Single-Strand Conformation Polymorphism Fingerprint Method for Dictyostelids

Overview

Journal Front Microbiol

Specialty Microbiology

Date 2021 Sep 13

PMID 34512585

Citations 2

Authors

Phongthana Pasookhush

Asmatullah Usmani

Kowit Suwannahong

Prasit Palittapongarnpim

Kamolchanok Rukseree

Kanchiyaphat Ariyachaokun

Sureemas Buates

Suradej Siripattanapipong

Pravech Ajawatanawong

Affiliations

Soon will be listed here.

Abstract

Dictyostelid social amoebae are a highly diverse group of eukaryotic soil microbes that are valuable resources for biological research. Genetic diversity study of these organisms solely relies on molecular phylogenetics of the gene, which is not ideal for large-scale genetic diversity study. Here, we designed a set of PCR-single-strand conformation polymorphism (SSCP) primers and optimized the SSCP fingerprint method for the screening of dictyostelids. The optimized SSCP condition required gel purification of the SSCP amplicons followed by electrophoresis using a 9% polyacrylamide gel under 4°C. We also tested the optimized SSCP procedure with 73 Thai isolates of dictyostelid that had the gene sequences published. The SSCP fingerprint patterns were related to the genus-level taxonomy of dictyostelids, but the fingerprint dendrogram did not reflect the deep phylogeny. This method is rapid, cost-effective, and suitable for large-scale sample screening as compared with the phylogenetic analysis of the gene sequences.

Citing Articles

Limitations of 18S rDNA Sequence in Species-Level Classification of Dictyostelids.

Chittavichai T, Sathitnaitham S, Utthiya S, Prompichai W, Prommarit K, Vuttipongchaikij S Microorganisms. 2025; 13(2).

PMID: 40005642 PMC: 11857562. DOI: 10.3390/microorganisms13020275.

Recent advancements in nanomaterial-based biosensors for diagnosis of breast cancer: a comprehensive review.

Yazdani Y, Jalali F, Tahmasbi H, Akbari M, Talebi N, Shahrtash S Cancer Cell Int. 2025; 25(1):50.

PMID: 39966938 PMC: 11834589. DOI: 10.1186/s12935-025-03663-8.

Circulating Nucleic Acids in Colorectal Cancer: Diagnostic and Prognostic Value.

Igder S, Zamani M, Fakher S, Siri M, Ashktorab H, Azarpira N Dis Markers. 2024; 2024:9943412.

PMID: 38380073 PMC: 10878755. DOI: 10.1155/2024/9943412.

Detection of SARS-CoV-2 spike protein D614G mutation using μTGGE.

Juma K, Morimoto K, Sharma V, Sharma K, Biyani R, Biyani M Mol Biol Rep. 2024; 51(1):289.

PMID: 38329653 DOI: 10.1007/s11033-023-09065-1.

References

Mathews D, Turner D . Prediction of RNA secondary structure by free energy minimization. Curr Opin Struct Biol. 2006; 16(3):270-8. DOI: 10.1016/j.sbi.2006.05.010. View

King J, Insall R . Chemotaxis: finding the way forward with Dictyostelium. Trends Cell Biol. 2009; 19(10):523-30. DOI: 10.1016/j.tcb.2009.07.004. View

Barroso A, Dunner S, Canon J . Technical note: Detection of bovine kappa-casein variants A, B, C, and E by means of polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP). J Anim Sci. 1998; 76(6):1535-8. DOI: 10.2527/1998.7661535x. View

Sunnucks P, Wilson A, Beheregaray L, Zenger K, French J, Taylor A . SSCP is not so difficult: the application and utility of single-stranded conformation polymorphism in evolutionary biology and molecular ecology. Mol Ecol. 2000; 9(11):1699-710. DOI: 10.1046/j.1365-294x.2000.01084.x. View

Stamatakis A . RAxML version 8: a tool for phylogenetic analysis and post-analysis of large phylogenies. Bioinformatics. 2014; 30(9):1312-3. PMC: 3998144. DOI: 10.1093/bioinformatics/btu033. View

