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biomphalaria genome initiative

A high quality BAC library for (BB02) Biomphalaria glabrata


Literature reference

Coen M Adema, Mei-Zhong Luo, Ben Hanelt, Lynn A Hertel, Jennifer J Marshall, Si-Ming Zhang, Randall J DeJong, Hye-Ran Kim, David Kudrna, Rod A Wing, Cari Soderlund, Matty Knight, Fred A Lewis, Roberta Lima Caldeira, Liana K Jannotti-Passos, Omar dos Santos Carvalho, Eric S Loker (2006) A bacterial artificial chromosome library for Biomphalaria glabrata, intermediate snail host of Schistosoma mansoni. Mem Inst Oswaldo Cruz, 101(Suppl. I): 167-177, 2006


Order BAC library resources

Goto http://www.genome.arizona.edu/orders/ .
In left menu bar View BAC/EST Resources click "Genomic - Libraries, Clones, & Filters"
Scroll to BG_BBa and click on Biomphalaria glabrata cv. BB02
make your choice and order individual clones, high density filters or the complete library




Construction of the BAC library for Biomphalaria glabrata

Background

On behalf of the Biomphalaria glabrata genome initiative, a white paper requesting the construction of a bacterial artificial chromosome (BAC) library  for the snail Biomphalaria glabrata, was submitted to the National Human Genome Research Institute (NHGRI). In July 2002, review by the Genome Resources and Sequencing Priorities Panel (GRASPP) of the NHGRI BAC Resource Steering Panel resulted in a "high priority" recommendation, concluding that the proposal was "worth accepting based on the medical importance alone". This project was assigned to the laboratory of Rod Wing, Arizona Genomics Institute, University of Arizona, Tucson AZ, which belongs to the National Institutes of Health (NIH) BAC Resource Network. The library was produced to meet a predetermined set of BAC Library Quality Assessment Standards. Details regarding this Biomphalaria glabrata project are publicly posted on the BAC Library Proposals and Information web page of NHGRI. Since the positive recommendation from the Prioritization Panel, NHGRI and the Biomphalaria glabrata Genome Initiative have interactively agreed to employ a Biomphalaria glabrata strain (field isolate) that is susceptible to Schistosma mansoni to generate the BAC library. Thus, molecular data collected from the Biomphalaria glabrata BAC library will provide a relevant context for study of the intramolluscan biology of schistosomes.


Working with the Arizona Genomics Institute in Tucson AZ, methods were adapted for isolation of high molecular weight genomic DNA from B. glabrata. Exploratory efforts using M-line snails identified the ovotestis of adult snails (shell diameter at least 10mm) as preferred tissue for the required DNA.

For production of the BAC library, template DNA was isolated from multiple BB02 snails (F2) bred from a single parent snail by selfing. HindIII restriction fragments were cloned into pAGIBAC. In total 61,824 clones were picked and the average insert size is estimated at ~136kb, this yields about 9.05x genome coverage.

For quality assesment both ends of 96 BAC clones were sequenced (GenBank DX360154-360158, DX360097-360114), and ptobing of high density filters of the BAC library with (single- and) low copy genes indicated adequate coverage of the genome by the library.


BB02BACfilters

The first set of filters was probed at UNM for quality control (February 2004). Shown is a BB02 snail placed on the stack of filters in a sealed plastic bag. The label with the library name "BG_BB" is visible from the top filter, the library set contains 4 filters (a,b,c,d) of 25 x25 cm.


The first round of QC probing with pooled probes was completed April 2004. The autorad above shows several positive (double) spots. Next, characterization of individual BAC clones confirmed the occurrence of target sequences.


The quality control phase was completed satisfactorily and the production of the BAC library was reported in Memórias do Instituto Oswaldo Cruz. At this time, individual clones, filters and replicate libraries are available to international (non-profit) researchers at cost.

The genome sequencing strategy by WUGSC includes the sequencing of the ends of essentially all BAC clones in the library. The contig assembly of BAC ends will provide further necessary groundwork for larger scale sequencing efforts.


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