Chromosome in situ hybridization to localize single copy genes

Lorraine Sohlberg lorraine at NATURE.BERKELEY.EDU
Wed Oct 9 15:32:19 EST 1996


Hello, I am developing techniques at the NSF Center for Plant and Microbial
Developmental Biology at Univ of Calif., Berkeley. I have used in situ
hybridization to identify the chromosomal position of a tandem repeat
sequence and am now I am interested in using in situ hybridization to
detect single copy genes on maize, corn, meiotic pachytene chromosomes.
This hasn't been done much in plants and I was wondering if I could get
feedback from the world of bionet about successful techniques. I have a few
questions regarding the probes used. I understand that the cloned sequence
used to create the probe must be rather large in order to get a decent
signal. In addition it can not contain any repeated sequence in an intron
or spacer region that my light up elsewhere in the genome. I was also
wondering how the probes are synthesized. I have been using DIG labelled
RNA probes and they work well to locate the tandem repeat.
Has anyone used PRINS or other PCR techniques?  Thanks for your help, in
advance, Lorraine





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