metmap

DNA methyltransferase binding motif plasmid assembler

Overall purpose:

• To identify the motifs of multiple DNA methyltransferases (DNA MTase) simultaneously.

Quick start

• This script requires python 3.6 or newer.
• Install using "pip install metmap"
• This will put a script named "run_metmap.py" in your pythons bin folder.
  • Linux: You should just be able to type "run_metmap.py" from terminal
  • Windows: The script will be placed in your python folders "Scripts" subfolder and you can run it with "python path\to\python\Scripts\run_metmap.py" from command line
  • Mac: Who knows..probably works like linux?

Test

You can download the tests/test_data.txt and run
run_metmap.py test_data.txt
This should generate a cas1.fa and cas1.gb file.

Overview:

• You have an organism with multiple identified DNA MTases and you wish to know their individual motifs.
• You use NGS to obtain motifs of all methylated DNA sites.
  • Some of those motifs will contain ambiguous bases and some will not
• You submit those motifs to this program
• This program then stitches these motifs together in random order
• Motifs may contain ambiguous bases per the IUPAC nucleotide code
  • We can't synthesize lots of ambiguous bases, so we "de-ambigulate" them before putting them into the final construct.
  • De-ambigulation happens according to 1 of 2 rules:
    • Rule 1:
      • Pick L random variants of the motif. E.g. Motif ATGNNTTA have a total of 16 possible actual sequences. If L<16 then the program will random pick L variants (without duplicates). If L>16 then each possible variant will be picked at least L/16 times and some will be picked 1 more than that.
    • Rule 2:
      • Make K copies of each completely "de-ambigulated" variant: E.g. the sequence "SATC" will then be treated as 2 sequences: "GATC" and "CTAC" that will each appear in K copies.
• We put M N's between each motif
• And the program will output P versions of these cassettes
• You then clone this cassette into a plasmid with 1 DNA MTase in each plasmid.
• You then transform this library into an organism that doesnt natively methylate DNA.
• Grow, Harvest, Sequence plasmids.
• ?
• Profit!

Motif file format

• The motifs should be stored in a standard text file
• One motif per line, then a comma then a 1 or a 2 to indicate whether either rule 1 or 2 should be used for this motif Example:
  ATGCATGCATGC, 1
  STGCAGTCATCGTTK, 1
  ATCNNNNAAA, 2
  CGTAGCANNNATCGATGC, 2

IUPAC nucleotide code:

code	nucs
R	A or G
Y	C or T
S	G or C
W	A or T
K	G or T
M	A or C
B	C or G or T
D	A or G or T
H	A or C or T
V	A or C or G
N	any base