P. micropora ChIP-purified DNA
Paulinella micropora
P. micropora MYN1 strain was cultured under 14 hours light /10 hours dark photoperiodic cycle at 25 degrees C in the modified Waris-H+Si medium. Light intensity was set at 30-40 microE m-2s-1.
sample_name
P. micropora ChIP-purified DNA
biomaterial_provider
Mami Nomura, Ken-ichiro Ishida, Graduate School of Life and Environmental Science, University of Tsukuba
biotic_relationship
free living
collection_date
2009-10-09
env_broad_scale
fresh water
env_local_scale
pond
env_medium
missing
estimated_size
1.35 Gb
geo_loc_name
Japan:Ibaraki, Tsukuba
isol_growth_condt
10.1111/jeu.12102
lat_lon
36.0485 N 140.1190 E
num_replicons
missing
ploidy
missing
project_name
Genome- and transcriptome-analysis of the photosynthetic testate amoeba Paulinella micropora
propagation
eukaryote: asexual
samp_collect_device
P. micropora cells were harvested by low-speed centrifugation (500 x g, 2 minutes) at 4 degrees C. The harvested cells were passed through miracloth (Merk Millipore, Darmstadt, Germany) and 20 micrometer mesh-filter (HD-20, Nippon Rikagaku kikai co. Ltd, Tokyo, Japan) to eliminate contaminating bacteria that were enriched in the dead P. micropora cell aggregates larger than 20 micrometer, and followed by the washing with 10 mM Tris-HCl (pH8.0) three times, and with 10 mM Tris-HCl (pH8.0) plus 10 mM EDTA six times. Next, to remove the cell debris smaller than 5 micrometer, the cells were trapped on 5 micrometer mesh-filter (PP-5n, Kyoshin Rikoh Inc, Tokyo, Japan) and washed with 5 ml TE buffer. The recovered cells were kept at -80 degrees C.
samp_mat_process
P. micropora cells were homogenized in presence of 500 microliter homogenizing buffer (20 mM Tris-HCl (pH 7.6), 10 mM NaCl, 10 mM KCl, 2.5 mM EDTA, 250 mM sucrose, 0.1 mM spermine, 0.5 mM spermidine, 1 mM DTT) with 30 micrometer clearance glass homogenizer (RD440911, Teraoka co., Ltd, Osaka, Japan). The homogenates were centrifuged at 1000 x g for 10 minutes at 4 degrees C, and the pellets were suspended in 300 microliter ChIP buffer (50 mM Tris-HCl (pH 8.0), 500 mM NaCl, 10 mM EDTA, 0.1% SDS, 0.5% Na-deoxycholate, 1% Triton X-100, 1 mM DTT, 10% glycerol) with 20 microliter Dynabeads protein G (Life technologies) charged with 1 microgram anti-histone H3 antibody (ab1791, Abcam plc, Cambridge UK), and incubated at 4 degrees C for 20 minutes. The Dynabeads were recovered, and washed with ChIP buffer twice, and with glycerol-free ChIP buffer twice, and was finally suspended in DNA extraction buffer (10 mM Tris-HCl (pH 8.0), 1 mM EDTA, 1% SDS). After RNaseA (10 microgram/ml)- and ProteinaseK (200 microgram/ml)- treatments, the ChIP-purified DNAs were extracted by using Plant DNeasy mini kit (Qiagen).
strain
MYN1
trophic_level
photoautotroph
BioSampleModel
MIGS.eu