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. 1986 Nov 11;14(21):8615–8624. doi: 10.1093/nar/14.21.8615

Efficient construction of cDNA libraries in plasmid expression vectors using an adaptor strategy.

H Haymerle, J Herz, G M Bressan, R Frank, K K Stanley
PMCID: PMC311881  PMID: 3024111

Abstract

We describe a method for the construction of large DNA fragment libraries in plasmid vectors, in which complementary, single-stranded extensions are ligated onto both vector and insert DNA using un-phosphorylated adaptor oligonucleotides. Special consideration has been taken of the requirements of expression screening as follows: cDNA synthesis using random oligonucleotide primers is described which maximises the probability of obtaining open reading frame fragments from large mRNA molecules, the adaptors use codons found in high abundance E. coli proteins to minimise problems of premature termination when using strong promoters, and the sequence encoded by the adaptors, when cloned into the bacterial expression vector pEX1, promotes a surface location for the foreign antigenic determinant where it is accessible to antibodies used for screening.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

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