An efficient and reliable method for cloning PCR-amplification products: a survey of point mutations in integrin cDNA.

Starr L, Quaranta V
Biotechniques. 1992 13 (4): 612-8

PMID: 1476731

A highly efficient, non-labor-intensive method for cloning DNA fragments produced by PCR amplification was used to carry out a rapid survey of potential point mutations in integrin alpha 6 cDNA from 17 different cell-type sources. The method includes glass powder purification of the PCR reaction mixture, followed by simultaneous treatment with T4 polynucleotide kinase and DNA polymerase I, and another glass powder purification. Sequences from multiple subclones of each cell type were readily generated, aligned and checked for mismatches. Several commonly used alternative procedures were compared for cloning efficiency and size-fidelity of inserted DNA fragments.

MeSH Terms (16)

Amino Acid Sequence Animals Base Sequence Cloning, Molecular DNA DNA Polymerase I Humans Integrins Mice Molecular Sequence Data Point Mutation Polymerase Chain Reaction Polynucleotide 5'-Hydroxyl-Kinase Rats Sequence Alignment Sequence Homology, Nucleic Acid

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