These pages provide a "minitutorial" for CODDLe'

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Updated March2007
These pages provide a "mini-tutorial" for
CODDLe. All TILLING requests must be initiated
through CODDLe. Why? CODDLe performs several
types of analyses on your gene target and stores
that information in our database. Inside, are
helpful hints for using this program, including
updates based on what we have learned over the
past few years.
CODDLe was developed by the PROWEB group in
Seattle, www.proweb.org
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CODDLe's pre-processing utility, the sequence
input page
Each gene target in our database must have a
unique name. You may be asked to use a different
name when you actually submit your request.
Your gDNA sequence must correspond directly to
the cDNA and/or protein sequence. A one base
difference, such as one due to a SNP, will give
you an error message.
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CODDLe does a BLAST search of known proteins to
find conserved regions, or BLOCKS
If an amino acid is conserved in related
proteins, it probably is important for function.
If you are working with a "hypothetical" or
uncharacterized protein, please submit your own
alignment to BLOCKS (see FAQ on our website).
This will allow the bioinformatic programs to
assess the TILLING mutations.
Proceed with CODDLe
PARSESNP Project Aligned Related Sequences and
Evaluate SNPs. This is the output you will get
when we notify you that we have found mutations
in your gene target!
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CODDLe - Step 3
You can change some of the parameters on this
page.
If you are not satisfied with the BLOCKS output
and if you have a better protein alignment for
your gene, then you can enter it at this point.
See the FAQ link on our website for more details.

We can TILL targets from 750 - 1500 bp. If you
have a large intron, it may be better to select a
smaller target.
Be sure the mutation method "TILLING w/ EMS
(plants fly)"
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Selecting the region to TILL, some things to
consider
Conserved regions from BLOCKS
Mutations in these regions are likely to be
damaging
Best Region to TILL
Mutations in these regions not likely to be
damaging
SIFT Sorting Intolerant From Tolerant PSSM
Position Specific Scoring Matrix
Both these algorithms weight the identity and
frequency of the amino acids found at each
position in the protein.
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CODDLe will let you TILL different regions of
your gene target
Be sure to look at the spectrum of mutations
Try to minimize the amount of intron in your
target. If necessary, TILL a region lt 1.5 kb.
Use this section to CODDLe different regions of
your gene. Be sure the "REDO" button is
highlighted.
CODDLe will determine the amino acid change for
every G and C in your sequence
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Primers for the region you want to TILL are
automatically designed by a modified version of
Primer3
Primer3/CODDLe will let you select the of
primer pairs to return
Fields marked with a red dot are the default
parameters for TILLING PCR conditions. Please do
not change them.
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Save your outputs as a .html file for future
reference, or to place your order at a later date
Try to TILL a region where 50 of the mutations
are non-silent.
If lt 25 of all possible mutations are
non-silent you will get a warning message. Most
of the mutations found by TILLING will be silent
or in introns, and it will take us longer to
complete your order!
Ready to order? Click here and you will be
directed to a "billing" page.
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See next page
This spring (2007) the TILLING seed stocks will
be deposited at the maize Co-Op.
To order seed, click on the link or email
maizetilling_at_purdue.edu
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A PARSESNP report is sent to you when we find
mutations in your gene target.
A list of restriction sites gained and lost in
the mutant lines.
Mutations with a SIFT score lt 0.05 are predicted
to be damaging based on bioinformatics.
However, any missense mutation may prove to be
interesting!
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Your TILLING request is considered complete when
we have obtained an allelic series where there is
95 probability (based on the bioinformatics)
that at least one of the mutations will be
damaging to the protein.
Go to the Seattle TILLING Project's website for a
more detailed explanation. http//tilling.fhcrc.o
rg9366/files/user_fees.html
For more information on CODDLe, BLOCKS, SIFT,
PARSESNP please check out these links
http//www.proweb.org/coddle/coddle_help.html
http//blocks.fhcrc.org/
http//blocks.fhcrc.org/sift/SIFT_help.html
http//www.proweb.org/parsesnp/parsesnp_help.html
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