IMPROVED RACE TECHNIQUE FOR THE FULL SEQUENCE IDENTIFICATION AND AMPLIFICATION OF mRNA ENDS

Rapid amplification of cDNA ends is widely used to identify and obtain nucleotide sequences from the 3' and 5' terminus of genes. Various RACE techniques have been developed to meet different requirements. The presented RACE in this study, is a new procedure to obtain a full cDNA sequence of the mRNA with partially known sequence. Using the extracted mRNA of the Aeluropus littoralis plant and PCR technique, the regions between the known parts and the end regions of the cDNA of the SOS1 gene was specifically amplified. In this method, two types of adapters were anchored to the end of the 5' cDNA at different stages, thus, only 5' end of full-sequenced cDNAs were selected and amplified. The efficiency of this technique was evaluated using amplification of 5' end of the SOS1 in Aeluropus littoralis. Sequencing of the amplified fragment confirmed that the amplified region in the PCR belongs to the AlSOS1 gene. This technique is highly efficient for separation of the unknown regions of cDNA ends, when, only a short sequence of cDNA is available. Furthermore, working with a very small amount of total RNA (about 1μg) and no amplification of the damaged or incomplete sequence of the desired gene as well as no background in PCR product are the advantages of the modified technique.