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CODDLe
(Codon Optimized to Discover Deleterious Lesions)
is a web-based interface developed by the Proweb
group that performs several types of analyses on your
gene target to determine the region with the highest
number of potential damaging mutations. We have a "CODDLe
Tutorial" that will explain the different steps
of CODDLe and highlights the parameters that you can
change. This tutorial can be downloaded as either a
PowerPoint or PDF file.
CODDLe uses the BLOCKS
program to compare your protein to the InterPro
12.0 protein database and identifies conserved domains.
CODDLe will also determine all the possible EMS-induced
mutations for your target and then analyze those mutations
with SIFT
(Sorting Intolerant From Tolerant) to identify which
amino acid changes would not be tolerated at any given
position. Using the above information, the region(s)
of your gene target with the highest number of potentially
damaging mutations will be identified and suggested
for TILLING. You have the option of selecting primers
for this region or you can select another region of
your gene. Primers are designed using Primer3
that has been modified for our TILLING PCR conditions.
The default is set to return five primer pairs, but
you can increase this number. CODDLe outputs can be
saved as .html files until you are ready to place your
order.
We highly recommend that
you select 3-4 primer sets suggested by CODDLe/Primer3
and test them in your lab on both B73 and W22 genomic
DNA using our PCR
conditions to ensure that you get a product of the
expected size. It is also a good idea to sequence the
PCR product. We have noted the common reasons why primers
fail our pre-screening step and those are listed here.
If you have questions about the primers, you can email
maizetilling
for advice. Once you have identified a good primers
set for TILLING go ahead and submit your request.
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