Isolation of 2 novel RFLP markers and their localization at 2q35 by microdissection and subsequent enzymatic amplification

We previously constructed a chromosome 2q-specific genomic libary and isolated a number of microclones. In the present study, we first analyzed with Southern hybridization whether any of the microclones represent restriction fragment length polymorphisms (RFLPs) and then tried to map RFLP markers physically, using the recently developed chromosome microdissection/enzymatic amplification method. Of 13 clones analyzed, two were RFLP markers; a clone, pM2C83, showed a fouralleleMspI RFLP, and the other, pM2C8, a two-alleleRsaI RFLP. In order to assign the two polymorphic markers, two chromsomal segments, 2q32-q35 and 2q35-qter, on the chromosome 2 from a karyotypically normal person were microdissected, and the DNA from each segment was amplified with the polymerase chain reaction (PCR) using marker sequence-specific primers. With this method, both of the clones were assigned to 2q35. These two RFLP markers must be useful for linkage analysis of genetic diseases whose loci are at around 2q35.