DNA sequencing (Forensic Science)
In some instances, biological samples are not suitable for performing the relatively fast and standard nuclear analysis and typing of DNA (deoxyribonucleic acid). On occasion, the copy number of nucleated cells is too low, and the analyst cannot obtain enough nuclear DNA to acquire a strong enough signal to interpret after typing the DNA. DNA sequencing, although more time-consuming and complicated than the standard analysis, may provide the necessary clues when other methods to obtain a good profile using conventional typing methods are exhausted.
The first step in sequencing a DNA sample is to determine what fraction of the genome is to be read (usually between three hundred and one thousand base pairs) and perform an amplification reaction using unlabeled primers that target the region of choice. After the DNA is amplified, the polymerase chain reaction (PCR) product is cleaned to remove any unbound nucleotides and residual, nonincorporated primers. This purified product is subsequently quantified and diluted to a predetermined concentration that is dependent on the length of the fragment of DNA being sequenced. The sequencing template is amplified with a single primer (forward or reverse) so that the end products are single-stranded DNA molecules. Unlike in the more common PCR reactions, a pair of primers cannot be used because the end product would result in a double-stranded molecule that could not be interpreted. This would be...
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