A method for isolating functional RNA from callus of Dendrobium candidum contented rich polysaccharides.
Isolation of high-quality RNA from Dendrobium candidum is particularly difficult. D. candidum contains considerable amounts of polysaccharides that coprecipitate with RNA, which render RNA unsuitable for either cDNA synthesis and/or PCR amplification. In this paper, a rapid and efficient method was described for functional RNA isolation from the callus of D. candidum. The procedure included: (i) an extraction with phenol and isopropyl alcohol, to remove proteins and polyphenols; (ii) purifications by lithium chloride, pre-cooled (-20 degrees C) ethanol successively to remove polysaccharides. The method resulted in high-quality RNA suitable for DDRT-PCR and cDNA library analysis finally.
Wanqian
Liu
L
Key Laboratory for Biomechanics and Tissue Engineering under the State Ministry of Education, College of Bioengineering, Chongqing University, Chongqing 400044, China.
Bochu
Wang
W
Chuanren
Duan
D
Biao
Li
L
D018076:DNA, Complementary / Q000096:biosynthesis / Q000235:genetics; D031667:Dendrobium / Q000737:chemistry* / Q000235:genetics; D004587:Electrophoresis, Agar Gel; D005419:Flavonoids / Q000737:chemistry; D018021:Lithium Chloride / Q000737:chemistry; D010636:Phenols / Q000737:chemistry; D010940:Plant Proteins / Q000737:chemistry; D059808:Polyphenols; D011134:Polysaccharides / Q000737:chemistry*; D018749:RNA, Plant / Q000235:genetics / Q000302:isolation & purification*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012996:Solutions; D012997:Solvents; D013056:Spectrophotometry, Ultraviolet
D018076:DNA, Complementary; D005419:Flavonoids; D010636:Phenols; D010940:Plant Proteins; D059808:Polyphenols; D011134:Polysaccharides; D018749:RNA, Plant; D012996:Solutions; D012997:Solvents; D018021:Lithium Chloride