Determination of Rates and Patterns of Recombination at the Maize Red Color (r1) Locus

Abstract

DETERMINATION OF RATES AND PATTERNS OF RECOMBINATION AT THE MAIZE RED COLOR (R 1) LOCUS By William R. Dietrich, B .A. A thesis submitted in partial fulfil lment of the requirements for the degree of Master of Science at Virginia Commonwealth University Virginia Commonwealth University, 1998 Director: Dr. William B. Eggleston, Ph.D. Assistant Professor Department of Bio logy Homologous recombination has been studied in many p lant and animal systems. In maize, most recombination occurs intragenically. The current study assessed the frequency and location of meiotic recombination at the maize red color (r1) gene. Three independent mutant (colorless seed) alleles derived from the colored seed al lele, R-sc: 124, were made heterozygous with the colored plant al lele, r-r:n 142. Each mutant resulted from the insertion of a Dissociation (Ds) transposable element into the 3' end of R-sc: 1 24. In the presence of Activator (Ac), each Ds insertion al lele (r-sc:mutables) produced spotted seeds and germinal reversion to ful ly colored seeds due to the vi excision of Ds. In the absence of Ac, each insertion al lele stable and produces vii colorless or very faintly pigmented seeds. The r-sc:mutablelr-r:n 142 heterozygotes were pol l inated with R-g:Bpale, an r1 allele that produces pale brown seeds, in the absence of Ac, to recover revertant progeny l ikely resulting from recombination rather than Os excision. Revertant progeny were self­ pol l inated to verify paternity. Molecular analysis using polymerase chain reaction, Southern blot and DNA sequence analyses showed that 92 out of 94 of the revertant al leles arose via a cross over event between the r-sc:mutable and r-r:n 142 chromosomes. The remaining two revertant al leles l ikely arose via gene conversion, double cross over or cryptic Ac activity. The ratio of genetic to physica l distance ( 1 /p) calculated for the 3' end of r1 is approximately 0 .07 cM/kb, which is comparable to 1 / p values calculated for a 1 , b 1 , b z 1 and wx 1 loci. Given the average value for 1 /p throughout the genome is 0. 00021 cM/kb, this analysis supports the hypothesis that recombination in maize primari ly occurs i ntragenical ly and that r1 serves as a recombinat ion hot spot in the maize genome. Molecular analysis also revealed that the majority of exchanges occurred at the 3' end of r1 as has been previously observed. The pattern of recombination observed at r1 is different from those observed at other maize loci. The recovery of substantial ly more cross over events (92/94) relative to non-cross over events (2/94) or the nature of the recombination event is consistent with previous observations at a 1, b 1 and bz 1 . The frequency of potential gene conversion is estimated at 3. 97 x 1 o-s and the gene conversion viii tract length is maximally 2 .5 kb. The possibi l ity of the influence of insertional mutations, amount and structure of DNA sequence homology, cisand trans­ factors and preference for cross over versus non-cross over events could explain the observed pattern at r1 .