Samples were acquired by the scientific drilling vessel JOIDES Resolution at IODP Site U1385A on November 25th, 2011 by Advanced Piston Coring (APC).
|Created||December 2, 2015|
|Modified||September 30, 2016|
|State||Final no updates expected|
Iberian Margin Anaerobic Sediment Metagenomes.
Samples were acquired by the scientific drilling vessel JOIDES Resolution at IODP Site U1385A (37°34.285’N, 10°7.562’W) on November 25th, 2011 by Advanced Piston Coring (APC). Whole-round sections meant for molecular analysis were immediately frozen at -80 degrees C for the remainder of the cruise and shipped at this temperature. DNA was extracted using the MO BIO PowerMax Soil DNA Isolation Kit (MO BIO Laboratories Inc., Carlsbad, CA, USA) with a minor adjustment to manufacturer’s protocol. Briefly, 10 g of sediment extruded from the interior (>1cm from core liner) of whole-round core samples was processed according to manufacturer’s instructions, save for an additional step of incubation in a 65 degree C water bath for 15 minutes prior to step 4 (10 minute vortex). Extractions did not yield enough genomic DNA for direct Illumina library preparation. Thus, 2 ng of genomic DNA from each sample was added to the GenomePlex Whole Genome Amplification (WGA) kit (Sigma-Aldrich, Inc., St. Louis, MO, USA). Reactions were run for 20 cycles at manufacturer suggested denaturing and annealing temperatures. Triplicate reactions were pooled and products were purified using the GenElute PCR Clean-Up kit (Sigma-Aldrich Inc., St. Louis, MO, USA). Samples were sent for Illumina library preparation and 2 x 150 bp paired-end-sequencing was performed on 2 lanes of a Illumina HiSeq2500 machine at the University of Delaware Sequencing and Genotyping Center in the Delaware Biotechnology Institute according to manufacturer’s instructions. Over 85,000,000 reads were generated across 6 samples.
Reads were quality checked in the CLC-BIO Genomics Workbench. Poor quality scores were detected over the first 30 bp of the majority of the reads, likely an artifact of identical nucleotides in those positions due to the 30 bp universal primer sites ligated to each read in the WGA step. Thus, the first 30 bp of each read was removed and assembly was performed on resulting 120 bp paired end reads. Sequences were assembled using CLC-BIO and IDBA-UD (Peng et al. 2012) up to 120 kmer length. CLC-BIO assembly was run with default parameters. IDBA-UD assembly was run with the following flags called: –mink 40, –maxk 120, — step 20, –num_threads 18, –min_contig 300. N50 scores and maximum contig length were consistently higher using IDBA-UD and further analysis was performed on assembled data from IDBA-UD.
General term for a laboratory apparatus commonly used for performing polymerase chain reaction (PCR). The device has a thermal block with holes where tubes with the PCR reaction mixtures can be inserted. The cycler then raises and lowers the temperature of the block in discrete, pre-programmed steps.
unique sample identification or number; any combination of alpha numeric characters; precise definition is file dependent
taxonomic group or entity. This may be a family, class, genus, species, etc.; usually this parameter will contain a mixture of taxonomic entities.
brief description, open ended, specific to the data set in which it appears
|Joseph A. Russell||University of Delaware|
|Jennifer F. Biddle||University of Delaware|
|Shannon Rauch||University of Delaware||✓|
BCO-DMO Project Info
|Project Title||Genomic analyses and microbial cultivations in unexplored sub-seafloor ridge flank and continental margin environments|
|Acronym||Subseafloor Microbial Ecology|
|Created||November 19, 2015|
|Modified||November 19, 2015|
Project description from C-DEBI:
Over the course of two years of C-DEBI support, I have investigated subseafloor microbial ecology in three separate environments; the basaltic crust aquifer underneath the sediments of North Pond, the sediments of North Pond, and the sediments of the Iberian Margin at IODP site U1385.
At North Pond, my research was primarily cultivation-based, with enrichments for multiple metabolisms across basalt and sediment samples. Shallow and deep heterotrophic isolates from the sediment column at site U1382B offer an opportunity to ask unique research questions regarding the breakdown of fresher, more labile organic carbon vs. older, more refractory organic carbon.
At the Iberian margin, my research was primarily molecular-based, with several enrichment and cultivation efforts initiated after compelling evidence for particular metabolisms associated with individual groups of microbes. Diversity studies using high-throughput sequencing of 16S/18S rRNA amplicons examined the distribution and abundance of bacteria, archaea, and microbial eukaryotes. To further investigate ecological trends and the biology of particular community members, metagenomes were generated from the same DNA pools as the amplicon data.
This project was funded by a C-DEBI Graduate Student Fellowship.