote Host: htgenomics.com
Subject: Batch
Dear Dr Zucker,
I have been using your algorithms since the late 1980's when I was working on the structure of HIV TAR RNA. Thank you for your exceptional contributions to the scientific community.
I have been using your hybridization server to screen for unwanted hybridizations between oligos in our microarrays. The program is very useful, however, it is rather tedious because I need to enter each pair of sequences one set at a time. Is there any way to run this program in batch mode where I could evaluate potential cross-hybridizations between multiple pairs of oligos?
Regards,
Matt
Date: Fri Sep 3 09:27:16 EDT 2004
Name: acox30
Email:acox30@hydromail.com URL:http:// Remote Host: pcp06940631pcs.nrockv01.md.comcast.net
Subject: site
This is an awesome site
Date: Tue Sep 7 03:51:23 EDT 2004
Name: Huajiang
Email:weihuajiang@tsinghua.org.cn URL:http:// Remote Host: 211.151.91.194
Subject: how
My major work is probe deisgn and hybridization data analysis. Our probe is designed for specific purpose instead of gene expression and our probe and their target sequences are very short, about or more than 50 bp. Specificity is quite import to us and cross hybridization should be avoid! Althrough Sequence alignment and similiarity can be criterion to score specificity, I think hybridization energy is more reasonable.
2-state hybridization is part of mfold 3.1, but I can't find it and use it in mfold 3.1. could you tell me how to use 2-state hybirdization through mfold!
Thank you!
Best wishes
----------
Department of Biological Sciences & Biotechnology,
Tsinghua University,
Beijing 100084, P.R. China
Date: Tue Sep 7 19:36:45 EDT 2004
Name: Changbong
Email:hyeoncb@glue.umd.edu URL:http:// Remote Host: rnadream.umd.edu
Subject: Installing
Dear Prof. Zuker:
I installed mfold-3.1.2 under /usr/local/lib in my local linux machine.
I set the variable in Makefile as
and created directory called MFOLD in my home directory.
The installation seemed to be fine.
But when I run the mfold giving a sequence in FASTA format as
mfold SEQ='1GID.fasta'
I got the following results.
cat: /dat/begin.dat: No such file or directory
REUSE= NO
cp: `1GID.fasta' and `./1GID.fasta' are the same file
1GID.fasta.pnt created.
Sequence length is 159
Folding at 37 degrees using version 3.0 dat files.
Save file created using nafold2.
Minimum folding energy is -63.1 kcal/mole.
Energy increment is 3.155 kcal/mole.
H-num file created from plot file.
1,
Suboptimal foldings created.
Energy dot plot created.
cp: cannot stat `/dat/*.dat': No such file or directory
miscloop missing
One of the data files is missing
Free energies re-evaluated using efn2 and added to ct file.
cat: /dat/bases.nav: No such file or directory
Structure plots generated.
All done.
I'm concerned about the lines with
"cat: /dat/begin.dat : No such file or directory"
and
"cat: /dat/bases.nav: No such file or directory"
Is anything wrong with the modification in Makefile?
I tried another fasta file which looks like the following
>1F27:A RNA
ACCGUCAGAGGACACGGUU
but in this case mfold SEQ='1F27.fasta gives the output as follows.
cat: /dat/begin.dat: No such file or directory
REUSE= NO
cp: `1F27.fasta' and `./1F27.fasta' are the same file
1F27.fasta.pnt created.
Sequence length is 19
Folding at 37 degrees using version 3.0 dat files.
Save file created using nafold2.
Save file is empty. No foldings.
Job Aborted
I guess I made a simple mistake when I set up the path in Makefile.
I would appreciate you for the help in advance.
Changbong Date: Sun Sep 12 10:30:11 EDT 2004
Name: Mr.sarayoot
Email:sarayoot@dmsc.moph.go.th URL:http:// Remote Host: 203.157.14.246
Subject: Thank
This is a second time for to use your software for folding analysis. The first time is to fold the RNA sequence. This software
are very useful for me including there are many analuzed information for further analysis.
Thank you very much of your kindly provinding me the help.
Mr. Sarayoot Subpasu
sarayoot@dmsc.moph.go.th
ssubpasu2004@yahoo.com Date: Mon Sep 13 16:40:04 EDT 2004
Name: Roland
Email:rgaboury@inuktun.com URL:http:// Remote Host: s0106000d88f1df8b.no.shawcable.net
Subject: Cycling...
Hey... who's that big old guy you're standing next to on the bridge?
M. Zuker replies:
Thanks for the flattery. The picture above is what I actually looked like in 1990. I see by your E-mail address that you're back in Candada. In fact, you're back to your home town. I'll visit if I ever get to BC. I have an open invitation to visit the CS Department at UBC. I'm glad you made contact.
Date: Mon Sep 13 18:02:46 EDT 2004
Name: Wei
Email:wei.guo@viagen.com URL:http:// Remote Host: yahoo.com
Subject: Automatic
Hi Professor Zuker,
My name is Wei Guo, a researcher at a small biotech start-up Viagen. I am wondering if I can use your DNA mfold server to test 2nd structures of a bunch of PCR primers. I want to write a script to make automatic requesting using url-encoded data, such as http://www.bioinfo.rpi.edu/applications/.../form1.cgi?sequence=actggcccaattttccaaggcttacc&email_addr=wei.guo@viagen.com&action=fold
Particularly I need to know what the key-value pair is returned when the Fold DNA button is clicked to submit a request.
Your time and help is greatly appreciated.
Sincerely,
Wei Guo M. Zuker replies:
Many users submit sequences or groups of sequences automatically to my
various servers. Your message indicates that you do not understand
how to do this properly. I will give you some information and
suggestions.
You are confusing a cgi script that generates a FORM with
the cgi-script that processes the information submitted by that
form. In the case of RNA and DNA folding, the request should be sent to
"http://www.bioinfo.rpi.edu/applications/old/cgi-bin/nph-mfold-3.1.cgi"
and not to a cgi script that generates a form.
The form page shows clearly what the "NAME" and "VALUE" pairs
are. The "NAME"s are case sensitive. No deviation is permitted. If you
use, for example, "sequence=ctagggcgact" instead of
"SEQUENCE=ggctagcgactttg", the script will act as if no sequence was
submitted. In other cases, you may end up with default values even
though you think that you have set a parameter. For example, in DNA
folding, if you use "t=50" instead of "T=50" for setting the
temperature, then "t=50" will be ignored and you will end up with
folding at 37° C, which is the default. In cases where the values
are rigidly defined within the form, any deviation in spelling or case
will generate an error or a default value that you may not wish.
