Preparing long probes by an asymmetric polymerase chain reaction-based approach for multiplex ligation-dependent probe amplification.

摘要: Abstract To clearly discriminate the results of simultaneous screening and quantification up to 40 different targets–DNA sequences, long probes from 100 500 nt, rather than smaller or similar-sized synthetic ones, were adopted for multiplex ligation-dependent probe amplification (MLPA). prepare probes, asymmetric polymerase chain reaction (PCR) was employed introduce non-complementary stuffers in between two parts MLPA with specially designed primers, then restriction enzymes selected digest double-stranded DNAs, finally polyacrylamide gel electrophoresis used purify single-stranded DNAs (i.e., probes). By using this approach, 12 prepared identify genetically modified (GM) maize. Our experimental show that full accordance ones could be assembled 4-, 7-, 10-plex analysis without losing result specificity accuracy, showing they as effective reliable those M13-derived vectors. This novel PCR-based approach does not need expensive equipment, special reagents, complicated operations when compared previous methods. Therefore, our new make more independent, efficient, economical.