Reverse transcription loop-mediated isothermal amplification of DNA for detection of Barley yellow dwarf viruses in China

Abstract

A reverse transcription loop-mediated isothermal amplification of DNA (RT-LAMP) for detection of Barley yellow dwarf viruses (BYDVs) was developed. In this procedure, three sets of four primers matching a total of six sequences of the coat protein or read-through protein genes of BYDVs - one each for three species, namely BYDV-GAV, -GPV and -PAV were synthesized for developing a specific and sensitive RT-LAMP assay for total RNA extracts from field-infected wheat plants in such a way that a loop could be formed and elongated during DNA amplification. RT-LAMP assays for each of three species of BYDV/CYDVs in China exhibited high specificity and could detect viral sequences in total RNA extracts from infected oat tissues at dilutions of 1 x 10(-5). All field samples collected from different regions of China showed the same result using both RT-LAMP and RT-PCR. This relatively simple and sensitive technique showed excellent potential with field-collected samples and for use in grassroots agencies in developing countries with limited resources.