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2010•Zhonghua laonian xin-nao-xueguanbing zazhiRequires access

Construction and preliminary identification of full-length cDNA library of human pituitary tissue

Huiling Huang

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Abstract

Objective To construct a full-length cDNA library of human normal pituitary tissue. Methods The total RNA was extracted for synthesizing the first strand of cDNA using SMART technique.The double-strand cDNA was amplified by LD-PCR and digested by SfiI restriction enzyme. The cDNA fragments were collected and ligated with modified pBluescript Ⅱ SK vector after fractionation through spin-400 column.The full-length cDNA library was constructed successfully and identified after transforming plasmid to DH-5α bacteria.Results The library contained 1×10~6 clones,in which 93.45% were recombinant.The average size of inserted fragment was more than 1000 bp,the rate of full-length cDNA was 44.4%.Conclusion The cDNA library is eligible and is reliable material for further study of tissue-specific genes of human pituitary.

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What this paper is about

Objective To construct a full-length cDNA library of human normal pituitary tissue. Methods The total RNA was extracted for synthesizing the first strand of cDNA using SMART technique.The double-strand cDNA was amplified by LD-PCR and digested by SfiI restriction enzyme. The cDNA fragments were collected and ligated with modified pBluescript Ⅱ SK vector after fractionation through spin-400 column.The full-length cDNA library was constructed successfully and identified after transforming plasmid to DH-5α bacteria.Results The library contained 1×10~6 clones,in which 93.45% were recombinant.The average size of inserted fragment was more than 1000 bp,the rate of full-length cDNA was 44.4%.Conclusion The cDNA library is eligible and is reliable material for further study of tissue-specific genes of human pituitary.

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Available abstract

Objective To construct a full-length cDNA library of human normal pituitary tissue. Methods The total RNA was extracted for synthesizing the first strand of cDNA using SMART technique.The double-strand cDNA was amplified by LD-PCR and digested by SfiI restriction enzyme. The cDNA fragments were collected and ligated with modified pBluescript Ⅱ SK vector after fractionation through spin-400 column.The full-length cDNA library was constructed successfully and identified after transforming plasmid to DH-5α bacteria.Results The library contained 1×10~6 clones,in which 93.45% were recombinant.The average size of inserted fragment was more than 1000 bp,the rate of full-length cDNA was 44.4%.Conclusion The cDNA library is eligible and is reliable material for further study of tissue-specific genes of human pituitary.

Key concepts: Complementary DNA, cDNA library, Molecular biology, Recombinant DNA, Plasmid, Biology, Rapid amplification of cDNA ends, Restriction enzyme

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