Automatic requests using the "GET" protocol are automatically
blocked in the "nph-mfold-3.1.cgi" script. You must use "POST"
instead.
The cgi script that handles full RNA and DNA folding generates
images of foldings and a web page that is meant to be used
interactively to examine results in details or to redraw foldings or
parts of foldings. It seems that you do not wish to work interactively
with the results or to produce images of foldings. If so, then you
should use more appropriate servers.
"quikfold" server: Fold many RNA or DNA sequences in a single
run. Optimal and suboptimal foldings are predicted. No images.
"zipfold" server: Predict the minimum folding energies for many
RNA or DNA sequences. No foldings are computed.
"Tm server: Predict the minimum folding energies for
many RNA or DNA sequences together with the enthalpy and entropy of
minimum energy foldings and a melting temperature based on a simple
2-state model. No foldings are shown although they could easily be
made available.
You should read:
M. Zuker
Mfold web server for nucleic acid folding and hybridization prediction. Nucleic Acids Res.31 (13), 3406-15, (2003) [Abstract][Full Text][Supplementary Material][Additional
Information] I apologize for my error in the Tm formula for
2-state hybridization. A correction appears under the "Additional
Information" link.
Finally, the "quikfold" server needs a password (the NAME is
"VERIFY") that changes daily and can only be found within the form
generated by
"http://www.bioinfo.rpi.edu/applications/mfold/old/rna/form3.cgi".
This was added to discourage use of the server by commercial software
packages.
Date: Wed Sep 15 12:09:43 EDT 2004
Name: Sarah
Email:snahmias@alumni.princeton.edu URL:http:// Remote Host: 146.203.53.105
Subject: scoring
When the energy is calculated, a number is given, but it says [initially....] what does that mean? Why has the number changed?
thanks
Sarah M. Zuker replies:
For RNA folding, versions 3.x
of mfold use free energy rules as described in:
D.H. Mathews, J. Sabina, M. Zuker & D.H. Turner
Expanded Sequence Dependence of Thermodynamic Parameters Improves
Prediction of RNA Secondary Structure
J. Mol. Biol. 288, 911-940 (1999)
The rules for multi-branch loops are too complicated to be used in an
efficient algorithm. Therefore, the folding is done using slightly
simpler rules. This gives the "initial free energies". The free
energies are then re-evaluated using the best rules that we have.
In particular, the revised energies are determined by:
using Jacobson-Stockmeyer theory to assign free energies to
multi-branch loops that includes a term that grows logarithmically
with the number of unpaired bases in the loop,
and by computing coaxial stacking of adjacent helices in multi-branch
loops or in the "exterior" loop. This latter feature has been
implemented by David Mathews in his RNAstructure and related software.
The re-evaluated energy is what is given as "dg = ..." in the figures.
The only purpose of this re-evaluation is to re-order the foldings
according to "better" energies. We reported that this improves folding
predictions. That is, the optimal folding according to the revised energy
rules tends to contain more "correct" base pairs.
Date: Wed Nov 17 08:41:18 EST 2004
Name: sujun
Email:zheng003@sina.com URL:http:// Remote Host: sina.com
Subject: dear Michael Zuker
Thanks for your web service of RNA folding !
about 2 days ago,i have input twuo rna sequence for rna strcuture rediction .I appled for the batch processing form .but i have not receive the results.Can you help me?
your friend
sujun zheng M. Zuker replies:
According to the log files, you submitted six large folding jobs recently. The four jobs submitted at around 04:00 EST on November 15 have already been erased. The two jobs submitted at round 08:00 EST on November 17 are still on the system and may be found at: 08-17-20 and here. Both jobs finished in less than 15 minutes. You should receive E-mail from the server when your job is complete. However, when you submit a batch job, you are given the URL for the results. You should check this URL after a hour or so. Do not forget to refresh the page, if necessary.
Date: Wed Nov 17 13:14:18 EST 2004
Name: Kordai
Email:M.Sowa@ed.ac.uk URL:http:// Remote Host: ed.ac.uk
Subject: RNAfold
Hi,
The server keeps telling me to do a batch job, (even though I have chosen batch job) and it
won't fold my 1499 bases. Could you please tell me what to do to rectify this? (it says I
can fold up to 6000 bases as a batch job, so I'm not over the limit). Thanks.
Kordai
Date: Sun Nov 28 11:17:58 EST 2004
Name: LY
Email:winecar@sohu.com URL:http:// Remote Host: 220.163.54.191
Subject: The
I have used Mfold to abtain RNA secondary structures of genus Thermus species. Most of them were agreeable with the model published in 1993 made using 16S rRNA of E.coli. But when folding 16S rRNA of T.scotoductus, most of its strains lost their Helix11, which could be shown in windows version of Mfold -RNASTRUCTURE 4.1. How should I resolve such problem?
Great appreciation to you for your reply.
Thank you! M. Zuker replies:
About the worst thing to do with an RNA folding program that folds
single sequences based on minimizing free energy is to expect it to
predict a correct folding for a large molecule without any additional
information. This is all the more true for small and large subunit
rRNA, where ribosomal proteins probably serve as guides or chaperones
in the folding process. mfold, RNASTRUCTURE and other such programs
attempt to simulate equilibrium conditions in solutions in the
absence of other molecules.
It is remarkable that mfold gives good answers for RNA viruses, in the
sense that separate folding domains are often robustly predicted and
many portions of predicted secondary structures are "well-determined"
in the sense that there are few alternatives for those predictions. It
is also remarkable that a minimum energy folding of, for example,
Thermococcus celer 16S rRNA contains about 80% of the base pairs
determined from phylogenetic methods. Predictions for rRNA of
Eubacteria are not nearly as good. Predictions for Eukaryotes are much
worse and those for mitochondrial rRNA are total nonsense. In joint
publications with the Turner group, you should note that we have not
attempted to fold entire rRNAs. For small subunit rRNA, the sequence
is broken into known domains of size up to about 500 bases and these
are folded separately.
If you wish to determine the secondary structure for, I presume, a new
rRNA, or at least one whose structure is unpublished or otherwise not
available in a public database, then by all means use mfold, but add
constraints that force certain known interactions or that exclude
incorrect foldings. Eukaryotes often have extra regions in their rRNA
that are presented as unstructured. It is appropriate to use mfold to
determine foldings for such inserted regions for which phylogenetic
data are either absent or insufficient to determine secondary
structure.
