Direct construction of a chromosome-specific NotI linking library from flow-sorted chromosomes.
A linking library consists of genomic DNA fragments which contain a specific rare restriction enzyme site; such clones are very useful as probes in pulsed field gel electrophoresis and in mapping and cloning large regions of DNA. However, identifying those linking clones which map to a certain chromosomal region can be laborious. Therefore, we have developed a straightforward procedure for constructing a linking library directly from flow-sorted chromosomes. As a test of the approach, a NotI linking library was constructed from the chromosome 17 fraction of a flow-sort of human chromosomes, using only 70 ng of DNA. Thirteen of sixteen linking clones were mapped to chromosome 17, suggesting that the library is highly enriched for this chromosome. This method should be generally applicable to other chromosomes and enzymes as well.
Wallace
M R
MR
Howard Hughes Medical Institute, University of Michigan, Ann Arbor.
Fountain
J W
JW
Brereton
A M
AM
Collins
F S
FS
D002874:Chromosome Mapping; D002886:Chromosomes, Human, Pair 17; D003001:Cloning, Molecular; D015252:Deoxyribonucleases, Type II Site-Specific; D008040:Genetic Linkage; D005822:Genetic Vectors; D006801:Humans; D008214:Lymphocytes; D009693:Nucleic Acid Hybridization; D015183:Restriction Mapping
D015252:Deoxyribonucleases, Type II Site-Specific; C038565:GCGGCCGC-specific type II deoxyribonucleases