Evaluation of molecular techniques in characterization of deep terrestrial biosphere

Malin Bomberg (Corresponding Author), Mari Nyyssönen, Aura Nousiainen, J. Hultman, L. Paulin, P. Auvinen, Merja Itävaara

Research output: Contribution to journalArticleScientificpeer-review

Abstract

A suite of molecular methods targeting 16S rRNA genes (i.e., DGGE, clone and high-throughput [HTP] amplicon library sequencing) was used to profile the microbial communities in deep Fennoscandian crystalline bedrock fracture fluids. Variation among bacterial 16S rRNA genes was examined with two commonly used primer pairs: P1/P2 and U968f/U1401r. DGGE using U968f/ U1401r mostly detected ß-, ?-proteobacteria and Firmicutes, while P1/P2 primers additionally detected other proteobacterial clades and candidate divisions. However, in combination with clone libraries the U968f/U1401r primers detected a higher bacterial diversity than DGGE alone. HTP amplicon sequencing with P1/P2 revealed an abundance of the DGGE bacterial groups as well as many other bacterial taxa likely representing minor components of these communities. Archaeal diversity was investigated via DGGE or HTP amplicon sequencingusing primers A344F/ 519RP. The majority of archaea detected with HTP amplicon sequencing belonged to uncultured Thermoplasmatales and Pendant 33/DHVE3, 4, 6 groups. DGGE of the same samples detected mostly SAGMEG and Methanosarcinales archaea, but almost none of those were revealed by HTP amplicon sequencing. Overall, our results show that the inferred diversity and composition of microbial communities in deep fracture fluids is highly dependent on analytical technique and that the method should be carefully selected with this in mind.
Original languageEnglish
Pages (from-to)468-487
JournalOpen Journal of Ecology
Volume4
Issue number8
DOIs
Publication statusPublished - 2014
MoE publication typeA1 Journal article-refereed

Fingerprint

biosphere
microbial community
clone
fluid
gene
targeting
analytical method
bedrock
library
evaluation
method

Keywords

  • DGGE
  • 16S rRNA Gene
  • amplicon library
  • sequencing
  • deep subsurface

Cite this

Bomberg, Malin ; Nyyssönen, Mari ; Nousiainen, Aura ; Hultman, J. ; Paulin, L. ; Auvinen, P. ; Itävaara, Merja. / Evaluation of molecular techniques in characterization of deep terrestrial biosphere. In: Open Journal of Ecology. 2014 ; Vol. 4, No. 8. pp. 468-487.
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abstract = "A suite of molecular methods targeting 16S rRNA genes (i.e., DGGE, clone and high-throughput [HTP] amplicon library sequencing) was used to profile the microbial communities in deep Fennoscandian crystalline bedrock fracture fluids. Variation among bacterial 16S rRNA genes was examined with two commonly used primer pairs: P1/P2 and U968f/U1401r. DGGE using U968f/ U1401r mostly detected {\ss}-, ?-proteobacteria and Firmicutes, while P1/P2 primers additionally detected other proteobacterial clades and candidate divisions. However, in combination with clone libraries the U968f/U1401r primers detected a higher bacterial diversity than DGGE alone. HTP amplicon sequencing with P1/P2 revealed an abundance of the DGGE bacterial groups as well as many other bacterial taxa likely representing minor components of these communities. Archaeal diversity was investigated via DGGE or HTP amplicon sequencingusing primers A344F/ 519RP. The majority of archaea detected with HTP amplicon sequencing belonged to uncultured Thermoplasmatales and Pendant 33/DHVE3, 4, 6 groups. DGGE of the same samples detected mostly SAGMEG and Methanosarcinales archaea, but almost none of those were revealed by HTP amplicon sequencing. Overall, our results show that the inferred diversity and composition of microbial communities in deep fracture fluids is highly dependent on analytical technique and that the method should be carefully selected with this in mind.",
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Evaluation of molecular techniques in characterization of deep terrestrial biosphere. / Bomberg, Malin (Corresponding Author); Nyyssönen, Mari; Nousiainen, Aura; Hultman, J.; Paulin, L.; Auvinen, P.; Itävaara, Merja.

In: Open Journal of Ecology, Vol. 4, No. 8, 2014, p. 468-487.

Research output: Contribution to journalArticleScientificpeer-review

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T1 - Evaluation of molecular techniques in characterization of deep terrestrial biosphere

AU - Bomberg, Malin

AU - Nyyssönen, Mari

AU - Nousiainen, Aura

AU - Hultman, J.

AU - Paulin, L.

AU - Auvinen, P.

AU - Itävaara, Merja

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N2 - A suite of molecular methods targeting 16S rRNA genes (i.e., DGGE, clone and high-throughput [HTP] amplicon library sequencing) was used to profile the microbial communities in deep Fennoscandian crystalline bedrock fracture fluids. Variation among bacterial 16S rRNA genes was examined with two commonly used primer pairs: P1/P2 and U968f/U1401r. DGGE using U968f/ U1401r mostly detected ß-, ?-proteobacteria and Firmicutes, while P1/P2 primers additionally detected other proteobacterial clades and candidate divisions. However, in combination with clone libraries the U968f/U1401r primers detected a higher bacterial diversity than DGGE alone. HTP amplicon sequencing with P1/P2 revealed an abundance of the DGGE bacterial groups as well as many other bacterial taxa likely representing minor components of these communities. Archaeal diversity was investigated via DGGE or HTP amplicon sequencingusing primers A344F/ 519RP. The majority of archaea detected with HTP amplicon sequencing belonged to uncultured Thermoplasmatales and Pendant 33/DHVE3, 4, 6 groups. DGGE of the same samples detected mostly SAGMEG and Methanosarcinales archaea, but almost none of those were revealed by HTP amplicon sequencing. Overall, our results show that the inferred diversity and composition of microbial communities in deep fracture fluids is highly dependent on analytical technique and that the method should be carefully selected with this in mind.

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KW - DGGE

KW - 16S rRNA Gene

KW - amplicon library

KW - sequencing

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DO - 10.4236/oje.2014.48040

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JO - Open Journal of Ecology

JF - Open Journal of Ecology

SN - 2162-1985

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