A method for identifying genes within yeast artificial chromosomes: application to isolation of MLL fusion cDNAs from acute leukaemia translocations.
The majority of chromosomal translocations breakpoints are within regions of the genome where few DNA probes are available. The use of yeast artificial chromosomes (YACs) containing long stretches of human DNA allows dispersed DNA markers to be used to identify the position of breakpoints but does not readily allow subcloning of the precise breakpoint within the YAC DNA nor the cDNAs containing the affected genes. We describe a procedure allowing rapid isolation of cDNAs corresponding to genes within a YAC clone. Random cDNA is hybridised to PCR-generated biotinylated fragments of total DNA from a yeast strain harbouring a YAC clone. The hybrids can be recovered to facilitate subsequent cloning of the cDNA molecules. The application of this method to the cloning of cDNA molecules carrying sequences involved in the translocation t(4;11)(q21;q23) is illustrated.
Forster
A
A
MRC Laboratory of Molecular Biology, Cambridge, UK.
Rabbitts
T H
TH
D000208:Acute Disease; D000595:Amino Acid Sequence; D001483:Base Sequence; D018244:Chromosomes, Artificial, Yeast; D002880:Chromosomes, Human, Pair 11; D002894:Chromosomes, Human, Pair 4; D018076:DNA, Complementary / Q000302:isolation & purification*; D004268:DNA-Binding Proteins / Q000235:genetics*; D011495:Histone-Lysine N-Methyltransferase; D006801:Humans; D007938:Leukemia / Q000235:genetics*; D008969:Molecular Sequence Data; D051788:Myeloid-Lymphoid Leukemia Protein; D011519:Proto-Oncogenes; D014157:Transcription Factors; D014178:Translocation, Genetic
D018076:DNA, Complementary; D004268:DNA-Binding Proteins; C497422:KMT2A protein, human; D014157:Transcription Factors; D051788:Myeloid-Lymphoid Leukemia Protein; D011495:Histone-Lysine N-Methyltransferase