New technique for rapid mutation detection

December 23, 2008

The gold standard for mutation screening for many years has been

amplification of target sequences by polymerase chain reaction (PCR)

followed by Sanger sequencing. However, when it comes to conditions

where the mutation may be in one of several genes or polygenic

conditions where a number of different genes may be involved (e.g.

breast cancer), gathering results by this technique can often be

labour intensive and time consuming as numerous different genes have

to be sequenced. Although a number of parallel sequencing techniques

have been developed, along with " front-end " methods to amplify a

subset of genes (see previous news), these procedures are still too

expensive and complex to be applied in a routine diagnostic or

screening setting.

Recently, VIB researchers at the University of Antwerp have developed

a new method of detecting mutations and copy number variations that

may make diagnosis of inherited diseases quicker (see press release).

The technique involves a combination of multiplex PCR (amplification

of multiple targets by using more than one set of primers) with high-

throughput parallel sequencing, in order to analyse multiple genes

simultaneously. In a paper published this month, Goossens et al.

describe the use of this technique to identify mutations in seven

genes associated with Charcot-Marie Tooth Disease (CMT) – an

inherited disorder that affects the peripheral nervous system

[Goossens et al 2008 Hum Mutat. 0(1-6)]. The researchers amplified

and sequenced seven genes from 31 individuals and compared the

mutations detected with the results obtained by traditional PCR-based

Sanger sequencing. All the variants that were detected by traditional

methods were also identified using the new technique. They were also

able to determine the copy number variation (CNV) of two genes

associated with CMT (GJB1 and PMP22).

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