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《Journal of Fruit Science》 2004-03
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Construction and Application of A Genomic DNA Library of Actinidia deliciosa

Xu Wenping, Xu Changjie, Chen Kunsong*, and Zhang Shanglong (Lab of Fruit Molecular Physiology and Biology/The State Agriculture Ministry Laboratory of Horticultural Plant Growth,Development & Biotechnology, Huajiachi Campus, Zhejiang University, Hangzhou,Zhejiang 310029 China)  
Genomic DNA was extracted from Actinidia deliciosa with a modified CTAB method, and purified by CsCl density gradient centrifugation. The purified DNA was then digested with Sau3A I. In order to remove the fragments less than 14 kb, the partially digested DNA was further size-fractioned by preparative gel electrophoresis and then the fragments between 14 to 23 kb were recovered from the gel by electroelution. The fragments were filled in with dATP and dGTP before ligated to Lambda FIX II vector, and the ligation products were incubated with "Gigapack III Gold" (Stratagene) packaging extracts. As a result,we constructed a genomic DNA library with a primary titer of 1.05×106 and the titer of amplified library is 2.5×109 pfu/mL. To evaluate the representation of the constructed library, the phages in the library were served as PCR template as well as genomic DNA with 6 pairs of specific primers designed from either reserved regions of plant genes or kiwifruit genes. Fragments of expected size were amplified for both templates.
【Fund】: 国家重点基础研究发展规划项目(973项目)(G2000046806);; 国家自然科学基金(30270917);; 浙江省自然科学基金重点资助项目(ZD0004)。
【CateGory Index】: S663.4
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