Finally, although RNASTRUCTURE (by David Mathews) began as a
"translation into C++" of the RNA folding program (nafold) in the
mfold package, it has been evolving more or less independently of my
old code and the new software being developed by my student, Nick
Markham, for folding and hybridization prediction. It is not realistic
to expect identical predictions.
Date: Thu Dec 2 18:53:36 EST 2004
Name: Ming
Email:mwu@qdots.com URL: Remote Host: 68-123-139-98.ded.pacbell.net
Subject: Thermodynamic
Dear Prof. Zuker,
Would it be possible to show dH and dS for the RNA mfold? Currently only free energy dG is shown. dH and dS will help to determine Tm and equilibrium constant K at different temperatures. What about salt effect on dH and dS for RNA folding?
Thanks.
Ming M. Zuker replies:
I cannot offer dG and dS for RNA folding (or hybridization) using the
latest, version 3 free energy rules at 37° C because they are not
available. It is for this reason that I offer an alternative for RNA
folding that uses the older, version 2.3 parameters, for which
enthalpies are also available. This site, RNA mfold version 2.3 server, is linked from the
default RNA folding form, which uses version 3 free energies. Use
version 2.3 RNA folding only if you really want to predict
Tm as well as dH and dS, or if you wish to investigate how
folding changes with increasing temperature. However, if you attempt
folding RNA from a thermophile by setting the temperature to 70°
C or higher, you will get nonsense.
Salt concentrations affect dS only. I have corrections for DNA folding
and hybridization which depend on the number of phosphate pairs in
hybridized regions. These corrections are independent of GC content
and the subject of some controversy. I have no such corrections for
RNA. However, the RNA folding parameters, derived for
[Na+] = 1 M and [Mg++] = 0 M,
corresond to physiological conditions where [Na+] =
125±25 mM and [Mg++] = 10±5 m M.
In the future, I hope to have a more comprehensive set of RNA
parameters that will permit melting predictions and corrections for
salt concentrations. They may come from my colleagues and long term
collaborators at the University of Rochester (Doug Turner's lab and
David Mathews) or from a company such as DNA Software, which
produces high quality commercial software based on theory and
measurements that is elegantly packaged, user friendly, and oriented
to the solution of specific problems.
Date: Thu Dec 2 22:21:34 EST 2004
Name: Du
Email:tonydudu@21cn.com URL:http:// Remote Host: pku.edu.cn
Subject: about
It seems that the server is down. Would you please have a check on it?
In addition, is the local version of hybridization server available?Thank you!
Date: Wed Dec 15 10:57:53 EST 2004
Name: diederik
Email:wdsmilde@casema.nl URL:http:// Remote Host: 535300be.cable.casema.nl
Subject: comments
Great stuff this is, but is it really necessary to type in my email address every time I submit another small job?
M. Zuker replies: I agree. I have removed the annoying demand for an E-mail address for DNA folding. The server will continue to demand an E-mail address for batch jobs. In these cases, a message is sent informing the user that the job is complete.
Date: Wed Dec 22 23:50:52 EST 2004
Name: Mr.Sarayoot
Email:sarayoot@dmsc.moph.go.th URL:http:// Remote Host: 203.157.14.245
Subject: Thank
Dear Prof. Michael ZUKER
I have no comment, however, I thank you very much for your kingly providing me the tools for DNA folding analysis. This very useful for predicting the Genotyping Analysis via SSCP.
Your Sincerely
Mr.Sarayoot Subpasu
Enteric Bacteriology Laboratory
National Institute of Health
Department of Medical Sciences
Minstry of Public Health
Tivanont, Nonthaburi 11000, THAILAND.
Name: Erik
Email: iepag@yahoo.ca
URL: http://
Remote Host: yahoo.ca
Subject: Meaning
Bonjour Dr Zuker,
I'm totally novice in your field. Your web site is very user friendly and I appreciate your tutorial notes a lot. They were extremely helpfull to introduce myself to basics of such a large field.
I'm doing full-length RT-PCR and I was wondering if the difficulties I'm having with one of clones (aptNR2C) are due to the fact of stable conformation in the mRNA for that subunit. So, for fun, I came on your web site and tested my three clones (aptNR2A:/04Jan21-16-14-38/, aptNR2B:/15/04Jan21-15-40-59/ and aptNR2C:/11/04Jan21-11-27-22/).
My interest was to verify if aptNR2C has a more stable conformation than the two other clones. I checked the most stable conformational structure for the three of them. Here's are the global values for these most stable structures:
aptNR2A => dG= -1792.2 [initially -1907.2]
aptNR2B => dG= -1892.48 [initially -2023.3]
aptNR2C => dG= 98492.62 [initially -1622.9]
What should I understand from these? What does "initially" mean? Does dG mean "deltaG"? If so, why aptNR2C has a positive value? Shouldn't be an unstable conformation which unlikely happens at this temperature and conditions? I missed that in your tutorial notes? Actually, could I compared the data in such a way? Is it legitimate? I think I understood that the minimum energy allow to calculate the probability of occurance of a given conformational structure for a given RNA sequence. However, my interest is to compare the conformational structure stability of different RNA molecules. Again, is it legitimate?
Well,...to my understanding of the situation, I think I'm lost! Could you help me find my way out?
=====
Erik Harvey-Girard
Université d'Ottawa
Laboratoire de Leonard Maler
Département de Médecine Cellulaire et Moléculaire, Faculté de Médecine
Local #2232, 451 Smyth Road, Ottawa (Ontario), Canada, K1H 8M5
(613) 562-5800 #8257
<;)))>< <;)))>< <;)))>< <;)))><
Name: Kayvan
Email: kayvan@mit.edu
URL: http://
Remote Host: tang-five-ninety-one.mit.edu
Subject: MFOLD
Hello,
I entered my sequence by don't know how to make sense of the output. Could somebody spend a few minutes with me to help make sense of this data? I'd really appreciate it. I'de like to include the mfold results (if applicable) in a paper I'm planning to submit to Oncogene in 2-3 weeks.
