Cycling gradient capillary electrophoresis: a low-cost tool for high-throughput analysis of genetic variations.
In the present work, we introduce a new type of DNA variation detection. This method represents a transfer of melting gel technique onto multicapillary electrophoresis DNA sequencing instrument with further improvements to achieve maximum sample throughput while maintaining a high performance. The main improvement comes from application of cycling (revolving) temporal temperature gradient in place of a single-sweep gradient, commonly used in similar gel-based techniques. This improvement enables utilization of multiple-injection technique, in which multiple samples are injected into the same capillary (or sets of capillaries) separated by predefined time intervals of partial electrophoresis. The periodic oscillation of the temperature results in identical separation conditions of all samples injected in such series. Using this novel approach, we demonstrate a dramatic increase in separation throughput by turning a standard commercial 96-capillary array instrument into a semicontinuous flow mutation detection system capable to screen over 15 000 samples in 24 h of operation on a single 96-capillary commercial instrument. This represents a 10-fold increase in sample throughput over the current comparable technology.
Minarik
Marek
M
Genomac International, Prague, Czech Republic. mminarik@genomac.com
Minarikova
Lucie
L
Bjørheim
Jens
J
Ekstrøm
Per Olaf
PO
D004252:DNA Mutational Analysis / Q000295:instrumentation / Q000379:methods*; D019075:Electrophoresis, Capillary / Q000295:instrumentation / Q000379:methods*; D014644:Genetic Variation / Q000235:genetics*; D011108:Polymers; D020641:Polymorphism, Single Nucleotide; D017422:Sequence Analysis, DNA / Q000295:instrumentation / Q000379:methods; D013696:Temperature; D013997:Time Factors
D011108:Polymers