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© 1993 Oxford University Press

RESEARCH-ARTICLE

Identification of human chromosome 9 specific genes using exon amplification

Deanna M. Church, Laura T. Banks, Anne C. Rogers1, Sharon L. Graw1,{dagger}, David E. Housman1, James F. Gusella and Alan J. Buckler*

Molecular Neurogenetics Unit, Massachusetts General Hospital and Department of Genetics, Harvard Medical School Boston, MA 02114 1Center for Cancer Research, Massachusetts Institute of Technology Cambridge, MA 02139, USA

To whom correspondence should be addressed

Received June 25, 1993; Revised August 29, 1993; Accepted August 29, 1993

We have recently developed a method, exon amplification, that is designed for isolation of exon sequences from genomic DNA. To assess the efficacy of this method we have analyzed cosmid genomic clones derived from human chromosome 9, and have cloned several products from this analysis. Approximately 63% of cosmids produced at least one product derived from functioning splice sites within the target genomic fragment, and In many cases multiple products were isolated. In addition, an easily identifiable class of false positives was produced from 56% of cosmids analyzed; these are readily eliminated from subsequent study. Sequence analysis and database searches revealed that the majority (87%) of the putative exon clones were unique, the remainder being derived from repetitive sequences. Analysis of sequence conservation by Southern blotting in addition to cDNA screening experiments suggested that most, If not all, of these unique sequences represent true exons. The results of these studies indicate that exon amplification is a rapid and reliable approach for isolation of exon sequences from mammalian genomic DNA.

{dagger}Present address: Eleanor Roosevelt Institute, Denver, CO 80206, USA


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