Thank you,
Kayvan Zainabadi
617-680-8746
kayvan@mit.edu
Name:
Email: mick@cict.fr
URL: http://
Remote Host: ondine.cict.fr
Subject: OligoArray
I Have a problem with OlogoArrayAux
trace of the problem
OligoArrayAux installed? NO
Why
˙y
sh -x /usr/local/bin/OligoArray -i chr1.fas -d yeast_orf.>
+ export HYBRIDLIB=/usr/local/OligoArray2_1/OligoArrayAux-1.7/Dat
+ export PATH=/usr/local/blast:/users/sysadmin/mick/bin:/usr/local/bin:/usr/sbin:/usr/bsd:/sbin:/usr/bin:/usr/bin/X11::/usr/local/bin:/usr/bsd:/usr/bin/X11:/usr/craysoft/nqe/bin:/usr/java/bin
+ export CLASSPATH=/usr/local/OligoArray2_1
+ java -jar /usr/local/OligoArray2_1/OligoArray2.jar -i chr1.fas -d yeast_orf.fas -o oligo.txt -n 2 -l 45 -L 47 -D 1500 -t 82 -T 88 -s 65 -x 65 -p 35 -P 50 -m AAAAA;GGGGG;CCCCC;TTTTT
*** OligoArray 2.1.1 ***
OligoArray 2.1.1 will start to process sequences from the file chr1.fas using the following parameters :
Blast database: 'yeast_orf.fas'
Oligo data will be saved in: 'oligo.txt'
Sequence without oligo will be saved in: 'rejected.fas'
The log file will be: 'OligoArray.log'
Maximum number of oligo to design per input sequence: '2'
Size range: '45' to '47'
Maximum distance between the 5' end of the oligo and the 3' end of the input sequence: '1500'
Minimum distance between the 5' ends of two adjacent oligos: '69'
Tm range: '82' to '88'
GC range: '35.0' to '50.0'
Threshold to reject secondary structures: '65.0'
Threshold to start to consider cross-hybridizations: '65'
Sequence to avoid in the oligo: 'AAAAA;GGGGG;CCCCC;TTTTT'
Number of sequence to run in parallel: '1'
Data initialization: DONE
Is yeast_orf.fas a valid Blast database? YES
Is OligoArrayAux installed? NO
Design aborted due to a failure in the test above
Name: Agung
Email: trisetyarso@yahoo.com
URL: http://www.agungtri.cjb.net
Remote Host: 202.155.151.71
Subject: About
Dear Professor Michael Zuker,
My name is Agung Trisetyarso, I am a PhD student candidate at Research Center Juelich Biophysics group. I have a strong background in theoretical physics and lack of biology knowledge.
I have read a lot of information from your website and it is very useful for my PhD research.
I interested in quantifying properties of biochemical network. The ambitious goal of my PhD research is to build a self-consistent theory of cell locomotion.
Sincerely yours,
Agung Trisetyarso
-----------------
http://www.agungtri.cjb.net
Name: Paul
Email: pgottl@med.cuny.edu
URL: http://
Remote Host: 134.74.54.158
Subject: salt
The server does not allow the NaCl concentration to be changed. Is there a way to do this?
M. Zuker replies: You can change the mono-valent ion concentration (Na+, Li+, K+ or NH4+) and the di-valent ion conentration (Mg++ or Ca++) for DNA only. We have no such corrections for RNA. However, the RNA folding parameters have been determined at [Na+] = 1M and [Mg++] = 0M, which actually mimic physiological conditions.
Name: Mike
Email: portereiko@biology.utah.edu
URL: http://
Remote Host: biol-81.biology.utah.edu
Subject: mfold
I used your program to fold an RNA for me, and I got a returned document entitled
nph-mfold-3.1.cgi. How do I open this document? I click on it, and it opens my
endnote program!
I am on a Mac with osx.
Thanks.
M. Zuker replies: According to my log files, you submitted 4 sequences today. All were folded successfully and the results have not yet been erased (22:07, April 20, 2004). You may always view your results if you have a static IP address by using the following link. Your problem has nothing to do with using a Mac, but everything to do with using the crappy Safari Internet browser. However, you used the Safari browser with only the first folding. The others were made using MicroSoft Internet Explorer version 5.22. You should not have had problems in the last three cases. Did you?
I will ask my systems person, Alex Yu yua@rpi.edu, to look into this problem. You are the second person who has reported such a problem. We may have to change things at this end, or you might need to change settings on your browser.
April 21: The mfold web-server script will now deal with submissions from people who use the Safari web browser.
Name: Steen
Email: swknudsen@biostud.ku.dk
URL: http://
Remote Host: pc2006.aki.ku.dk
Subject: Output
Thanks for a very nice homepage.
I have tried to fold some 16S sequences in various ways, and I'm impressed by the variety of output-formats.
But I have not been able to find an output-format in which the structure is not shown in a figure, but just presented as a simple linear text-string, with colored bases indicating loops and stems in the structure.
M. Zuker replies: I can't easily give you colored bases, but I can easily give you a simple linear text string showing the sequence followed by another simple linear text string that gives base pair information. This is known as the Vienna format.
When using 16S sequences in a phylogeny-analysis, you are often interested in the secondary structure of the sequences in order to better align the sequences, in order to make a better phylogeny.
Mfold gives me the structure of my sequence - but it's a hard job printing all structures for every taxa, and then afterwards marking regions of bases in my sequence-text-file corresponding to loops and stems.
My software is a very poor tool to compute ab initio foldings of entire rRNAs. I suggest that you familiarize yourself with what Robin Gutell has to offer. In particular, you should register at his Comparative RNA Web Site.
I don't know if this feature I request is allready avialable in mfold.
The Vienna format is available as of today.
If not. You should surely consider making this output-format available. The file format could be a simple RTF or word-file, in which the sequence is presented as a linear text-string with different background-colorings denoting bases which are likely/unlikely to form stems.
If this feature is allready available, please let me know how.
What you request is harder, and it will have to wait until I can raise money specifically to support this web server. Up to now, I have used some resources from an existing grant and my University has been very helpful as well.
Thanks
Best Regards
Steen Knudsen (master student)
Name: claire
Email: claire_hoede@yahoo.fr
URL: http://
Remote Host: st159-10.bichat.inserm.fr
Subject: Trouble
Hello Mr M. Zucker,
I am writing a mail because I have got some problem to install mfold-3.1 for Mac OS X.
