Lake Finsevatn sediment surface
diversity investigated by
pyrosequencing
Jon Bråte, Dan KristoferRee, KamranShalchian-Tabrizi and Dag Klaveness
Microbial Evolution Research Group (MERG)
Department of Biology, University of Oslo
MERG
Microbial Evolution Research Group
Outline
Lake Finsevatn
PCR, pyrosequencing and sequence handling
Eukaryotic sediment diversity
Freshwater Perkinsea
MERG
Microbial Evolution Research Group
Lake Finsevatn
Photo: Dag Klaveness
MERG
Microbial Evolution Research Group
Lake Finsevatn
Catchment area
From NIVA 1987
MERG
Microbial Evolution Research Group
Sampling
Sampling in March 2009
24 m depth
~2 g of surface sediment
DNA isolated with
PowerSoil (MoBio, USA)
MERG
Microbial Evolution Research Group
454 Pyrosequencing
Massively parallel sequencing
Generates 1.000.000 reads
about 450 bp length
454 Life Sciences (Roche)
MERG
454 Life Sciences (Roche)
Microbial Evolution Research Group
PCR primers
Match a broad diversity of eukaryotes
Target the V4 region of the 18S rDNA gene
Amplicon size ~450 bp
Suitable for the 454 Titanium
pyrosequencing technology
Pyrosequencing
Run 1
Run 2
Combined
Total nr of reads
9937
32557
42494
Nr of reads after
trimming
3755
14642
18397
Nr of reads after
clustering
1640
4511
6049
Removing single
singletons
306
306
306
MERG
Microbial Evolution Research Group
BLAST results
BLAST results displayed with MEGAN (Huson et al. 2007)
MERG
Microbial Evolution Research Group
Perkinsea
Få inn en slide omperkinseadiversitetførvårtfunn Leander?
Modified from Hoppenrath& Leander. J EukaryotMicrobiol. 2009
MERG
Microbial Evolution Research Group
Freshwater Perkinsea
Bråte et al. 2010 ISME
MERG
Microbial Evolution Research Group
Freshwater Perkinsea
MERG
Microbial Evolution Research Group
Acknowledgements
Ramiro Logares, Institut de Ciències del Mar, Barcelona
DipankarBachar and Richard Christen, CNRS & University of Nice
HamidReza Shahbazkia, University of Algarve
Field station at Finse(www.finse.uio.no)
UiO sequencing platform (www.sequencing.uio.no)
MERG
Microbial Evolution Research Group