Development of a highly sensitive nested RT-PCR method for Beet necrotic yellow vein virus detection

Development of a highly sensitive one-tube nested real-time PCR for detecting Mycobacterium tuberculosis

“…One-tube nested real-time PCR was approximately 100 times more sensitive than conventional real-time PCR and AdvanSure TB/NTM real-time PCR. These results were consistent with reports indicating that nested PCR greatly enhanced sensitivity compared to conventional PCR (Abrahao et al, 2009;Llop et al, 2000;Morris et al, 2001) due to 2 sequential amplification steps of the target gene. The detection limit of one-tube nested real-time PCR was 1 fg/μL of M. tuberculosis DNA, which is supposed to be the lowest quantity of nucleic acid capable of detection by PCR (Eing et al, 1998;Hashimoto et al, 1996).…”

“…Over the past several decades, nucleic acid amplification (NAA) tests, especially polymerase chain reaction (PCR), have been implemented for the detection of pathogens (Armand et al, 2011;Bosshard et al, 2004;Llop et al, 2000;Morris et al, 2001). PCR has been utilized widely for the detection of M. tuberculosis due to its rapidity and specificity (Chan et al, 1996;Cousins et al, 1992;Hashimoto et al, 1996;Kim et al, 2008;Sankar et al, 2011).…”

Construction of Plant Transformation Vectors Carrying Beet Necrotic Yellow Vein Virus Coat Protein Gene (I) - Transformation Vectors

“…Isolation of BNYVV coat protein gene was easily achieved, since all samples gave PCR products after RT-PCR with total RNAs as well as with IC RT-PCR. These results are not unexpected because in previous research isolation of BNYVV CP gene from sugar beet tissue was possible only by twostep PCR (12), or RT-PCR with "nested" primers (20). Even when BNYVV RNA fragments were multiplied with single PCR their amount was so low that they could be analyzed only with polyacrilamide gels and stained with silver (14), which is more sensitive than ethidium bromide.…”

Multiplex Reverse Transcription-PCR for Simultaneous Detection of Beet Necrotic Yellow Vein Virus , Beet Soilborne Virus , and Beet Virus Q and Their Vector Polymyxa betae KESKIN on Sugar Beet

“…The technique is a new tool for sugar beet breeders wanting to test their soils for the presence of different soilborne viruses. Though probably not as sensitive as a nested-PCR method for BNYVV detection (26), the new technique gives results that demonstrate that the sensitivity is much higher than that observed for the BNYVV DAS-ELISA and highly similar to that of RT-PCR. No RT-PCR technique has been described for the detection of BSBV and BVQ.…”