CHORI-255
Gorilla (Gorilla gorilla gorilla) (Male)
BAC Library
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The CHORI-255 gorilla 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). To obtain DNA from this endangered species,
a proposal was submitted by Evan Eichler (Case Western Reserve
University, Cleveland) to the Species Survival Plan Coordinator
(SSCP). After favorable review, Dr. Lisa Faust (the SSCP)
kindly provided us a gorilla blood sample. This gorilla was
a male Western lowland gorilla (“Frank”, #327,
Gorilla gorilla gorilla) housed at the Lincoln
Park Zoo (Chicago). High-molecular-weight DNA was isolated
from white blood cells by embedding in agarose. The embedded
DNA was partially digested with a combination of EcoRI restriction
enzyme and EcoRI Methylase and size fractionated by pulsed-field
electrophoresis. DNA fragments from the appropriate size fraction
were cloned into the pTARBAC2.1 vector between the two EcoRI
sites. The ligation products were transformed into DH10B (T1
resistant) electro-competent cells (Invitrogen). The library
has been arrayed into 576 “384-well” microtiter
dishes and has subsequently been gridded onto twelve 22x22cm
nylon high-density filters for probe hybridization screening.
Each hybridization membrane represents over 18,000 distinct
gorilla BAC clones (48 dishes), stamped in duplicate. Library
characterization was performed by Qing Cao and Kazutoyo Osoegawa.
BAC library construction was funded by NIH grants HG01165-07SI
and HG025323-01 as part of the NIH-funded
BAC Resource Network.
click here for a legend of
the previous tables.
| Segment |
1 |
2 |
Total |
| Cloning Vector |
pTARBAC2.1 |
pTARBAC2.1 |
|
| DNA Source |
White Blood Cell |
White Blood Cell |
|
| Restriction Enzyme |
EcoRI |
EcoRI |
|
| Plate Range |
1-288 |
289-576 |
1-576 |
| Total 384-Well Dishes |
288 |
288 |
576 |
| Empty Wells (%) |
1,800 (1.6%) |
1,600 (1.4%) |
3400 (1.5%) |
| Non-insert Clones (%) |
4/278 (1.4%) |
6/277 (2.1%) |
10/555 (1.8%) |
| Vector Clones (%) |
2/110K (0) |
1/110K (0) |
3/220K (0) |
| Recombinant Clones |
>107K |
>106K |
>213K (>96% of wells) |
| Insert size (Average) |
~200Kbp |
~170Kbp |
~185Kbp |
| Standard Deviation |
39Kbp |
34Kbp |
36Kbp |
| Library Redundancy |
~7 fold |
~6 fold |
~13 fold |
Data on the CHORI-255 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 the recipients 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 (pdejong@chori.org,
fax: (510) 450-7924). For hybridization membranes, please
contact BACPAC Resources (BACPACorders@chori.org
Please contact Pieter
J. de Jong or Kazutoyo
Osoegawa for questions related to this library.
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