Many strategies for isolating novel genes produce only a segment of the target DNA sequences, usually located towards 3' ends of the cDNAs. Thus, polymerase chain reaction (PCR)-based techniques were developed to aid isolation of full-length cDNAs. However, these techniques are often difficult, with low rates of success. We describe improvements to DNA ligase- dependent amplification PCR applied using 3' sequence information derived from cloned differential gene-display PCR products. We have also modified Alu-PCR, using primers specific for mammalian interspersed Alu repetitive sequence, to permit rapid cloning of upstream regulatory DNA sequences from genomic DNA. In combination, the PCR techniques described provide a complete strategy for successful cloning of full-length cDNA and corresponding upstream regulatory DNA sequences.
|Number of pages||6|
|Journal||Methods in Molecular and Cellular Biology|
|Publication status||Published - 1995|