Isolation of poly (A)+ mRNA for downstream reactions from some medicinal and aromatic plants.
In the present protocol for extraction of RNA, hexadecyltrimethylammoniumbromide (CTAB) and insoluble polyvinylpyrrolidone were used followed by LiCl precipitation, CsCl ultracentrifugation and finally poly (A)+ mRNA was isolated with the help of oligo(dT)-cellulose columns. The isolated poly (A)+ mRNA was found to be suitable for cDNA-AFLP and suppression subtractive hybridization applications. It is a modified and consolidated protocol based on previously described methods for isolated steps and works better for medicinal and aromatic plants. High yield of poly (A)+ mRNA coupled with its amenability for downstream reactions like RT-PCR, northern blotting and cDNA synthesis for library construction is a key feature of the present protocol.
Shukla
Ashutosh K
AK
Genetic Resources and Biotechnology Division, Central Institute of Medicinal and Aromatic Plants, Lucknow 226 015, India.
Shasany
Ajit K
AK
Khanuja
Suman P S
SP
D030008:Catharanthus / Q000737:chemistry; D010936:Plant Extracts / Q000737:chemistry; D010946:Plants, Medicinal / Q000737:chemistry*; D012333:RNA, Messenger / Q000302:isolation & purification*; D018749:RNA, Plant / Q000302:isolation & purification*; D012015:Reference Standards
D010936:Plant Extracts; D012333:RNA, Messenger; D018749:RNA, Plant