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Capillary electrophoresis ribosomal RNA single-stranded conformation polymorphism: a new approach for characterization of low-diversity microbial communities

journal contribution
posted on 2023-05-17, 13:26 authored by Nai, YH, Zemb, O, Gutierrez-Zamora, M-L, Manefield, M, Shane PowellShane Powell, Michael BreadmoreMichael Breadmore
Capillary electrophoresis (CE) has been the principle system for nucleic acid analysis since the early 1990s due to its inherent advantages such as fast analysis time, high resolution and efficiency, minimal sample requirement, high detection sensitivity, and automation. In the past few decades, microbial community fingerprinting methods such as terminal restriction fragment length polymorphism and single-stranded conformation polymorphism (SSCP) have migrated to CE to utilize its advantages over conventional slab gel electrophoresis. Recently, a gel-based direct rRNA fingerprint method was demonstrated. Different from other existing microbial community characterization approaches, this novel approach is polymerase chain reaction free and capable of providing information on the relative abundance of rRNA from individual phylotypes in low-diversity samples. As a gel-based method, it has a long analysis time and relatively large reagent and sample requirements. Here, we addressed these limitations by transferring the RNA fingerprint approach to the CE platform. Analysis time significantly improved from 24 h to 60 min, and the use of a fluorescently labeled hybridization probe as the detection strategy decreased the sample requirement by ten-fold. The combination of fast analysis time, low sample requirement, and sensitive fluorescence detection makes CE-RNA-SSCP an appealing new approach for characterizing low-diversity microbial communities.

History

Publication title

Analytical and Bioanalytical Chemistry

Volume

404

Issue

6-7

Pagination

1897-1906

ISSN

1618-2642

Department/School

Tasmanian Institute of Agriculture (TIA)

Publisher

Springer-Verlag Heidelberg

Place of publication

Tiergartenstrasse 17, Heidelberg, Germany, D-69121

Rights statement

CopyrightSpringer-Verlag 2012

Repository Status

  • Restricted

Socio-economic Objectives

Expanding knowledge in the agricultural, food and veterinary sciences