The CHORI-14 library was generated by Chung Li Shu and Kazutoyo Osoegawa. Construction was funded by a grant from the National Human Genome Research Institute (NHGRI, NIH) (#HG01165-06). This library was generated according to the new NHGRI/DOE "Guidance on Human Subjects in Large-Scale DNA Sequencing".
Male blood was obtained via a double-bind selection protocol. Male blood DNA was isolated from one randomly chosen donor (out of 10 male donors) and partially digested with a combination of EcoRI and EcoRI Methylase. Size selected DNA was cloned into the pTARBAC2 vector between the EcoRI sites. The ligation products were transformed into DH10B electrocompetent cells (BRL Life Technologies). The library has been arrayed into 384-well microtiter dishes and also gridded onto 22x22cm nylon high density filters for screening by probe hybridization.

 

Segment
Cloning
Vector
DNA
Restriction Enzyme
Plate Numbers
Total Plates
Empty Wells
Empty Wells (%)
1
pTARBAC2

Male blood
EcoRI
1-178, 185-288
282
2,694
2.5
2
pTARBAC2

Male blood
EcoRI
289-528
240
2,147
2.3
total
1-178, 185-528
522
4,841
2.4

 

Segment
Non-insert Clones (%)
Non-insert clones
Non-recombinant clones (BAC vector)
Recombinant clones
Insert size (Average)
Redundancy (Genome size:
3,3 Mb)
1
4.7
approx. 4,963
N/A
approx. 100,631
163 Kbp
5.0
2
2.9
approx. 2610
N/A
approx. 87,403
168 Kbp
4.5
total
3.7
approx. 7,573
N/A
approx. 188,034
165 Kbp
9.5


click here for a legend of the previous tables.


Data on the CHORI-14 clone average insert size has been determined by Pulsed Field Gel Electrophoresis. Clone size distribution has been plotted graphically.