Indeed, when I install, it doesn't succeed to copy a file. I present below the display that I obtain:
cp bin/batgen bin/distance bin/ct2rnaml bin/ct_compare bin/ct_boxplot_ng bin/efn bin/newtemp bin/scorer bin/auxgen bin/sav2plot bin/sav2p-num bin/h-num.exe bin/ss-count bin/add-dHdSTm bin/nafold bin/nafold2 bin/boxplot_ng bin/naview.exe bin/plt22ps bin/quikfold bin/plt22gif bin/genss bin/tgd bin/sav2plot2 bin/sav2p-num2 bin/add-dHdSTm2 bin/naview bin/mfold bin/auto_ct2ps bin/h-num bin/filter-sort bin/efn2 /Users/apple/bin
usage: cp [-R [-H | -L | -P]] [-f | -i | -n] [-pv] src target
cp [-R [-H | -L | -P]] [-f | -i | -n] [-pv] src1 ... srcN directory
make: *** [install] Error 64
Thank you for your help ...
Very cordially
Claire Hoede
NB : Sorry for my english...
M. Zuker replies: Your problem seems to be simple. Either the directory "/Users/apple/bin" does not exist or else you do not have permission to write into that directory. You could do this as superuser, or else you could copy all executable files into another directory that is on your exec path (contained in the environment variable $PATH).
Name: Jian
Email: jian@lanl.gov
URL: http://www.bioinfo.rpi.edu/applications/mfold/old/rna/form5.cgi
Remote Host: wwwcache.lanl.gov
Subject: Using
Dear Dr. Zuker,
Thank you for the wonderful tools you have developed for predicting the fold of RNA and DNA. I am writing a program to design probes for our microarray study. One of the tasks is to eliminate that probes that form stable hairpin with Tm > 50 degree. I found your Tm server does exactly what I want to do. But can I use your code in my program instead of using your webserver?
Thank you very much!
Jian
Name: Ebbe
Email: esa@mb.au.dk
URL: http://www.rna.dk/esa/
Remote Host: mbio-38.mbio.au.dk
Subject: RNAML
Dear Michael,
It would be nice if you could provide RNAML as an output file for each structure. I would like to use the RNAML file to annotate the RNA structure and to import the structure in RnaViz.
Best regards,
Ebbe
Name: roya
Email: roya1969@hotmail.com
URL: http://
Remote Host: 81.91.144.61
Subject: rna
Hi dear sir (or as a muslim salam dear sir !)
it was my good luck to find yr interesting site it was really helpful for my lecture ,i am an iranian woman and also doing my m.s in genetics. and i needed something about RNA structure,anyway i am thankful of your reply ,it was my proud, and my lecture will indebt of you!
best regard-Roya
Name: john
Email:
URL: http://www.optimates.us/robyn-matthew.htm
Remote Host: adsl-66-142-43-153.dsl.kscymo.swbell.net
Subject:
nice site
Name: Tony
Email: xxu@danforthcenter.org
URL:
Remote Host: 65.254.111.250
Subject: batch
Dear Dr. Zuker,
Very nice server!
If I run Mfold for RNA sequences (short), I can obtain "jpg" file for the prediction,
which is very nice for visualization. However, when if I run Quickfold for a batch of
short RNA sequences, I could not have any visualized result. What I could have is ct
file or text output. Is there any program I can use to convert the ct file or text output
into jpg format?
Thanks?
Tony
M. Zuker replies: Yes. Download the ct file. You must split it into separate files for each folding. You can produce images using my "sir_graph" program from the "mfold_util" package which you may download from: http://www.rpi.edu/~zukerm/export/.
Take: mfold_util-3.1.tar.gz This software currently works under:
Mac OS X: Darwin-Fink-X11, Darwin-Panther
PC: Windows-Cygwin, Windows-MingW
Linux: RedHat
SGI IRIX, 32 and 64 bit
Name: Xu
Email: xuhy@genomics.org.cn
URL: http://
Remote Host: 211.152.157.234
Subject: the
Dear Prof. Zuker:
I have downloaded the mfold-3.1.2(http://www.rpi.edu/~zukerm/mfold-3.1/all-OJb3E4D8krAAAAlp3yY/)and made it run successfully on Redhat Linux 9.0. Now through rewriting MAXN=16383 in Makefile and MAXN.inc, I know that RNA sequence length=16383bases(i think the number is based on 1024*16bit) is the upper limit that the wonderful software can do. But the problem is that I need to deal with RNA sequence length>=30000bases, and I have been trying sorts of different ways but failed. Would you give me some advices? Thanks a lot!!
M. Zuker replies: The problem with 16bit (short) integers is not the sequence length. The sequence length is stored as a 32bit integer. It is the enengies, expressed as integers, that cause problems. Once folding energies approach 3,200 kcal/mole, problems begin. In "Makefile", you must set MAXN2 to be the maximum sequence size. This means that only "nafold2" should be used for folding very large sequences. One solution is a slight alteration in the "mfold" script. Change: to In order to fold about 30Kb (such as the SARS virus), you will need a 64bit computer with a lot of RAM. Good luck.
Name: Daen
Email: guestbooks@daen.dk
URL: http://www.daen.dk
Remote Host: nuevolution.com
Subject: Linköping
Hi - great to meet you in Sweden - very much enjoyed the talk, although I'm not entirely clear on your explanation of the database search algorithm that you gave - kindest regards Daen
Name: Odd
Email: odd_bres@hotmail.com
URL: http://
Remote Host: lab12.zoo.umanitoba.ca
Subject: thanks
Dear Sirs, thank you for making mfold available on the www. It is well set up and it ran fast. I used it to identify a putative selenocysteine insertion element on a cDNA for a deiodinase gene in trout. I'll make the appropriate citation.
Odd Bres, Ph.D.
Department of Zoology
University of Manitoba
Winnipeg, Manitoba, CANADA R3T 2N2
obres@cc.umantitoba.ca
204 474 9245
Name: Gérard
Email: gniyonizigiye@hotmail.com
URL: http://
Remote Host: d51a5e345.kabel.telenet.be
Subject: Température
Bonjour Monsieur le Professeur et merci pour les informations que vous nous donnez à travers vos pages Web.
En effet, j'aimerais différentier deux séquences par la technique PCR basée sur l'ELISA en utilisant un oligonucléotide qui va reconnaître la séquence spécifique. Mais comme les deux séquences présentent une forte homologie, la sonde spécifique d'une longueur de 17 paires de bases avec seulement une différence de 2 bases a été choisie:
actggcgcagatggacc.
