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 Home > Resources > Libraries
   CHORI-237: European Ferret (Mustela furo) BAC Library   

Background information

The CHORI-237 “Ferret” BAC library has been constructed by Boudewijn ten Hallers and Dr. Baoli Zhu in Pieter de Jong’s laboratory at BACPAC Resources, Children’s Hospital Oakland Research Institute. The library preparation followed the general cloning approach developed in our laboratory (Osoegawa et al., 1998). DNA was isolated from frozen internal fetus organs obtained from a female ferret with sample ID FIO-2, kindly provided by John F. Engelhardt, Ph.D., Department of Anatomy and Cell Biology, University of Iowa, Room 1-111, Bowen Science Building, 51 Newton Road, Iowa City, Iowa 52242-1109. Additional sample information: The ferret internal organs were collected from E28 fetuses; father: sable-color coat (Day of birth: 5/15/03) and mother: sable-color coat (Day of birth: 3/2/04).

The frozen kidney tissue was ground under liquid nitrogen into fine powder using pestle and mortar. The powder was suspended in chromatin isolation buffer and further homogenized using a Dounce homogenizer with a tight pestle (7ml homogenizer with a tight pestle with 0.0010"-0.0030" clearance). The chromatin suspension was embedded in 0.5% agarose to generate gel plugs prior to deproteinizing the DNA with cell lysis buffer. The agarose embedded DNA was partially digested with a combination of EcoRI and EcoRI Methylase and size fractionated by pulsed-field electrophoresis. DNA fragments from the appropriate size fraction were cloned into the pBACGK1.1 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 528, 384-well microtiter dishes and has subsequently been gridded onto 11 22x22cm nylon high-density filters for screening by probe hybridization. Each hybridization membrane represents over 18,000 distinct BAC clones, stamped in duplicate.

The BAC library construction was funded by grant (#HG025323-03) under the auspices of the NIH Resource Network.

Segment

1

2

All

Vector

pBACGK1.1

pBACGK1.1

pBACGK1.1

Restriction Enzyme

EcoRI/EcoRI Methylase

EcoRI/EcoRI Methylase

EcoRI/EcoRI Methylase

DNA Source

N/A

N/A

N/A

Plate Numbers

1-288

289-528

1-528

Plate Count

288

240

528

Empty Wells

1250 (1.13%)

1659 (1.8%)

N/A

Non-Recombinant Clones

N/A

N/A

N/A

Non-Insert Clones

Approx. 0 (0/277, 0%)

N/A

N/A

Recombinant Clones

109342

90501

202752

Average Insert Size

147 Kbp

157 Kbp

152 Kbp

Genomic Coverage

N/A

N/A

N/A

Click here for a legend of the previous tables.
No clones were found to be non-recombinant after analysis of CHORI-237 using overgo probes specific for the puc19 fragment.

 

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