CDNA cloning a)Red blood cell precursors (c)Digest mRNA with RNase cDNA directs synthesis of second cDNA strand in the presence of four nucleotides and dNa polymerase. Release mRNA from cytoplasm and purify. NIILEEEE cDNA TTTTIIIIIIIIIIAAAA mRNA S1 nuclease DNA polymerase AAAA mRNA cuts hairpin oop EEELE AAAA mRNA 3'CDNA DOuble (b) Add oligo(dT) primer. 5 helix reat with reverse transcriptase in presence of four nucleotides. Insert cDNA into vector 1.[IIIIIIIIIIIIILIAAAA Primer mRNA Growing CDNA 2. AlAJA mRNA 3. A-Halrpin loop Reverse transcriptase