Abstract
Single-strand conformation polymorphism (SSCP) analysis is a sensitive and rapid technique for detecting DNA polymorphisms and mutations in PCR-amplified fragments. Due to its technical simplicity, it is widely used as a screening tool in various investigations, ranging from clinical diagnosis of human hereditary diseases to the characterization of microbial communities. This method can also be used successfully on phytoplasmas as a tool for the detection of molecular variability in conserved housekeeping genes such as 16S rRNA and tuf, as well as in more variable genes, revealing the presence of polymorphisms undetected by routine RFLP analyses. The reliability of SSCP has been confirmed by multiple alignments and phylogenetic analyses of representative sequences showing different SSCP profiles. However, it is not broadly applied in phytoplasma research yet. The technique provides an inexpensive, convenient, and sensitive method for determining sequence variation and to differentiate phytoplasma strains, and is particularly suitable for epidemiological studies or as a fast screening, typing tool when dealing with a large number of field samples.
This is a preview of subscription content, log in via an institution.
Buying options
Tax calculation will be finalised at checkout
Purchases are for personal use only
Learn about institutional subscriptionsReferences
Orita M et al (1989) Detection of polymorphisms of human DNA by gel electrophoresis as single-strand conformation polymorphisms. Proc Natl Acad Sci U S A 86:2766–2770
Orita M et al (1989) Rapid and sensitive detection of point mutations and DNA polymorphisms using the polymerase chain reaction. Genomics 5:874–879
Telenti A (1994) Detection of mutations by PCR-SSCP (single-strand conformation polymorphism). In: Dale JW, Sanders PG (eds) Methods in gene technology, vol 2. JAI Press Ltd, Middlesex, UK, pp 241–252
Rubio L et al (1996) Differentiation of citrus tristeza closterovirus (CTV) isolates by single-strand conformation polymorphism analysis of the coat protein gene. Ann Appl Biol 129:479–489
Ravnik-Glavač M, Glavač D, Dean M (1994) Sensitivity of single-strand conformation polymorphism and heteroduplex method for mutation detection in the cystic fibrosis gene. Hum Mol Genet 3:801–807
Hall JS et al (2001) Structure and temporal dynamics of populations within wheat streak mosaic virus isolates. J Virol 75:10231–10243
Yap EP, McGee JO (1992) Nonisotopic SSCP detection in PCR products by ethidium bromide staining. Trends Genet 8:49
Hongyo T et al (1993) ‘Cold SSCP’: a simple, rapid and non-radioactive method for optimized single-strand conformation polymorphism analyses. Nucleic Acids Res 21:3637–3642
Palacio A, Duran-Vila N (1999) Single-strand conformation polymorphism (SSCP) analysis as a tool for viroid characterisation. J Virol Methods 77:27–36
Ren J-C (2000) High-throughput single-strand conformation polymorphism analysis by capillary electrophoresis. J Chromatogr B 741:115–128
Kozlowski P, Krzyzosiak WJ (2001) Combined SSCP/duplex analysis by capillary electrophoresis for more efficient mutation detection. Nucleic Acids Res 29:e71
Papp T et al (2007) Mutational analysis of Chk1, Chk2, Apaf1 and Rb1 in human malignant melanoma cell lines. Oncol Rep 17:135–140
Silveira APF et al (2011) Gene mutations and polymorphisms of TP53 and FHIT in chronic esophagitis and esophagealcarcinoma. Anticancer Res 31:1685–1690
Mackiewicz V et al (2005) Rapid investigation of hepatitis A virus outbreak by single strand conformation polymorphism analysis. J Med Virol 76:271–278
Di Liberto G et al (2006) Clinical and therapeutic implications of hepatitis C virus compartmentalization. Gastroenterology 131:76–84
Simsek S et al (2011) Molecular discrimination of sheep and cattle isolates of Echinococcus granulosus by SSCP and conventional PCR in Turkey. Vet Parasitol 178:367–369
Schaschl H, Wegner KM (2006) Polymorphism and signature of selection in the MHC class I genes of the three-spined stickleback Gasterosteus aculeatus. J Fish Biol 69(suppl B):177–188
Dassanayake RS, Samaranayake LP (2003) Amplification-based nucleic acid scanning techniques to assess genetic polymorphism in Candida. Crit Rev Microbiol 29:1–24
Nahiyan ASM et al (2011) PCR-SSCP analysis of Fusarium diversity in asparagus decline in Japan. Eur J Plant Pathol 130:197–203
Oto M, Suda W, Shinoyama H (2006) 16S rRNA gene-based analysis of microbial community by whole-genome amplification and minigel-single-strand conformation polymorphism technique. J Biosci Bioeng 102:482–484
Turturo C et al (2005) Genetic variability and population structure of Grapevine leafroll-associated virus 3 isolates. J Gen Virol 86:217–224
Cerni S et al (2008) Stem pitting and seedling yellows symptoms of Citrus tristeza virus infection may be determined by minor sequence variants. Virus Genes 36:241–249
Šeruga Musić M, Krajačić M, Škorić D (2008) The use of SSCP analysis in the assessment of phytoplasma gene variability. J Microbiol Methods 73:69–72
Schneider B, Seemüller E (2009) Strain differentiation of Candidatus Phytoplasma mali by SSCP- and sequence analyses of the hflB gene. J Plant Pathol 91:103–112
Igloi GL (1983) A silver stain for the detection of nanogram amounts of tRNA following two-dimensional electrophoresis. Anal Biochem 134:184–188
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2013 Springer Science+Business Media, LLC
About this protocol
Cite this protocol
Musić, M.Š., Škorić, D. (2013). Single-Strand Conformation Polymorphism Analysis for Differentiating Phytoplasma Strains. In: Dickinson, M., Hodgetts, J. (eds) Phytoplasma. Methods in Molecular Biology, vol 938. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-089-2_18
Download citation
DOI: https://doi.org/10.1007/978-1-62703-089-2_18
Published:
Publisher Name: Humana Press, Totowa, NJ
Print ISBN: 978-1-62703-088-5
Online ISBN: 978-1-62703-089-2
eBook Packages: Springer Protocols