Single-Strand Conformation Polymorphism Fingerprint Method for Dictyostelids

Phongthana Pasookhush; Asmatullah Usmani; Kowit Suwannahong; Prasit Palittapongarnpim; Kamolchanok Rukseree; Kanchiyaphat Ariyachaokun; Sureemas Buates; Suradej Siripattanapipong; Pravech Ajawatanawong

doi:10.3389/fmicb.2021.708685

Single-Strand Conformation Polymorphism Fingerprint Method for Dictyostelids

Front Microbiol. 2021 Aug 27:12:708685. doi: 10.3389/fmicb.2021.708685. eCollection 2021.

Affiliations

¹ Division of Bioinformatics and Data Management for Research, Research Division, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand.
² Department of Microbiology, Faculty of Science, Mahidol University, Bangkok, Thailand.
³ Department of Biology, Faculty of Education, Kandahar University, Kandahar, Afghanistan.
⁴ Department of Environmental Health, Faculty of Public Health, Burapa University, Chonburi, Thailand.
⁵ National Science and Technology Development Agency (NSTDA), Thailand Science Park, Khlong Nueng, Thailand.
⁶ Department of Sciences and Liberal Arts, Mahidol University, Amnatcharoen Campus, Bung, Thailand.
⁷ Department of Biological Sciences, Faculty of Science, Ubon Ratchathani University, Ubon Ratchathani, Thailand.

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 SSU rDNA 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 SSU rDNA 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 SSU rDNA gene sequences.

Keywords: SSCP; SSU rDNA gene; dictyostelids; diversity; fingerprint.