Optimization of RNA isolation from the archaebacterium Methanosarcina barkeri and validation for oligonucleotide microarray analysis

Journal of Microbiological Methods
David E CulleyWeiwen Zhang

Abstract

The recent completion of a draft genome sequence for Methanosarcina barkeri has allowed the application of various high throughput post-genomics technologies, such as nucleic acid microarrays and mass spectrometry of proteins to detect global changes in transcription and translation that occur in response to experimental treatments. However, due to the production of a thick heteropolysaccharide outer layer, M. barkeri usually grows in large aggregates of cells rather than as individual, planktonic cells. Complete disruption of these aggregates and lysis of the released cells presents technical difficulties in ensuring the isolation of intact RNA from the entire population of cells. Initial attempts at isolating RNA from M. barkeri using several standard extraction protocols gave incomplete lysis of cells and resulted in low yields of poor quality RNA. In this study, we tested several chemical and mechanical disruption modifications of standard RNA extraction methods to optimize the extraction efficiency and minimize the number of unlysed cells remaining after extraction. As a further test of the quality of the resulting RNAs, their performance in replicate microarray analyses were determined. The results showed that inclusion o...Continue Reading

References

May 1, 1983·The Journal of Clinical Endocrinology and Metabolism·G G KrishnaJ R Sowers
Mar 8, 2000·Current Microbiology·G N JarvisK N Joblin
Aug 26, 2000·Journal of Bacteriology·S Karlin, J Mrázek
Apr 5, 2002·Genome Research·James E GalaganBruce Birren
Nov 19, 2002·Applied Microbiology and Biotechnology·U DeppenmeierC Prust
Apr 1, 1997·The Journal of General and Applied Microbiology·Kazunari UshidaJean-Pierre Jouany
Jun 5, 2004·Journal of Microbiological Methods·W Van DesselJ Anné
Oct 27, 2004·BMC Biotechnology·Ulrika LiljedahlAnn-Christine Syvänen
May 20, 2005·Molecular Genetics and Genomics : MGG·Raymond HoveyUwe Deppenmeier

❮ Previous
Next ❯

Citations

Nov 10, 2009·Canadian Journal of Microbiology·Wei TangZongbao Kent Zhao
Mar 28, 2013·Archaea : an International Microbiological Journal·Patrick D Browne, Hinsby Cadillo-Quiroz
Aug 31, 2013·Folia Microbiologica·Simon Rittmann, Peter Holubar
Mar 28, 2013·Archaea : an International Microbiological Journal·Elisabeth W VissersHendrikus J Laanbroek
Feb 26, 2011·Archaea : an International Microbiological Journal·Jasper WaltherJohn van der Oost
Sep 17, 2008·Journal of Cellular and Molecular Medicine·Fei GaoLining Zhang
May 20, 2014·Applied and Environmental Microbiology·Pengfei LiuYahai Lu

❮ Previous
Next ❯

Related Concepts

Related Feeds

Archaeogenetics

Recent advances in genomic sequencing has led to the discovery of new strains of Archaea and shed light on their evolutionary history. Discover the latest research on Archaeogenetics here.