Construction Of Genomic Dna Library For Volvariella Volvacea

Volvariella volvacea is the fifth edible fungus of the tropics and subtropics. This paper is the construction of genomic DNA library for Volvariella volvacea V23, which will contribute to construct the cold-resistant engineering strain by molecular biological techniques. This library was constructed by partial digestion and the titer is 1.5 *105 pfu/ml. A cloning strategy is used with Lambda GEM-11 phage being choning vector and E..coli LE392 being host strain.The main steps involved in genomic DNA Lambda vector library are: Preparation of high molecular wight genomic DNA (> 100kb) Sau3AI partial digestion of genomic DNA 15-20kb size fractionation of genomic DNA Double digestion of vector with BamHI & EcoRI Ligation of genomic DNA with vector arms Packaging of recombinant DNA and titering of packaged phage Amplification and conservation of genomic DNA library DIG-labeled 5.8s+ITS DNA by PCR was used as probes tohybridizate in situ, and a positive clone was obtained, which shows the good representation of this library.