A quelle température d'hybridation dois-je faire l'incubation pour la différentier avec: actggcgcagatgtaca
Merci
Name: Johnny
Email: lokecj@muohio.edu
URL: http://
Remote Host: ip134-053-027-184.s27.muohio.edu
Subject: mFold
Dear Dr. Michael Zuker,
I am a grad student here at Miami University, Ohio studying Arabidopsis polyadenylation signals. I have came to you site to fold some RNA sequences, and I have about 1000 sequences, the problem with my summission using your quickfold is that it does not give jpg images output. Is there a way I can do a big summission with jpg image output?
Please advice. I appreciate for your help in this.
Thanks.
Johnny Loke
Name: chabane
Email: tibichec@iro.umontreal.ca
URL: http://
Remote Host: tweed.iro.umontreal.ca
Subject: mfod
Hye!
My comment is that when I enter a sequence where there is a mix of upper and lower case characters, the output is don't respect the case of the input sequence and give the result only in upper case characters.
Thanks
Name: Xu
Email: xuhy@genomics.org.cn
URL: http://
Remote Host: 211.152.157.234
Subject: puzzle
Dear Prof. Zuker,
Thank you very much for your replying last time. Now I am also puzzled by two questions: (1)why in /mfold-3.1.2/samples the energy value in mdv1.log is different from it in mdv1_*.gif, for example, the minimum energy (that is shown in the bottom of mdv1_1.gif)is dG=-143.45(initially -145.3),but in the mdv1.log, however, it(also mdv1_1) is dG=-139.86(initially -141.8)? In my opinion, they would have been the same value, because they are calculated for the same structure mdv1_1. (2) during installing mfold-3.1.2 under Red hat Linux 9.0 and 32bit CPU and 2.0G RAM, I found that if in Makefile£¬I set maxn(nafold)>22612 or maxn1(quickfold)>16383 or maxn2(nafold2)>16182, after ¡°make all¡±£¬the screen shown ¡°bigstuff is too large¡±£¬and installing failed. I wonder how the three different numbers are come from, and why they seem a bit strange? Thanks a lot!!
Name: Verónica
Email: mvr@agro.uba.ar
URL: http://
Remote Host: ol168-193.fibertel.com.ar
Subject: thanks!
I used your program to analize cDNA secondary structure of a sorghum gene and obtained a very complicated shape; I will use this information to design new primers for real time pcr, I hope this time things will work! Thanks to all the people who make this software available on the internet.
Kind regards,
Verónica
Name: mr
Email: aba_agu@yahoo.com
URL: www.aba
Remote Host: 62.56.189.138
Subject: good
i love this guest page.
Name: Fernando
Email: fgomez318@aol.com
URL: http://
Remote Host: b12-arbiter-d.net.nih.gov
Subject: Great
Dear Fellows;
I just wanted to drop you a note letting you know that I really enjoy using your RNAfolding service. I use the nucleic acid folding option when I want to see if my siRNA duplex components are going to form any energetically favorable secondary loops, and therefore should not be used for the siRNA KO I am designing. However, just for clarification, I wanted to confirm that the "dG" indicated in the thermodynamic profile you guys give when analyzing a particular sequence means "delta G". I ask this because other sites give delta E. Is "dG" delta G, and how should I view this value differently from the delta E value given in other sites?
Thanks for taking the time to answer my questions. I really appreciate it!
Sincerely,
Fernando Gomez
Summer Intern, National Institues of Aging
Name:
Email: saskia.rutjes@rivm.nl
URL: http://
Remote Host: proxy02.rivm.nl
Subject:
Name: Gabriela
Email: golmedo@ira.cinvestav.mx
URL: http://
Remote Host: ciea.ira.cinvestav.mx
Subject: suggestion
I wish there was a second button to start RNA folding right next to the box where the
sequence is paste. When testing many RNA segments with the same parameters I hate to
scroll down from the box to the clickable fold RNA botton.
Name: Dr
Email: bourdonv@mskcc.org
URL: http://
Remote Host: 140.163.188.72
Subject: mfold
Hi
I need to send a batch of short sequences to be fold and to get back the vienna file of each structure obtained. I can not find how to do, is it possible?
thanks
Name: Nikolai
Email: ns01@ic.ac.uk
URL: http://
Remote Host: ic.ac.uk
Subject: Folding
Dear Prof. Zuker,
We are currently working on a novel nucleic acid-based diagnostic assay involving the formation of DNA/RNA 3-way junctions. I was wondering if you are aware of any software capable of modelling such hybridisations between 3 or more single stranded nucleic acid sequences.
Many thanks,
Nikolai Szep
Name: Matthew
Email: mrounseville@htgenomics.com
URL: http://
Remote Host: htgenomics.com
Subject: Batch
Dear Dr Zucker,
I have been using your algorithms since the late 1980's when I was working on the structure of HIV TAR RNA. Thank you for your exceptional contributions to the scientific community.
I have been using your hybridization server to screen for unwanted hybridizations between oligos in our microarrays. The program is very useful, however, it is rather tedious because I need to enter each pair of sequences one set at a time. Is there any way to run this program in batch mode where I could evaluate potential cross-hybridizations between multiple pairs of oligos?
Regards,
Matt
Name: acox30
Email: acox30@hydromail.com
URL: http://
Remote Host: pcp06940631pcs.nrockv01.md.comcast.net
Subject: site
This is an awesome site
Name: Huajiang
Email: weihuajiang@tsinghua.org.cn
URL: http://
Remote Host: 211.151.91.194
Subject: how
My major work is probe deisgn and hybridization data analysis. Our probe is designed for specific purpose instead of gene expression and our probe and their target sequences are very short, about or more than 50 bp. Specificity is quite import to us and cross hybridization should be avoid! Althrough Sequence alignment and similiarity can be criterion to score specificity, I think hybridization energy is more reasonable.
2-state hybridization is part of mfold 3.1, but I can't find it and use it in mfold 3.1. could you tell me how to use 2-state hybirdization through mfold!
Thank you!
Best wishes
----------
Department of Biological Sciences & Biotechnology,
Tsinghua University,
Beijing 100084, P.R. China
Name: Changbong
Email: hyeoncb@glue.umd.edu
URL: http://
Remote Host: rnadream.umd.edu
Subject: Installing
Dear Prof. Zuker:
I installed mfold-3.1.2 under /usr/local/lib in my local linux machine.