Hauser P, Blanc D, Sudre P, Senggen Manoloff E, Nahimana A, Bille J . Genetic diversity of Pneumocystis carinii in HIV-positive and -negative patients as revealed by PCR-SSCP typing. AIDS. 2001; 15(4):461-6. DOI: 10.1097/00002030-200103090-00004. View

Bassam B, Gresshoff P . Silver staining DNA in polyacrylamide gels. Nat Protoc. 2007; 2(11):2649-54. DOI: 10.1038/nprot.2007.330. View

Sheikh S, Thulin M, Cavender J, Escalante R, Kawakami S, Lado C . A New Classification of the Dictyostelids. Protist. 2018; 169(1):1-28. DOI: 10.1016/j.protis.2017.11.001. View

Calvo-Garrido J, Carilla-Latorre S, Kubohara Y, Santos-Rodrigo N, Mesquita A, Soldati T . Autophagy in Dictyostelium: genes and pathways, cell death and infection. Autophagy. 2010; 6(6):686-701. DOI: 10.4161/auto.6.6.12513. View

10.

Olive D, Bean P . Principles and applications of methods for DNA-based typing of microbial organisms. J Clin Microbiol. 1999; 37(6):1661-9. PMC: 84917. DOI: 10.1128/JCM.37.6.1661-1669.1999. View

11.

Olsen G, McCarroll R, Sogin M . Secondary structure of the Dictyostelium discoideum small subunit ribosomal RNA. Nucleic Acids Res. 1983; 11(22):8037-49. PMC: 326558. DOI: 10.1093/nar/11.22.8037. View

12.

Hayashi K . PCR-SSCP: a method for detection of mutations. Genet Anal Tech Appl. 1992; 9(3):73-9. DOI: 10.1016/1050-3862(92)90001-l. View

13.

Gouy M, Guindon S, Gascuel O . SeaView version 4: A multiplatform graphical user interface for sequence alignment and phylogenetic tree building. Mol Biol Evol. 2009; 27(2):221-4. DOI: 10.1093/molbev/msp259. View

14.

Lorenz R, Bernhart S, Honer Zu Siederdissen C, Tafer H, Flamm C, Stadler P . ViennaRNA Package 2.0. Algorithms Mol Biol. 2011; 6:26. PMC: 3319429. DOI: 10.1186/1748-7188-6-26. View

15.

Mesquita A, Cardenal-Munoz E, Dominguez E, Munoz-Braceras S, Nunez-Corcuera B, Phillips B . Autophagy in Dictyostelium: Mechanisms, regulation and disease in a simple biomedical model. Autophagy. 2016; 13(1):24-40. PMC: 5240833. DOI: 10.1080/15548627.2016.1226737. View

16.

Bozzaro S, Eichinger L . The professional phagocyte Dictyostelium discoideum as a model host for bacterial pathogens. Curr Drug Targets. 2011; 12(7):942-54. PMC: 3267156. DOI: 10.2174/138945011795677782. View

17.

Heras J, Dominguez C, Mata E, Pascual V, Lozano C, Torres C . GelJ--a tool for analyzing DNA fingerprint gel images. BMC Bioinformatics. 2015; 16:270. PMC: 4549892. DOI: 10.1186/s12859-015-0703-0. View

18.

Romeralo M, Cavender J, Landolt J, Stephenson S, Baldauf S . An expanded phylogeny of social amoebas (Dictyostelia) shows increasing diversity and new morphological patterns. BMC Evol Biol. 2011; 11:84. PMC: 3073913. DOI: 10.1186/1471-2148-11-84. View

19.

Shannon C . The mathematical theory of communication. 1963. MD Comput. 1997; 14(4):306-17. View

20.

Chrambach A, Rodbard D . Polyacrylamide gel electrophoresis. Science. 1971; 172(3982):440-51. DOI: 10.1126/science.172.3982.440. View