Am. Chachulska et al., SYNTHESIS OF FULL-LENGTH POTYVIRUS CDNA COPIES SUITABLE FOR THE ANALYSIS OF GENOME POLYMORPHISM, Journal of virological methods, 67(2), 1997, pp. 189-197
Citations number
29
Categorie Soggetti
Virology,"Biochemical Research Methods","Biothechnology & Applied Migrobiology
New methods facilitating the synthesis and amplification of full-lengt
h cDNA copies of single-stranded viral RNA genomes have been developed
. A method is described for the efficient purification of potyviral RN
A and total RNA from infected plants and it is shown that they can ser
ve as templates for the efficient synthesis of a full-length, 10 kb lo
ng, genomic cDNA. Two different reverse transcriptases were used (AMV-
RT and MMLV-RT); only the first reverse transcriptase produced a good
quality, full-length cDNA using viral RNA as a template. Surprisingly,
MMLV-RT allowed for the full-length cDNA synthesis on virions rather
than viral RNA. The PW cDNA, synthesized using either RNA or virions,
can be amplified successfully by PCR with high yields of full-length p
roducts. Such products are good substrates for the study by RFLP of th
e total genome polymorphism of virus isolates. (C) 1997 Elsevier Scien
ce B.V.