The KB-1 consortium was maintained in batch culture in defined mineral medium
[65 (link)] with trichloroethene (TCE) as electron acceptor and methanol as electron donor. The culture was routinely allowed to dechlorinate TCE completely to ethene prior to a new amendment of acceptor/donor approximately every two weeks. The DonnaII reactor and ANAS semi-batch reactor were maintained as described previously
[19 (link),20 (link)]. See Table
1 for a summary comparison of maintenance conditions, and Table
2 for metagenome sizes and modes of sequencing used.
For the KB-1 metagenome, DNA was extracted from the T3 MP1 KB-1 culture just after completion of a dechlorination cycle using a Cetyl trimethylammonium bromide (CTAB) protocol
[66 ] with volumes scaled up for higher yield as described in the alternate protocol, omitting subsequent cesium chloride gradient centrifugation steps. Clone libraries with 35-kb and 3-kb inserts were created by the JGI using their in-house protocols (
http://www.jgi.doe.gov/sequencing/protocols ), and an additional 3 kb short insert library generated for construction of a shotgun metagenome microarray was constructed by Genome Atlantic. A total of 103 MB of metagenome sequence was generated on AB13730xl Sanger sequencers from the three clone libraries. The metagenome was assembled using the JGI’s in-house bacterial assembly pipeline, utilizing lucy
[67 (link)] for vector and quality screening. The KB-1 metagenome sequence and assembly were made publically available by the JGI on May 2nd, 2009 (
http://genome.jgi-psf.org/aqukb/aqukb.download.ftp.html ). Community composition of the three KB-1 DNA samples utilized for sequencing the KB-1 metagenome was determined by Dr. Alison S. Waller using qPCR
[30 ].
The DonnaII metagenome was generated from 454 Titanium libraries according to the JGI’s in-house protocols (
http://www.jgi.doe.gov/sequencing/protocols ). Metagenome assembly was conducted using the Newbler program from Roche
[68 (link)]. The sequence data, including an in-house assembly draft for the DonnaII metagenome was made publically available by the JGI on January 31st, 2012 (IMG-M taxon ID: 2032320001 (
http://img.jgi.doe.gov/cgi-bin/m/main.cgi )).
The ANAS metagenome was composed of a combination of Sanger sequencing and Titanium 454 sequencing as described above. The sequence data, including an in-house assembly draft for the ANAS metagenome was made publically available by the JGI on August 20th, 2009 (IMG-M taxon ID: 2014730001 (
http://img.jgi.doe.gov/cgi-bin/m/main.cgi )).
[65 (link)] with trichloroethene (TCE) as electron acceptor and methanol as electron donor. The culture was routinely allowed to dechlorinate TCE completely to ethene prior to a new amendment of acceptor/donor approximately every two weeks. The DonnaII reactor and ANAS semi-batch reactor were maintained as described previously
[19 (link),20 (link)]. See Table
For the KB-1 metagenome, DNA was extracted from the T3 MP1 KB-1 culture just after completion of a dechlorination cycle using a Cetyl trimethylammonium bromide (CTAB) protocol
[66 ] with volumes scaled up for higher yield as described in the alternate protocol, omitting subsequent cesium chloride gradient centrifugation steps. Clone libraries with 35-kb and 3-kb inserts were created by the JGI using their in-house protocols (
[67 (link)] for vector and quality screening. The KB-1 metagenome sequence and assembly were made publically available by the JGI on May 2nd, 2009 (
[30 ].
The DonnaII metagenome was generated from 454 Titanium libraries according to the JGI’s in-house protocols (
[68 (link)]. The sequence data, including an in-house assembly draft for the DonnaII metagenome was made publically available by the JGI on January 31st, 2012 (IMG-M taxon ID: 2032320001 (
The ANAS metagenome was composed of a combination of Sanger sequencing and Titanium 454 sequencing as described above. The sequence data, including an in-house assembly draft for the ANAS metagenome was made publically available by the JGI on August 20th, 2009 (IMG-M taxon ID: 2014730001 (
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