In general ,we order deletion strain from CGC or any other place.
But can someone tell me how the deletion strains are created in those Genome centers? I heard scientists use mutagens like EMS, UV / TMP etc…
But specifically how we get a strain deleted just specifically for a particular gene?
Any inputs will be highly appreciated…
Our lab has established the following protocol to isolate deletion mutations in specific genes:
http://www.wormbook.org/chapters/www_introreversegenetics/introreversegenetics.html#d0e928
I believe that the protocols used by the KO consortia are similar. It’s important to note that the method does not introduce mutations into your favorite gene, only that you can use PCR screening protocols to isolate from the library those specific strains that bear mutations.
Additional modifications/improvements have been made by other labs including the use of heat-stable restriction enzymes to reduce amplification of the wild type gene (http://www.ncbi.nlm.nih.gov/pubmed/12384612). A different method developed by the Jorgensen lab called MosDEL (http://www.ncbi.nlm.nih.gov/pubmed/20418868) should allow one to engineer deletion mutants starting from a strain bearing a Mos1 transposon insertion in or near your gene of interest.
-Kevin.
Thank you so much… Really appreciate it…