Rapid isolation of genomic clones for individual members of human multigene families: identification and localisation of UBE2L4, a novel member of a ubiquitin conjugating enzyme dispersed gene family.
We describe a rapid, PCR-based, screening procedure for the isolation of human genomic clones in lambda bacteriophage, containing sequences coding for individual homologous members of a multigene family. The approach is based upon the identification, by dilution, of sub-pools of the genomic library that contain members of the gene family, prior to phage isolation. The presence of specific genes is established by PCR of aliquots of individually amplified library pools, using consensus primers and subsequent sequencing. We have used the approach to isolate a fourth member of the UBE2L gene family, UBE2L4, and located it on chromosome 19q13.1-->q13.2. This PCR-based approach to library screening has wider applicability in that it could be used to isolate alternate-spliced products from cDNA libraries.
Ardley
H C
HC
Molecular Medicine Unit, St. James's University Hospital, Leeds, UK.
Moynihan
T P
TP
Thompson
J
J
Leek
J P
JP
Markham
A F
AF
Robinson
P A
PA
D010582:Bacteriophage lambda / Q000235:genetics; D001483:Base Sequence; D002874:Chromosome Mapping; D015698:Genomic Library; D006801:Humans; D008025:Ligases / Q000235:genetics*; D008969:Molecular Sequence Data; D005810:Multigene Family; D016133:Polymerase Chain Reaction; D044763:Ubiquitin-Conjugating Enzymes
C471619:UBE2L4 protein, human; D044763:Ubiquitin-Conjugating Enzymes; D008025:Ligases