The PrimeScript Double Strand cDNA Synthesis Kit contains all of the reagents necessary to synthesize double-stranded cDNA from purified poly A+ RNA templates using the Gubler-Hoffman method (Gubler and Hoffman 1983). First-strand cDNA is synthesized using PrimeScript RT and either oligo(dT) primers or random primers. After cDNA synthesis, E. coli RNase H7 is used to nick the RNA portion of mRNA-cDNA hybrids. E. coli DNA Polymerase I is subsequently used for second-strand synthesis by nick translation, and E. coli DNA Ligase repairs breaks in the second strand. Finally, T4 DNA Polymerase is used to blunt the ends. The resulting double-stranded cDNA can be cloned into an appropriate vector to generate a cDNA library.