Single-stranded conformational polymorphism analysis

Progress in molecular biology has provided a new perspective. Techniques such as the polymerase chain reaction and single-strand conformation polymorphism analysis have greatly facilitated the molecular analysis of erythroenzymopathies. These studies have clarified the correlation between the functional and structural abnormalities of the variant enzymes. In general, the mutations that induce an alteration of substrate binding site and/or enzyme instability might result in markedly altered enzyme properties and severe clinical symptoms. 

Kain, K.C., Craig, A.A. and Ohrt, C. (1996) Single-strand conformational polymorphism analysis differentiates Plasmodium falciparum treatment failures from re-infections. Molecular and Biochemical Parasitology 79, 167—175. 

Sheffield, V.C., Beck, J.S., Kwitek, A.E., Sandstrom, D.W. and Stone, E.M. (1993) The sensitivity of single strand conformation polymorphism analysis for the detection of single base substitutions. Genomics 16, 325-332. 

Beier DR, Dushkin H, Sussman DJ. Mapping genes in the mouse using single-strand conformation polymorphism analysis of recombinant inbred strains and interspecific crosses. Proc Natl Acad Sci USA 1992 89 9102-9106. 

Spire-Vayron de la Moureyre C, Debuysere H, Sabbagh N, Marez D, Vinner E, Chevalier ED et al. Detection of known and new mutations in the thiopurine S-methyltransferase gene by single-strand conformation polymorphism analysis. Hum Mutat 1998 12 177— 185. 

Snyder TM, McGown LB (2005) Multiplex single strand conformation polymorphism analysis by capillary electrophoresis with on-the-fly fluorescence lifetime detection. Appl Spectrosc 59 335-339... 

Maesawa, C Tamura, G Suzuki, Y., Ishida, K., Saito, K., and Satodate, R. (1992) Sensitive detection of p53 gene mutations in esophageal endoscopic biopsy specimens by cell sorting combined with polymerase chain reaction single-strand conformation polymorphism analysis. Jpn. J. Cancer Res. 83, 1253-1256. 

Several rapid and sensitive methods have been developed to detect mutations in cDNA and genomic DNA. These include ribonuclease (RNase) protection analysis, denaturing gradient gel electrophoresis, and single-strand conformation polymorphism analysis. These methods can be used in conjunction with the PCR technique to detect small deletions or insertions and single base substitutions. 

Single-strand conformation polymorphism analysis is the more popular method because of its simplicity and wide applicability to many different genes. This method makes use of... 

Single-strand conformation polymorphism analysis (SSCP, SSCA) Scanning... 

Figure 37-i I A schematic of single-strand conformation polymorphism analysis (SSCP). PCR amplicons carrying the polymorphic site (shown as x and y) are diluted and then denatured to form single-stranded fragments. When the fragments are electrophoresed, they assume secondary structures. When conditions are right, polymorphisms are detected by differential band patterns compared with a known sample. 

Highsmith WE Jr, Nataraj AJ, Jin Q, O Connor JM, Ei-Nabi SH, Kusukawa N, Garner MM, Use of DNA toolbox for the characterization of mutation scanning methods. II evaluation of single-strand conformation polymorphism analysis. Electrophoresis 1999 20 1195-203. 

Suzuki Y, Orita M, Shiraishi M, Hayashi K, Sekiya T. Detection of ras gene mutations in human lung cancers by single-strand conformation polymorphism analysis of polymerase chain reaction products. Oncogene 1990 5 1037-43. 

Cho, J.C. and Kim, S.J. (2000) Computer-assisted PCR-single-strand-conformation polymorphism analysis for assessing shift in soil bacterial community structure during bioremediational treatments. World Journal of Microbiology and Biotechnology, 16, 231-235. 

Figure 12.1. Single-strand conformation polymorphism analysis. Single-stranded DNAs are generated by denaturation of the PCR products and separated on a nondenaturing polyacrylamide gel. A fragment with a single-base modification generally forms a different conformer and migrates difter-ently when compared with the wild-type DNA. Reproduced with permission Ifom Ref 9.

Duthoit F, Godon JJ, Montel MC (2003) Bacterial community dynamics during production of registered designation of origin Salers cheese as evaluated by 16S rRNA gene single-strand conformation polymorphism analysis. Appl Environ Microbiol 69 3840-3848... 

Widjojoatmodjo MN, Fluit AC, Verhoef J (1995) Molecular identification of bacteria by fluorescence-based PCR-single-strand conformation polymorphism analysis of the 16S rRNA gene. J Clin Microbiol 33 2601-2606... 

Matrix-assisted laser desorption ionisation/time of flight (i.e., MALD-TOF) MS Electrospray ionization MS SSCP - Quantitative single-strand conformation polymorphism analysis... 

Duthoit F, Tessier F, Montel M-C (2005). Diversity, d3mamics and acitivity of bacterial populations in registered designation of origin salers cheese by single-strand conformation polymorphism analysis of 16S rRNA genes. J. Appl Microbiol, 98 1198-1208. 

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