DNA sequencing

[coffeebythelake]

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Not MCAT related, but thought I would get more responses here, esp. from people who took labs in bio.

I was wondering why DNA sequencing techniques have trouble detecting long deletions and duplications of the DNA.

 

[Foghorn]

Not MCAT related, but thought I would get more responses here, esp. from people who took labs in bio.

I was wondering why DNA sequencing techniques have trouble detecting long deletions and duplications of the DNA.

Sequencing of DNA containing deletions isn't particularly useful unless wild type DNA is initially known. It's more logical to 1st do restriction analyis of wild type DNA and a sample containing deleted DNA to qualitatively and quantitatively compare differences between the two prior to sequencing. The same reasoning can be applied to duplications. For eukaryotes, whether (wild-type) genomic DNA or cDNA is being analyzed also needs to be considered among other things.

What do you mean by long deletions? Do you mean long strectches of DNA that needs to be sequenced or a DNA sample known to have a large segment of DNA missing? If it's sequencing long segments of DNA, there are limits to how long DNA Polymerases can extend under non-extreme polymerization conditions. There are techniques such as long distance PCR, developed by Wayne Barnes in a 1991 PNAS paper, where 50Kbps can be synthesized. This isn't normally used for generic Sanger dideoxy sequencing method with fluorescent nucleotides.