A mouse genomic library in the bacteriophage P1 cloning system: organization and characterization
- 1 January 1992
- journal article
- Published by Springer Nature in Mammalian Genome
- Vol. 3 (10) , 550-558
- https://doi.org/10.1007/bf00350620
Abstract
Using the bacteriophage P1 cloning system, we have constructed a two to three times coverage, high-molecular-weight (HMW) genomic library from mouse C127 fibroblast cells. The library consists of about 127,500 clones with an average insert size of about 70 kb that are organized into 300 primary pools containing approximately 425 clones per pool. For screening purposes the primary pools are combined into secondary pools (∼4250 clones each) and tertiary pools (∼21,250 clones each). Screening is performed by the polymerase chain reaction (PCR) with DNA isolated from the secondary and tertiary pools. We have screened the library for 13 different mouse sequences and have detected 11. Clones generated from two of the eleven positive screens were isolated from the library (those containing the c-fos and Gαi2 genes) and were further characterized. Direct double-stranded sequencing of DNA from P1 clones with primers bordering the insert provided sequence information from each end of the cloned DNA.Keywords
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