RFLP linkage analysis of rough sheath-1 on chromosome 7S

--Shawndra D. Martinez, Philip W. Becraft and Michael Freeling

Rough sheath-1 (Rs1) was recently mapped courtesy of Pioneer Hi-Bred, Inc. to the short arm of chromosome 7 using recombinant inbreds with a Rs1 genomic clone. To further define the location of Rs1 on chromosome 7S, we undertook experiments to map Rs1 using RFLPs.

The experiments were performed with a population segregating 1:1 for Rs1-O. Fifty-six progeny of a cross between an A643 inbred female and a Rs1-0/+ heterozygote in a B73 inbred background (backcrossed 4X) were scored phenotypically for Rs1-O. DNA was extracted from each individual and cut with either EcoRI, BamHI, or HindIII enzyme. The gels were blotted and probed with PIO200581B, NPI400 (both courtesy of Pioneer Hi-Bred, Inc.) and opaque-2 (o2) (courtesy of Bob Schmidt, B). The established order of these loci is PIO200581B, NPI400, o2 proceeding from the telomere toward the centromere on chromosome 7S. The blots were also probed with a Rs1 probe which verified that none of the plants were misgenotyped based on phenotype. The results of this analysis follow:
 
Chromosome Segment Number of Individuals Number of Recombinants Map Distance (cM ± s.e.)
1. o2 - NPI400 55 7 12.7 ± 4.5
2. NPI400 - PIO200581B 52 10 19.2 ± 5.5
3. PIO200581B - Rs1 53 3 5.7 ± 3.2

Only two double recombinants were obtained; one involving chromosome segments 1 and 2, the other segments 2 and 3.

These data show that Rs1 maps to the distal region of the short arm of chromosome 7, distal to PIO200581B. Therefore, Rs1 defines a new zero point on the chromosome 7 RFLP map. Our data put Rs1 approximately 37.6 map units distal to o2. Because the position of o2 on the genetic map is 16, it appears likely that Rs1 may also define a new zero position on the chromosome 7 genetic map. This position is now defined by hairy sheath (Hs). In previous attempts to map Rs1-O, Hs and o2 in a three-point cross, we were unable to score the Hs phenotype. Until this is accomplished we cannot determine the relative positions of Rs1 and Hs with certainty.

This part of chromosome 7S appears to display substantial variability in recombination frequency. In an earlier RFLP mapping experiment, 4 out of 24 individuals were recombinant between Rs1 and o2, for a recombination frequency of 16.7 +/- 6.7%. The reported positions of these RFLP loci in the compiled data tables (MNL 65:145-153, 1991) would also indicate a lower recombination frequency between PIO200581B, NPI400 and o2 than was obtained in the current experiment.


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