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CHORI-253: Sumatran Orangutan (Pongo pygmaeus) BAC Library |
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The CHORI-253 Sumatran Orangutan BAC library has been constructed by Michael Nefedov in Pieter de Jong’s laboratory at BACPAC Resources, Children’s Hospital Oakland Research Institute. The preparation of the library followed the cloning approach developed in our laboratory (Osoegawa et al., 1998). Evan Eichler (Department of Genetics, Case Western Reserve University) facilitated contacts with Steve Taylor (Louisville Kentucky Zoo) to obtain a blood sample from a male Sumatran orangutan ("Segundo"). DNA was isolated from white blood cells by embedding the cells in agarose. Agarose embedded DNA was partially digested with a combination of EcoRI restriction enzyme and EcoRI Methylase and the fragments were size-fractionated by pulsed-field gel electrophoresis. DNA fragments from the appropriate size fraction were cloned into the pTARBAC2.1 vector between the EcoRI sites. The ligation products were then transformed into DH10B (T1 resistant) electro-competent cells (Invitrogen). The library has been arrayed into 384-well microtiter dishes and also gridded onto eleven 22x22cm nylon high-density filters for screening by probe hybridization. Each hybridization membrane represents over 18,000 distinct Orangutan BAC clones, stamped in duplicate. Library characterization was performed by Qing Cao and Kazutoyo Osoegawa. This work was funded by NIH grant HG01165-06.
| Segment | Cloning Vector | DNA | Restriction enzyme | Plate Number | Total Plates | Empty Wells | Empty Wells (%) | | 1 | pTARBAC2.1 | Blood | EcoRI | 1-288 | 288 | 2,126 | 1.9 | | 2 | pTARBAC2.1 | Blood | EcoRI | 289-528 | 240 | 2,096 | 2.3 | | Total Library | | | | | 528 | 4,222 | 2.1 |
| Segment | Non-insert clones (%) | Non-insert clones (deleted vector) | Non-Recombinant Clones (vector) | Recombinant clones | Insert Size (Average) | Redundancy (Genome Size: 3000 Mb) | | 1 | 0 | 0 | 0 | 108,000 | 177 Kbp | 6.4 | | 2 | 0.4 | ~400 | 0 | 90,000 | 178 Kbp | 5.3 | | Total library | 0.2 | ~400 | 0 | 198,000 | 177 Kbp | 11.7 |
Click here for a legend of the previous tables.
Data on the CHORI-253 clone average insert size has been determined by Pulsed Field Gel Electrophoresis. Clone size distribution has been plotted graphically.
Individual clones identified after screening can be obtained as stab cultures at a cost of $80 per clone in addition to a $35 set-up charge per shipment. Shipping will be done by Federal Express and charged to recipient's account number.
With respect to the scheduling of shipments of arrayed library copies, we would like to make you aware of our policy for array distribution.
Please visit the ordering information page if you want to place an order.
Academic and commercial users interested in a copy of the BAC library should contact Pieter J. de Jong ([email protected], fax: (510) 450-7924). For hybridization membranes, please contact BACPAC Resources ([email protected]).
Please contact Pieter J. de Jong for questions related to this library.
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