Chinese Academy of Agricultural Sciences

Construction of genic male sterile system marked by yellow-green seedlings
--Zhou, H; Li, J; Deng, Y; Sun , R; Jiang, W

Nuclear male sterile gene ms2 and a yellow-green seedling gene (virescent) were introduced into an elite maize inbred line. Then a genic male sterile system marked by yellow-green seedling was constructed. The problem of segregation of genic male sterility was overcome by pulling out the normal green seedlings and leaving all the yellow-green seedlings (virescent) at three-four leaf stage. This new way of utilizing nuclear male sterility of maize is reported for the first time in the World and would find widespread use in developing countries.

Heterosis is usually described in terms of superiority of F1 performance over some measures of its parents. It's one of the modern achievements to utilize heterosis to produce more agricultural products. Artificial detasseling is often used in production of hybrid seeds, but it's more effective to utilize male sterile lines. Cytoplasmic male sterility is destroyed by specific diseases, e.g. Southern corn leaf blight. Maize nuclear male sterile genes were first found in 1930. Now more than 20 genes have been found, and most of them are recessive. They segregate in progenies and have no maintainers, so it is difficult to use them in breeding. Patterson (Proc. 6th Meeting Maize and Sorghum Section, Eucarpia, 1973) developed a duplication-deficiency system, but the seeds of the maintainer were small. It's difficult to use it in production.

From 1982, Jingxiong Li (pp. 4-5 in Recent Advances in Maize Breeding, China Science Press, 1992) used the linkage of ms1 and a white endosperm gene y in chromosome 6 to make backcrosses with some inbreds in order to select ms1 y/Ms1 Y linkage in China. The white seeds were male sterile and the yellow seeds were male fertile. Seeds produced can be separated into white and yellow seeds, i.e. male sterile and fertile seeds, by eyes or by machines. But the linkage between ms1 and y is not very close. They have a 5.72 cM recombination value. That means a seed producer must pull out 5-6% fertile plant tassels in the early flowering stage. This also makes it difficult to be used in seed production.

We had been thinking of the problem. We found that the ms2 gene is very close to a yellow-green seedling gene virescent (v) in chromosome 9. So we designed a genic male sterile system marked by yellow-green seedlings in order to find a new way to utilize genic male sterility. This study is to construct the system.

The virescent gene (v) was sent from the maize stock center in Illinois in 1992. The phenotype of this gene is a yellow-green seedling in the 3-4 leaf stage, which then becomes a normal green seedling. Nine plants containing the ms2 gene were pollinated with v pollen in 1993. The hybrid seeds were planted and selfed and 10,000 of their progenies were planted. Normal green seedlings (V gene) were pulled out and all of the remaining seedlings were yellow-green (v gene). When most of the plants shed pollen, we looked for male sterile plants (v v ms ms). All the male sterile plants were pollinated with elite maize inbreds. We harvested the ears, planted 3/4 of the seeds and selfed. The selfed seeds and the remaining 1/4 seeds of the last generation were planted, and all of the normal green seedlings of the selfed seeds were pulled out. Male sterile plants were selected and pollinated with pollen of the remaining 1/4 plants. Their progenies segregate green-male sterile and yellow-male fertile seedlings 1:1.

Nuclear male sterile gene ms2 and the yellow-green seedling gene (virescent) were introduced into an elite maize inbred line after 5 generations of genetic manipulation and selection. The genic male sterility system marked by yellow-green seedlings was developed by us. The virescent gene has little effect on plant growth because it is yellow-green not white and becomes normal green after 3 leaves. The system employed seedling colour to distinguish male sterile and fertile plants, pulling out male fertile seedlings in one row and pulling out male sterile seedlings in another row. The male fertile row is the maintainer. The harvested seeds in sterile rows will be grown and manipulated in the same way next year. So, the system resolved the problem of maintainer in nuclear male sterility. To make hybrid seeds, the farmer can pull out green seedlings in the 4-5 female rows, the male row can be any normal inbred (Ms Ms V V), because they can all restore the male sterility. The hybrid seeds (Ms ms V v) will grow normally, with no yellow-green male sterile plants. The farmer can grow them like other normal seeds. But the system needs the seed company to increase by 100% female seeds for pulling out. This means increasing the cost of hybrid seeds. But farmers have the custom of planting 3 times more seeds than should be planted and pulling out some weaker seedlings after 4-6 leaves, so this problem is not a problem in China. This system can be used in China or some developing countries. The design and manipulation is first reported by us. We think this is a new way of utilizing genic male sterility of maize. 

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