I set the variable in Makefile as
BINDIR = /usr/local/bin
LIBDIR = /home/myname/MFOLD
and created directory called MFOLD in my home directory.
The installation seemed to be fine.
But when I run the mfold giving a sequence in FASTA format as
mfold SEQ='1GID.fasta'
I got the following results.
cat: /dat/begin.dat: No such file or directory
REUSE= NO
cp: `1GID.fasta' and `./1GID.fasta' are the same file
1GID.fasta.pnt created.
Sequence length is 159
Folding at 37 degrees using version 3.0 dat files.
Save file created using nafold2.
Minimum folding energy is -63.1 kcal/mole.
Energy increment is 3.155 kcal/mole.
H-num file created from plot file.
1,
Suboptimal foldings created.
Energy dot plot created.
cp: cannot stat `/dat/*.dat': No such file or directory
miscloop missing
One of the data files is missing
Free energies re-evaluated using efn2 and added to ct file.
cat: /dat/bases.nav: No such file or directory
Structure plots generated.
All done.
I'm concerned about the lines with
"cat: /dat/begin.dat : No such file or directory"
and
"cat: /dat/bases.nav: No such file or directory"
Is anything wrong with the modification in Makefile?
I tried another fasta file which looks like the following
>1F27:A RNA
ACCGUCAGAGGACACGGUU
but in this case mfold SEQ='1F27.fasta gives the output as follows.
cat: /dat/begin.dat: No such file or directory
REUSE= NO
cp: `1F27.fasta' and `./1F27.fasta' are the same file
1F27.fasta.pnt created.
Sequence length is 19
Folding at 37 degrees using version 3.0 dat files.
Save file created using nafold2.
Save file is empty. No foldings.
Job Aborted
I guess I made a simple mistake when I set up the path in Makefile.
I would appreciate you for the help in advance.
Changbong
Name: Mr.sarayoot
Email: sarayoot@dmsc.moph.go.th
URL: http://
Remote Host: 203.157.14.246
Subject: Thank
This is a second time for to use your software for folding analysis. The first time is to fold the RNA sequence. This software
are very useful for me including there are many analuzed information for further analysis.
Thank you very much of your kindly provinding me the help.
Mr. Sarayoot Subpasu
sarayoot@dmsc.moph.go.th
ssubpasu2004@yahoo.com
Name: Roland
Email: rgaboury@inuktun.com
URL: http://
Remote Host: s0106000d88f1df8b.no.shawcable.net
Subject: Cycling...
Hey... who's that big old guy you're standing next to on the bridge?
M. Zuker replies: Thanks for the flattery. The picture above is what I actually looked like in 1990. I see by your E-mail address that you're back in Candada. In fact, you're back to your home town. I'll visit if I ever get to BC. I have an open invitation to visit the CS Department at UBC. I'm glad you made contact.
Name: Wei
Email: wei.guo@viagen.com
URL: http://
Remote Host: yahoo.com
Subject: Automatic
Hi Professor Zuker,
My name is Wei Guo, a researcher at a small biotech start-up Viagen. I am wondering if I can use your DNA mfold server to test 2nd structures of a bunch of PCR primers. I want to write a script to make automatic requesting using url-encoded data, such as http://www.bioinfo.rpi.edu/applications/.../form1.cgi?sequence=actggcccaattttccaaggcttacc&email_addr=wei.guo@viagen.com&action=fold
Particularly I need to know what the key-value pair is returned when the Fold DNA button is clicked to submit a request.
Your time and help is greatly appreciated.
Sincerely,
Wei Guo
M. Zuker replies: Many users submit sequences or groups of sequences automatically to my various servers. Your message indicates that you do not understand how to do this properly. I will give you some information and suggestions.
"http://www.bioinfo.rpi.edu/applications/old/cgi-bin/nph-mfold-3.1.cgi"
and not to a cgi script that generates a form.
- "quikfold" server: Fold many RNA or DNA sequences in a single
run. Optimal and suboptimal foldings are predicted. No images.
- "zipfold" server: Predict the minimum folding energies for many
RNA or DNA sequences. No foldings are computed.
- "Tm server: Predict the minimum folding energies for
many RNA or DNA sequences together with the enthalpy and entropy of
minimum energy foldings and a melting temperature based on a simple
2-state model. No foldings are shown although they could easily be
made available.
You should read:M. Zuker
Mfold web server for nucleic acid folding and hybridization prediction.
Nucleic Acids Res. 31 (13), 3406-15, (2003)
[Abstract] [Full Text] [Supplementary Material] [Additional Information]
I apologize for my error in the Tm formula for 2-state hybridization. A correction appears under the "Additional Information" link.
Finally, the "quikfold" server needs a password (the NAME is "VERIFY") that changes daily and can only be found within the form generated by
"http://www.bioinfo.rpi.edu/applications/mfold/old/rna/form3.cgi".
This was added to discourage use of the server by commercial software packages.
Name: Sarah
Email: snahmias@alumni.princeton.edu
URL: http://
Remote Host: 146.203.53.105
Subject: scoring
When the energy is calculated, a number is given, but it says [initially....] what does that mean? Why has the number changed?
thanks
Sarah
M. Zuker replies: For RNA folding, versions 3.x of mfold use free energy rules as described in:
D.H. Mathews, J. Sabina, M. Zuker & D.H. Turner
Expanded Sequence Dependence of Thermodynamic Parameters Improves Prediction of RNA Secondary Structure
J. Mol. Biol. 288, 911-940 (1999)
The rules for multi-branch loops are too complicated to be used in an efficient algorithm. Therefore, the folding is done using slightly simpler rules. This gives the "initial free energies". The free energies are then re-evaluated using the best rules that we have. In particular, the revised energies are determined by:
- using Jacobson-Stockmeyer theory to assign free energies to
multi-branch loops that includes a term that grows logarithmically
with the number of unpaired bases in the loop,
- and by computing coaxial stacking of adjacent helices in multi-branch
loops or in the "exterior" loop. This latter feature has been
implemented by David Mathews in his RNAstructure and related software.
