Gene Mapping and Positional Cloning


Gene mapping and positional cloning are used to identify a gene or genes that, when variant or mutated, contribute to the causation of a genetically defined disorder or condition.

Keywords: disorders; genetic; single; multigenic; common; variations

Figure 1.

Complementary deoxyribonucleic acid (cDNA) selection. The target DNA is labeled with biotin and repetitive DNA removed by a prehybridization step. This leaves nonrepetitive, biotin‐labeled target DNA single stranded, which is then annealed to cDNA amplified up from an appropriate library, using vector‐primed polymerase chain reaction (PCR). Streptavidin magnetic beads have a high affinity for biotin and can be used simply to isolate all the DNA that contains biotin. PCR amplification with the cDNA vector primers will amplify only cDNA that is homologous to the target DNA – all other cDNA is lost, because it has no biotin. The process can be repeated for three or more cycles.

Figure 2.

Exon trapping. (a) The exon‐trapping vector, with inserted target DNA that contains no exons, is expressed in the appropriate human cells and its ribonucleic acid (RNA) spliced to the vector donor (D) and acceptor (A) sites; reverse transcriptase (RT)‐PCR of messenger RNA (mRNA) yields no product, because the inserted DNA transcript is spliced out. (b) If a fragment of DNA containing an exon is inserted into the vector, a new splice pattern occurs. RT‐PCR of the cellular mRNA will result in a PCR product that contains the inserted target DNA exon. This can be sequenced or cloned as required.


Further Reading

Church DM and Buckler AJ (1999). Gene identification by exon amplification. Methods in Enzymology 303: 83–99.

Deloukas P, Schuler GD, Gyapay G, et al. (1998) A physical map of 30000 human genes. Science 282: 744–746.

Flavell RA, Kooter JM, De Boer E, Little PF and Williamson R (1978) Analysis of the beta‐delta‐globin gene loci in normal and Hb Lepore DNA: direct determination of gene linkage and intergene distance. Cell 15: 25–41.

Riordan JR, Rommens JM, Kerem B, et al. (1989) Identification of the cystic fibrosis gene: cloning and characterization of complementary DNA. Science 245: 1066–1073.

Simmons AD and Lovett M (1999) Direct cDNA selection using large genomic DNA targets. Methods in Enzymology 303: 111–126.

The International Human Genome Mapping Consortium (2001) A physical map of the human genome. Nature 409: 934–941.

Web Links

Celera Genomics.

Ensemble Project.

NCBI (National Center for Biotechnology Information).

UCSC Genome Bioinformatics.

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How to Cite close
Little, Peter FR(Sep 2005) Gene Mapping and Positional Cloning. In: eLS. John Wiley & Sons Ltd, Chichester. [doi: 10.1038/npg.els.0005370]