cDNA Selection: Efficient PCR Approach for the Selection of cDNAs Encoded in Large Chromosomal DNA Fragments

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 88; no. 21; pp. 9623 - 9627
• Main Authors: Parimoo, Satish, Patanjali, Sankhavaram R., Shukla, Hridayabhiranjan, Chaplin, David D., Weissman, Sherman M.
• Format: Journal Article
• Language: English
• Published: Washington, DC National Academy of Sciences of the United States of America 01.11.1991; National Acad Sciences
• Subjects: Base Sequence; Biological and medical sciences; CDNA libraries; Cloning, Molecular - methods; Complementary DNA; Cosmids; DNA; DNA - genetics; DNA probes; Fundamental and applied biological sciences. Psychology; Gene Library; Genes. Genome; Genetic hybridization; Genetic Vectors; Genomic Library; Genomics; histocompatibility locus HLA-A; Humans; Libraries; Molecular and cellular biology; Molecular genetics; Molecular Sequence Data; Nucleic Acid Hybridization; Oligonucleotides - chemistry; Polymerase chain reaction; Polymerase Chain Reaction - methods; Repetitive Sequences, Nucleic Acid; yeast artificial chromosomes; Yeasts; Polymerase chain reaction; Screening; Molecular hybridization; Complementary DNA; Major histocompatibility system; Cosmid; Nucleotide fragment; Identification; Yeast artificial chromosome
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.88.21.9623

Identification of coding segments in large fragments of genomic DNA is a recurrent problem in genome mapping and positional cloning studies. We have developed a rapid and efficient protocol to achieve this goal, based on hybridization of cDNA fragments to immobilized DNA and recovery of the selected cDNAs by the PCR. The procedure permits rapid cloning of cDNA fragments encoded by large genomic DNA fragments, groups of yeast artificial chromosomes, or cosmids and has the potential to directly enrich cDNAs encoded in chromosome segments. By this approach we have been able to identify several non-major histocompatibility complex class I clones from a yeast artificial chromosome that includes the HLA-A locus.