Perpustakaan Gen: Bagaimana Mengonstruksinya?
Wahyudi, Aris Tri
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Many methods can be used to construct genomic DNA library in the cosmid vector. In this system, large fragments of genomic DNA generated by randomly digestion are ligated to the cosmid vector to form concatemers that can be packaged into bacteriophage λ tparticles. These particles serve as vehicles by which the recombinant DNA molecules are efﬁciently introduced into bacteria. The method for constructing libraries of bacterial genomic DNA in cosmid is to clone genomic fragments of appropriate size (30-45 Kb) obtained by partial digestion with Sau3AI that recognize a 4-bp sequence compat- ible with that of the vector. The partially digested DNA is cloned into the BamHI site of pJRD215 cosmid vector. The recombinant cosmids are subsequently packaged into bakteriphage 1 and infected into the E. coli NM554 as a host strain. Screening of the E. coli harboring recombinant cosmids containing gene(s) of interest is usually performed by colony hybridization.