2-DIMENSIONAL SEPARATION AND CLONING OF CHROMOSOME-1 NOTI-ECORV-DERIVED GENOMIC FRAGMENTS

The two-dimensional (2-D) separation of genomic digests has provided t he means to analyze over 2000 unique restriction fragments simultaneou sly in a single gel, for genetic variation as well as for genomic alte rations in cancer, Ey utilizing different combinations of restriction enzymes or different electrophoretic conditions, the number of analyza ble fragments in multiple 2-D patterns can be augmented, We have previ ously shown the feasibility of distinguishing between spot intensities representing fragments from one allele and from two alleles and have implemented approaches for the cloning of fragments of interest in 2-D gels, In this study, the 2-D separation and cloning of chromosome 1 N otI-EcoRV-derived genomic fragments was performed, Three hundred forty -six NotI fragments in whole genomic preparations were assigned to chr omosome I, To verify the reliability of the assignment, two of the Not I fragments attributed to chromosome 1 were cloned and sequenced. The fragments that contained CpG islands mere mapped by FISH to 1p35-p36.1 and to 1p13.3-p21, respectively. Our study indicates the feasibility of analyzing 2-D separations of whole genomic digests for the detectio n of alterations in specific chromosomes, The large number of restrict ion fragments attributed to chromosome 1 provides the means to screen 2-D patterns for chromosome 1 deletions and amplifications with a high marker density. (C) 1996 Academic Press, Inc.