Hi Guys,
I would like to find a tool that simulates the 'in silico' digestion of an eukaryote genome (not fully sequenced but EST/454 available) by the most frequently used restriction enzymes. My goal is to know, how many fragments and repetitive elements are generated, what are their average sizes, etc... depending of the enzyme. This will help me to know which enzyme is the best one to prepare gDNA libraries for NGS.
Cheers.
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written
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Bioch'Ti •
Brute force: the following java program prints all the restriction sites (type II) (chromosome, start, end, size) for all the chromosomes of the human chromosomes. Hope it helps:
Compilation:
javac Digest.java
Execution:
java Digest
Output:
chr1 AatII 0 47476 47476
chr1 AatII 47476 70600 23124
chr1 AatII 70600 164453 93853
chr1 AatII 164453 170555 6102
chr1 AatII 170555 175979 5424
chr1 AatII 175979 325093 149114
chr1 AatII 325093 360629 35536
chr1 AatII 360629 369596 8967
chr1 AatII 369596 568607 199011
chr1 AatII 568607 568665 58
chr1 AatII 568665 620090 51425
(...)
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Pierre Lindenbaum
As you said, Brute Force! Thnaks very much for you detailed and helpful answer!
This java program provided all the results I needed with less hassle than many of the web-based tools I had tried. Thanks!
This java script for in silico restriction digestion does not do reverse complementary search, am I right ?
https://en.wikipedia.org/wiki/Palindromic_sequence#Restriction_enzyme_sites
EMBOSS has an app that does this from REBASE database: http://emboss.open-bio.org/wiki/Appdoc:Restrict
In fact, EMBOSS has many tools to play with REBASE or restriction enzymes in nucleotide or amino acid spaces.
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Louis Letourneau
I just tried this Emboss package, but there are two limitations: the amount of sequences that you can upload (it has crashed with a 5Mb fasta file) and it didn't give me the length of the generated fragments by the list of the enzymes... However, it worked.
If you have Eukariotic sized genome (say 100Mb+) you will have to install EMBOSS on a local computer and use command line. On Ubuntu and Debian you can install it by typing: sudo apt-get install emboss
From an already sequenced organism?
You might want to look at : http://insilico.ehu.es/digest/ (ok that's prokaryotes!)
For eukaryotes: http://insilico.ehu.es/eukaryota/digest/
They also have a tool for long DNA sequence length digests here: http://insilico.ehu.es/restriction/long_seq/
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Thanks for the clarification on the sequencing status of the organism!
I also want to know how to save the output in a file, would you mind adding a few lines. Thanks!