Faculty Research 1990 - 1999

Title

The mouse Y chromosome: enrichment, sizing, and cloning by bivariate flow cytometry.

Document Type

Article

Publication Date

1998

Keywords

Animal, DNA, Gene-Library, In-Situ-Hybridization-Fluorescence, Interleukin-2, Mice, Mice-Inbred-C57BL, Molecular-Sequence-Data, Polymerase-Chain-Reaction, Repetitive-Sequences-Nucleic-Acid, Sequence-Homology-Nucleic-Acid, Spleen, Y-Chromosome, Chromosomes, Human, Pair 16, Cloning, Molecular, Flow Cytometry

JAX Source

Genomics 1998 Mar 15;48(3):304-13

Grant

CA34196/CA/NCI, RR01315/RR/NCRR

Abstract

In this report, we demonstrate the utility of interleukin-2 (IL-2) stimulation of spleen cell cultures and bivariate flow cytometry in the analysis and purification of the C57BL/6J mouse Y Chromosome. We determined that the DNA content of the C57BL/6J Y Chromosome is approximately 94.7 Mb, making it similar in size to human Chromosome 16 and significantly larger than previous estimates. In addition, we describe the bulk isolation of mouse Y Chromosomes and demonstrate enrichment of the isolated material using a fluorescence in situ hybridization strategy. We detail the construction of two small insert Y Chromosome-specific libraries, ideal for sampling Y Chromosome sequences. From these libraries 1566 clones were analyzed. We provide a detailed characterization of 103 clones, generating nearly 50 kb of sequence. For 30 of these clones, we identify regions of homology to known Y chromosomal sequences, confirming the enrichment of the sorted DNA. From the remaining characterized clones, we describe the development of 15 male-specific PCR assays and 19 male-female PCR assays potentially originating from the pseudoautosomal region or other areas of X-Y or autosome-Y homology.

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