A method for rapid isolation of total RNA of high purity and yield from Arthrospira platensis.
Arthrospira (Spirulina) platensis is widely used as a food supplement and has been an economically important species for centuries. However, the genetic aspect of studies of this particular organism has always been neglected, mainly because of the nonavailability of suitable methods for isolation of nucleic acids and the difficulties faced during further manipulations. Although total RNA has been isolated using commercially available kits, we present a method optimized to obtain DNA-free total RNA of higher yields and higher purity in less time than is required by other methods (<2 h). It involves hot phenol - chloroform - IAA extraction using an aqueous to organic phase ratio of 1:2 followed by lithium chloride precipitation and 70% ethanol wash. This method, optimized for the cyanobacterium Arthrospira (Spirulina) platensis, eliminates the need for DNase treatment and produces high-quality RNA, as validated by bioanalyzer, RT-PCR, and cloning. With the recent release of the Arthrospira genome, the current method will be of great value for carrying out high-throughput studies like microarray and real-time PCR.
Pathak
Ravi Ramesh
RR
G.G.S. Indraprastha University, Delhi, India.
Lochab
Sunila
S
D000458:Cyanobacteria / Q000235:genetics*; D005821:Genetic Techniques; D016133:Polymerase Chain Reaction; D012329:RNA, Bacterial / Q000235:genetics / Q000302:isolation & purification*
D012329:RNA, Bacterial