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Method Detail
Submitter Method Handle: TSC-CSHL
Submitter Method ID: TSC-SANGER-19-24
Submitted method description:
The following method definition consists of one or more
protocols used in The SNP Consortium project by the
participating laboratory SANGER.
See the TSC website (http://snp.cshl.org/cgi-bin/protocol)
for a complete list of TSC protocols.
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SNP production protocol TSCM0019 (
http://snp.cshl.org/cgi-bin/protocol?id=TSCM0019)
Title: Library preparation and sequencing for PvuII enzyme
Lab: Sanger Center, Cambridge
Contact: snp@sanger.ac.uk
Description:
A panel of 24 DNAs was digested with PvuII, size
fractionated on an agarose gel, and cloned into puc18-based vectors.
Size fractions were taken at 1.0-1.1kb, 1.1-1.2kb, 1.2-1.4kb, 1.4-1.6kb,
1.6-1.8kb, 1.8-2.1kb, 2.1-2.5kb, 2.5-3.0kb, 3.0-3.5kb and 3.5-4.0kb,
from which libraries were constructed. Samples from these libraries were
named such that the size insert can be identified. The sample names
start with p1_0, p1_1, p1_3, p1_5, p1_7, p1_9, p2_3, p2_7, p3_2 and
p3_7, with respect to the size fractions listed above. Sequences were
obtained primarily from ABI 3700 capillilary sequencers. Base-calling
was performed with Phrap, which provides the quality values upon which
the SNP detection is based.
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SNP detection protocol TSCM0024 (
http://snp.cshl.org/cgi-bin/protocol?id=TSCM0024)
Title: SNPs from reads detected against finished public human genomic sequence
Lab: Sanger Center, Cambridge
Contact: snp@sanger.ac.uk
Description:
All reads were clipped of sequencing vector and low quality ends,
which set a usable read length for each read. The clipped reads
were screened for repetitive sequence with RepeatMasker, using
the default human settings. Only reads with >=80 non-repetitive
bases and >= 100 Phred quality (Q) >=30 bases were used in this
analysis. The reads were cross-matched against available finished
public human genomic sequence which is available via anonymous
ftp at snp.cshl.org in the file pub/SNP/nrClean21-FEB-00. All
reads which aligned to genomic sequence within 40bp of the start
and 10bp of the end of their usable read length, were assembled
with the matching region of genomic sequence (Q was set to 40 for
finished genomic sequence). High quality base discrepancies
(Q>=23) were identified as candidate SNPs. Further restrictions
on the candidate SNPs were that its neighbouring 5 bases all had
Phred quality values of >=15, at least 9 of the 10 neighbours
match and that the trace chromatograms at the SNP base location
did not have an underlying base signal greater than 50% of the
called base. If the number of detected SNPs in one clique was
greater than 4 or the depth of the assembly (not including the
genomic sequence) was greater than 8, then all SNPs were discard-
ed for that clique.

This method was used in the following submission:

Submitter Handle Batch Type Submitter batch id Release build id
TSC-CSHL Assay TSC-SANGER-19-24-000905-RESUBMISSION 85
TSC-CSHL Assay 001018_11 89
TSC-CSHL Assay 001219_12 92
TSC-CSHL Assay TSC-SANGER-19-24_101012214 92
TSC-CSHL Assay TSC-SANGER-19-24_101060714 96
TSC-CSHL Assay TSC-SANGER-Sep-19-2002-SNP_SUBMISSION-19-24 107