The re-evaluated energy is what is given as "dg = ..." in the figures. The only purpose of this re-evaluation is to re-order the foldings according to "better" energies. We reported that this improves folding predictions. That is, the optimal folding according to the revised energy rules tends to contain more "correct" base pairs.Name: sujun
Email: zheng003@sina.com
URL: http://
Remote Host: sina.com
Subject:
dear Michael Zuker
Thanks for your web service of RNA folding !
about 2 days ago,i have input twuo rna sequence for rna strcuture rediction .I appled for the batch processing form .but i have not receive the results.Can you help me?
your friend
sujun zheng
M. Zuker replies: According to the log files, you submitted six large folding jobs recently. The four jobs submitted at around 04:00 EST on November 15 have already been erased. The two jobs submitted at round 08:00 EST on November 17 are still on the system and may be found at: 08-17-20 and here. Both jobs finished in less than 15 minutes. You should receive E-mail from the server when your job is complete. However, when you submit a batch job, you are given the URL for the results. You should check this URL after a hour or so. Do not forget to refresh the page, if necessary.
Name: Kordai
Email: M.Sowa@ed.ac.uk
URL: http://
Remote Host: ed.ac.uk
Subject: RNAfold
Hi,
The server keeps telling me to do a batch job, (even though I have chosen batch job) and it
won't fold my 1499 bases. Could you please tell me what to do to rectify this? (it says I
can fold up to 6000 bases as a batch job, so I'm not over the limit). Thanks.
Kordai
Name: LY
Email: winecar@sohu.com
URL: http://
Remote Host: 220.163.54.191
Subject: The
I have used Mfold to abtain RNA secondary structures of genus Thermus species. Most of them were agreeable with the model published in 1993 made using 16S rRNA of E.coli. But when folding 16S rRNA of T.scotoductus, most of its strains lost their Helix11, which could be shown in windows version of Mfold -RNASTRUCTURE 4.1. How should I resolve such problem?
Great appreciation to you for your reply.
Thank you!
M. Zuker replies: About the worst thing to do with an RNA folding program that folds single sequences based on minimizing free energy is to expect it to predict a correct folding for a large molecule without any additional information. This is all the more true for small and large subunit rRNA, where ribosomal proteins probably serve as guides or chaperones in the folding process. mfold, RNASTRUCTURE and other such programs attempt to simulate equilibrium conditions in solutions in the absence of other molecules.
It is remarkable that mfold gives good answers for RNA viruses, in the sense that separate folding domains are often robustly predicted and many portions of predicted secondary structures are "well-determined" in the sense that there are few alternatives for those predictions. It is also remarkable that a minimum energy folding of, for example, Thermococcus celer 16S rRNA contains about 80% of the base pairs determined from phylogenetic methods. Predictions for rRNA of Eubacteria are not nearly as good. Predictions for Eukaryotes are much worse and those for mitochondrial rRNA are total nonsense. In joint publications with the Turner group, you should note that we have not attempted to fold entire rRNAs. For small subunit rRNA, the sequence is broken into known domains of size up to about 500 bases and these are folded separately.
If you wish to determine the secondary structure for, I presume, a new rRNA, or at least one whose structure is unpublished or otherwise not available in a public database, then by all means use mfold, but add constraints that force certain known interactions or that exclude incorrect foldings. Eukaryotes often have extra regions in their rRNA that are presented as unstructured. It is appropriate to use mfold to determine foldings for such inserted regions for which phylogenetic data are either absent or insufficient to determine secondary structure.
Finally, although RNASTRUCTURE (by David Mathews) began as a "translation into C++" of the RNA folding program (nafold) in the mfold package, it has been evolving more or less independently of my old code and the new software being developed by my student, Nick Markham, for folding and hybridization prediction. It is not realistic to expect identical predictions.
Name: Ming
Email: mwu@qdots.com
URL:
Remote Host: 68-123-139-98.ded.pacbell.net
Subject: Thermodynamic
Dear Prof. Zuker,
Would it be possible to show dH and dS for the RNA mfold? Currently only free energy dG is shown. dH and dS will help to determine Tm and equilibrium constant K at different temperatures. What about salt effect on dH and dS for RNA folding?
Thanks.
Ming
M. Zuker replies: I cannot offer dG and dS for RNA folding (or hybridization) using the latest, version 3 free energy rules at 37° C because they are not available. It is for this reason that I offer an alternative for RNA folding that uses the older, version 2.3 parameters, for which enthalpies are also available. This site, RNA mfold version 2.3 server, is linked from the default RNA folding form, which uses version 3 free energies. Use version 2.3 RNA folding only if you really want to predict Tm as well as dH and dS, or if you wish to investigate how folding changes with increasing temperature. However, if you attempt folding RNA from a thermophile by setting the temperature to 70° C or higher, you will get nonsense.
Salt concentrations affect dS only. I have corrections for DNA folding and hybridization which depend on the number of phosphate pairs in hybridized regions. These corrections are independent of GC content and the subject of some controversy. I have no such corrections for RNA. However, the RNA folding parameters, derived for [Na+] = 1 M and [Mg++] = 0 M, corresond to physiological conditions where [Na+] = 125±25 mM and [Mg++] = 10±5 m M.
In the future, I hope to have a more comprehensive set of RNA parameters that will permit melting predictions and corrections for salt concentrations. They may come from my colleagues and long term collaborators at the University of Rochester (Doug Turner's lab and David Mathews) or from a company such as DNA Software, which produces high quality commercial software based on theory and measurements that is elegantly packaged, user friendly, and oriented to the solution of specific problems.
Name: Du
Email: tonydudu@21cn.com
URL: http://
Remote Host: pku.edu.cn
Subject: about
It seems that the server is down. Would you please have a check on it?
In addition, is the local version of hybridization server available?Thank you!
Name: diederik
Email: wdsmilde@casema.nl
URL: http://
Remote Host: 535300be.cable.casema.nl
Subject: comments
Great stuff this is, but is it really necessary to type in my email address every time I submit another small job?
M. Zuker replies: I agree. I have removed the annoying demand for an E-mail address for DNA folding. The server will continue to demand an E-mail address for batch jobs. In these cases, a message is sent informing the user that the job is complete.
Name: Mr.Sarayoot
Email: sarayoot@dmsc.moph.go.th
URL: http://
Remote Host: 203.157.14.245
Subject: Thank
Dear Prof. Michael ZUKER
I have no comment, however, I thank you very much for your kingly providing me the tools for DNA folding analysis. This very useful for predicting the Genotyping Analysis via SSCP.
Your Sincerely
Mr.Sarayoot Subpasu
Enteric Bacteriology Laboratory
National Institute of Health
Department of Medical Sciences
Minstry of Public Health
Tivanont, Nonthaburi 11000, THAILAND.
Cell-Phone: 66-01-2641064
¤