III. 1. The use of male sterile stocks in linkage analysis.
T. E. Haus, Department of Agronomy, Colorado State University, Ft. Collins, Colorado 80523. U. S. A.
Male sterile stocks offer the unique advantage of producing large numbers of testcross progeny with relative ease. The approach method of crossing (Curtis and Croy, 1958) eliminates the laborious procedure of emasculation and pollination normally required in making barley crosses.
The procedure will be illustrated by using 3 linked gene loci on chromosome 4 in barley. Hooded vs awned (K/k), green vs. chlorina (F9/9) and male fertile vs. male sterile (Msg24/msg24) have all been shown to be on chromosome 4. f9 and k are on the long arm (Haus 1983) and msg24 has been reported to be near the centromere (Wolfe 1984) but arm location is not known.
Stocks are synthesized having recessive genes. In using male sterile genes it is convenient to produce stocks that segregate 1:1 (male fertile: male sterile). This can be accomplished by crossing a male-sterile plant with a male-fertile one. F1 plants are crossed as the male with male-sterile plants. The resulting seeds will segregate in the desired 1:1 ratio of male fertile to male-sterile plants. Perpetuation of such stocks can be done by simply intercrossing fertile and sterile plants grown from this seed.
Using stocks segregating 1:1 (male fertile:male sterile) and recessive for kk, f9f9, male-sterile plants are crossed as female plants to normal plants (Msg24Msg24 KK F9F9). F1 plants are used as male parents in crosses with male-sterile, awned, chlorina plants (msg24msg24 kk f9f9). Progeny segregate into 8 phenotypic classes according to recombination frequencies.
Among the testcross progeny are the two parental types: Msg24msg24 Kk F9f9 and msg24msg24 kk f9f9 which are recognizable by phenotype, i.e., male-fertile, hooded, green and male-sterile, awned, chlorina. These male sterile progeny crossed with the parental type male-fertile, hooded, green plants produce seeds which segregate as a repeat of the testcross so large numbers of progeny can be produced to provide a reasonable estimate of map distance as measured by recombination value.
The one limitation of this sytem is the inability to use it to map gene loci of seedling lethal mutants and some others such as rattail (rt). Since such mutants do not survive to flowering or produce floral organs testcrosses of them cannot be made.
The male-sterile stock used in this type of study does not need to be linked with the mutants being mapped. The advantage of linkage of the male sterile locus and other genes in the experiment is that it (the male sterile locus) is an added position which has no marked effect on viability.
References:
Curtis, B. C. and L. I. Croy. The approach method of making crosses in small grains. Agron. Jour. 50:49-51.
Haus, T. E. Coordinator's report. BGN 13:93-94. 1983.
Wolfe, R. I. Personal communication. 1984.