ABSTRACT
Subcloning or insertion of a gene or DNA fragment into an appropriate vector is an essential technique that is widely used in molecular biology today. Major advantages include: (1) DNA fragments of interest can be subcloned into a high copy number of plasmid vector and can be ampli„ed up to 300-fold by plasmid replication in Escherichia coli; (2) vectors used for subcloning usually contain SP6, T7, or T3 promoters upstream from the polycloning sites, which allows one to prepare sense RNA or antisense RNA of the inserted cDNA as hybridization probes; (3) the SP6, T7, and T3 promoters enables an investigator to sequence the DNA insert on both strands in opposite directions; and (4) DNA constructs can be prepared by inserting a gene or cDNA of interest into an appropriate vector for gene transfer and expression in cultured cells and animals.
