BioSample

The poly (A)+ RNA was dephosphorylated with bacterial alkaline phosphatase (BAP) and then decapped with tabacco acid pyrophosphatase (TAP). The decapped intact mRNA was ligated with DNA-RNA linker including EcoR I site by treatment of T4 RNA ligase and the first strand cDNA was synthesized from oligo dT-selected mRNA by priming with dT-tailed vector. The dT-tailed vector was adjusted to have about 60nt. The cDNA vector was circularized with E. coli DNA ligase after digestion of EcoRI which site is also included in vector. An RNA strand converted to a DNA strand by Okayama-Berg method. The obtained cDNA vectors were used for transformation of competent cells E. coli Top10F' by electroporation method. The cDNA libraries constructed by this method are full-length enriched cDNA library.