Identification of Soybean mosaic virus Strains by RT-PCR/RFLP Analysis of Cylindrical Inclusion Coding Region

Abstract

A reverse-transcriptase polymerase chain reaction/restriction fragment length polymorphism (RT-PCR/RFLP) was employed successfully for detection and identification of Soybean mosaic virus (SMV) strains. A primer pair amplifying a 1,385-bp fragment of the cylindrical inclusion (CI) coding region was used for RT-PCR and the RFLP profiles of the RT-PCR products were compared after restriction digestion with RsaI, EcoRI, or AccI restriction endonucleases. These enzymes were chosen based on the nucleotide sequences of SMV strains G2, G5, G5H, G7, and G7H in the CI coding region. These five strains, as well as seedborne SMV isolates from local soybean cultivars, could be differentiated by RT-PCR/RFLP analysis. The results correlated well with strain identification by symptom phenotypes produced on differential cultivars inoculated with strains and isolates. The sensitivity of RT-PCR enabled detection of SMV from plants with necrotic symptoms in which the number of virus particles was too low to be detected by enzyme-linked immunosorbent assay.

Structure, expression, and mapping of two nodule-specific genes identified by mining public soybean EST databases

“…Ten microliters of PCR products was separated via electrophoresis on agarose gels with ethidium bromide. For the expression and alternative splicing analyses of AI973769 and AI973715, one-step RT-PCR protocol was used as described by Kim et al (2004). The primers used for the alternative splicing analyses were designed from the included putative intron and its flanking exon (Fig.…”

Biological, Molecular and Phiysiological Characterization of Four Soybean mosaic virus Isolates Present in Argentine Soybean Crops

“…Fragments corresponding to the CI and P1 genomic regions were amplified by RT-PCR, using the sets of primers described by Kim et al (2004) and Sherepitko et al (2011) (Table 2). RT-PCR was performed with the Access RT-PCR System (Promega Corporation Madison WI USA), using as template 1 μg of RNA of the different isolates.…”

SELEKSI GALUR DARI POPULASI F4 KEDELAI YANG TAHAN TERHADAP PENYAKIT MOSAIK (Soybean mosaic virus) DAN BERDAYA HASIL TINGGI

“…Masalah utama yang dihadapi adalah dari 191 aksesi kedelai yang telah diuji hanya diperoleh tujuh asesi kedelai (Lokal Jombang, Mlg 3288, Lokal Temanggung, Malabar, Pangrango, PI 200485, M8Grb 44) dengan kategori sangat tahan SMV secara visual dan bereaksi negatif dengan reaksi RT-PCR serta mempunyai penampakan agronomis yang baik. Sepasang universal primer mengamplifikasikan fragmen DNA berukuran sekitar 1687 bp pada posisi 4176 sampai 5560 (Kim et al, 2004;Somowiyarjo, 2004;Hwang et al, 2011;Andayanie & Adinurani, 2013a, 2013b. Sepasang oligonukleotida yaitu universal primer (Sprimer (F):5'GGNAAYAAYAG YGGNCARCC3' dan (M4) (R): 5'GTTTTCCCA GTCACGAC3' dirancang untuk mengamplifikasikan fragmen DNA berukuran sekitar 1687 bp (Kim et al, 2004;Somowiyarjo, 2004;Hwang et al, 2011;Andayanie & Adinurani, 2013a) Sepasang primer tersebut dirancang untuk gen CP dan Non Coding Region ujung 3 anggota famili Potyviridae (Chen et al, 2001).…”