Whole‐genome Amplification


Whole‐genome amplification (WGA) techniques utilize the principles of the polymerase chain reaction (PCR) to copy enzymatically the deoxyribonucleic acid (DNA) contained within a sample. However, unlike most PCR applications, the amplification is nonspecific and theoretically provides an increase in the copy number of all genomic sequences. WGA is used in areas of research where minute DNA samples are routinely encountered. Applications include preimplantation genetic diagnosis, the creation of probes from microdissected or flow‐sorted chromosomes, forensics and the study of ancient DNA samples.

Keywords: DOP‐PCR; T‐PCR; alu‐PCR; linker adapter‐PCR; primer extension preamplification

Figure 1.

Degenerate oligonucleotide primed polymerase chain reaction (DOP‐PCR). (a) Sample deoxyribonucleic acid (DNA) is added to a reaction mixture containing a semidegenerate primer. The primer consists of two specified ‘tag’ sequences separated by several random nucleotides. (b) Low annealing temperatures allow the primers to anneal at many sites throughout the genome. (c) Initiation of DNA synthesis from these sites permits the entire genome to be copied. (d) The fragments generated in this way can also serve as templates during later cycles. (e–g) After several rounds of amplification most fragments contain regions complementary to the tag sequences at both ends. Next the PCR annealing temperatures are increased. At these higher temperatures primers can only successfully anneal to sequences complementary to their tags. (h) Fragments that contain these sequences at each end are amplified in a highly efficient exponential fashion.

Figure 2.

Linker adapter polymerase chain reaction (LA‐PCR). (a) Genomic deoxyribonucleic acid (DNA) template. (b) Template is digested with a restriction enzyme. (c) Adapter oligonucleotides are ligated and (d) serve as sites for the annealing of PCR primers.



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Further Reading

Cotter FE, Das S, Douek E, Carter NP and Young BD (1991) The generation of DNA probes to chromosome 11q23 by Alu PCR on small numbers of flow‐sorted 22q‐derivative chromosomes. Genomics 9(3): 473–480.

Kristjansson K, Chong SS, Van den Veyver IB, et al. (1994) Preimplantation single cell analyses of dystrophin gene deletions using whole genome amplification. Nature Genetics 6(1): 19–23.

Lengauer C, Green ED and Cremer T (1992) Fluorescence in situ hybridization of YAC clones after Alu‐PCR amplification. Genomics 13(3): 826–828.

Mao YW, Liang SY, Song WQ and Li XL (1998) Construction of a DNA library from chromosome 4 of rice (Oryza sativa) by microdissection. Cell Research 8(4): 285–293.

Montgomery KD, Tedford KL and McDougall JK (1995) Genetic instability of chromosome 3 in HPV‐immortalized and tumorigenic human keratinocytes. Genes Chromosomes and Cancer 14(2): 97–105.

Telenius H, Pelmear AH, Tunnacliffe A, et al. (1992) Cytogenetic analysis by chromosome painting using DOP‐PCR amplified flow‐sorted chromosomes. Genes Chromosomes and Cancer 4(3): 257–263.

Voullaire L, Slater H, Williamson R and Wilton L (2000) Chromosome analysis of blastomeres from human embryos by using comparative genomic hybridization. Human Genetics 106(2): 210–217.

Weier HU, Polikoff D, Fawcett JJ, et al. (1994) Generation of five high‐complexity painting probe libraries from flow‐sorted mouse chromosomes. Genomics 21(3): 641–644.

Wells D and Delhanty JDA (2000) Comprehensive chromosomal analysis of human pre‐implantation embryos using whole genome amplification and single cell comparative genomic hybridization. Molecular Human Reproduction 6(11): 1055–1062.

Wells D and Delhanty JDA (2001) Preimplantation genetic diagnosis: applications for molecular medicine. Trends in Molecular Medicine 7: 23–30.

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Wells, Dagan(Sep 2005) Whole‐genome Amplification. In: eLS. John Wiley & Sons Ltd, Chichester. http://www.els.net [doi: 10.1038/npg.els.0005341]