Potential SNPs can be studied further, either by using a contig sequence or by using a sequence from a high quality section of a single read.
Programs such as phred and phrap and data management systems such as Geospiza's Finch -Server are used to:
- store sequencing data
- identify bases
- assemble sequences into contigs
- and to provide quality information about DNA sequences and assemblies
Phred processes data from sequencing instruments, identifes each base, and assigns a quality score to each base. The quality score is related to the probability that a base has been identified incorrectly. Poor quality DNA and/or poor quality run conditions make it more difficult to identify bases accurately, thus Phred scores are commonly used as a measure of overall quality in DNA sequencing.
Phrap is most commonly used to assemble shorter sequences into longer contiguous sequences (contigs).
More about Phred and PhrapIn this example, the sequences come from clone libraries of ESTs (expressed sequence tags) from CGAP (the National Cancer Institute's Cancer Genome Anatomy Project). EST libraries are made by isolating RNA, purifying the messenger RNA, and using reverse transcriptase to make a DNA copy.
Messenger RNAs are the RNA molecules that code for protein. EST libraries may also contain contaminating sequences from mitochondria or ribosomal RNA depending on the method used to prepare the library.
Each individual sequence (or "read") should represent an RNA molecule that was copied, cloned, and used as a template for sequencing.
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Funding for this project was provided by the National Science Foundation's Course, Curriculum, and Laboratory Improvement Program under grants DUE-0088153 and DUE-0127599
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