Doctoral Dissertations

Date of Award

8-2001

Degree Type

Dissertation

Degree Name

Doctor of Philosophy

Major

Biomedical Sciences

Major Professor

Dabney K. Johnson

Committee Members

Edward J. Michaud III, Rebecca A. Prosser, Gary A. Sega

Abstract

The psrt mutations, 723SJ and 1060SJ, are END-induced, non-complementing mutations that produce identical phenotypes and map to the same p-deletion interval on mouse chromosome 7. psrt stand for profound seizure and runting; animals exhibiting this phenotype are runted and have severe seizures that are first detected at seven to ten days of age. Homozygous and hemizygous mutants typically live 15 to 18 days. A positional cloning strategy was employed to identify the gene responsible for this phenotype. The region of the genome containing psrt was better defined by mapping molecular markers and determining the breakpoints of p3RD3ooH, a p-deletion that does not complement the phenotype. A physical map was constructed in the minimal deletion interval determined to contain the mutation, and a Bacterial Artificial Chromosome from the physical map was sequenced to identify candidate genes. Three genes were identified, two from sequence analysis (Tat-interacting protein (30 kDa), Tip30, and protein arginine N-methyltransferase 3, Prmt3) and one from a search of the genome sequencing databases from the human region of homology (glycine transporter type 2, GLYT2). No mutations were detected in the Tip30 cDNA and G/yt2 was not molecularly characterized. However, the 5' end of the 723SJ Prmt3 transcript could not be amplified, indicating a possible chromosomal rearrangement or deletion, and mutation screening by temperature-gradient capillary electrophoresis was utilized to detect mismatches between the control and both the 723SJ and 1060SJ cDNAs.

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