J Integr Plant Biol. ›› 1992, Vol. 34 ›› Issue (6): -.

• Research Articles •    

Construction of Alfalfa Genomic Library and Some Improvements in Methodology

Guan Chang-hui, Zheng Hong-gang,Wang Bin and Jing Yu-xiang   

Abstract: A genomic library of alfalfa (Medicago sativa L.) was constructed using λ bacteriophageEMBL4 as vector. The number of the phage recombinants obtained was 8.75× l05 pfu which reached the desired capacity of the library, with some modifications of the frequently adopted methods in the experiments, high quality of the library was obtained. The use of urea and CTAB (cetyltrimethylammonium bromide) in the preparation of genomic DNA increased the yield and removed the polysaccharides that inhibit many enzymes. The vector and donor DNAs were recovered by centrifugation through a sucrose density gradient, which could not only protect the cohesive ends of the DNA molecules, but also avoid the agarose contamination produced easily from electric elution or Iow melting point agarose in DNA recovery. Both the DNA insertion identification of the recombinant phages and the preliminary screening with an alfalfa leghemoglobin cDNA probe, proved the efficiency of the library.

Key words: Alfalfa, Genomic library, Leghemoglobin gene

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