title: Marine microbial communities 0.26 K
status: Public on Apr 24 2010
contact.city: Tucson
contact.first: Virginia
contact.middle: Isabel
contact.last: Rich
contact.email: vrich@email.arizona.edu
contact.state: AZ
contact.address: 1041 E. Lowell St.
contact.country: USA
contact.institute: University of Arizona
organism: uncultured crenarchaeote 4B7
  Prochlorococcus marinus subsp. pastoris str. CCMP1986
  uncultured marine alpha proteobacterium
  uncultured marine gamma proteobacterium EBAC31A08
  uncultured marine group II euryarchaeote 37F11
  uncultured Pseudomonadota bacterium
  uncultured crenarchaeote 74A4
  uncultured marine bacterium 440
  uncultured marine bacterium 577
  uncultured marine bacterium 583
  uncultured gamma proteobacterium eBACHOT4E07
  uncultured marine bacterium EB0_41B09
  uncultured proteobacterium 60D04
  uncultured proteobacterium 65D09
  uncultured marine group II euryarchaeote EF100_57A08
  uncultured marine bacterium EB000_55B11
description: update to platform GPL6012. DeLongLab_GenomeProxyArray_v2.0_268targets. Note supplementary platform for full array design and sequences of probes.
manufacture_protocol: Glass slides with poly-lysine coating. The expanded genome proxy array was designed as in (Rich et al., 2008). Briefly, each genotype was targeted using suites of ~20 70-mer oligonucleotide probes designed using the program ArrayOligoSelector (Zhu et al., 2003). Probes had approximately the same %GC (40%) and were distributed across the target genome or genome fragment, with no more than one probe per gene and avoiding 16S and 23S rRNA genes. The probes were selected solely based on theoretical thermodynamic properties and GC content; that is, probe selection did not focus on specific genes or regions, but simply produced the “optimal” probes for each genome proxy based on the probes’ predicted hybridization properties. The array included positive and negative control probes designed using the same method, to Halobacterium salinarum NRC-1 and a random genome sequence, respectively.    The expanded array had a broader scope than the prototype of Rich et al., 2008 (268 target genotypes, as opposed to the prototype’s 14) and included a co-spot oligo for spot alignment and gridding purposes (using the “alien” oligo sequence of (Urisman et al., 2005). The targets were selected from fully-sequenced marine microbial genomes, publicly-available marine-derived BAC and fosmid clone sequences, and fully-sequenced clones from the lab’s Monterey Bay and Hawaii environmental BAC- and fosmid-based genomic libraries. Targeted genotypes are detailed in Table S1, summarized in Table S2, and presented in a schematic phylogenetic overview in Fig. 1. Previously-unpublished sequences used for array design were submitted to Genbank under accession numbers GU474833-GU474949.    Oligonucleotides were synthesized (Illumina, San Diego, CA), suspended in 3X SSC to a concentration of 40 pmol ml-1, and spotted on homemade poly-L-lysine-coated glass slides using a QArray 2 microarraying robot (Genetix, Hampshire, UK). Six replicates of each probe were spotted.