II.19 Evidence for two independent high lysine genes in barley.
S. R. Muench, A. J. Lejeune, R. A. Nilan, and A. Kleinhofs. Department of Agronomy and Soils, Washington State University, Pullman, Washington 99163, USA.
The discovery of Hiproly, CI3947, by Munck, Karlsson, and Hagberg (6), in 1968 initiated considerable research to increase the lysine content in the protein of commercial barley varieties. Genetic aspects of this research led to the conclusion (7) that the Hiproly lysine content was controlled by a single recessive gene.
In 1970, Ingverson, Køie, and Doll (4) identified several high lysine mutants. One of these mutants, 1508, was of particular interest since it resulted in a 45% increase in lysine content with only a 10% decrease in yield compared to its parent variety Bomi. Genetic studies conducted by Doll (3) proved that the high lysine content in the protein of Mutant 1508 was due to a single recessive gene.
For future genetic and plant breeding studies it is important to know if the two genes in CI3947 and Mutant 1508 are allelic or non-allelic. The literature reveals that the action of the high lysine gene on protein synthesis appears not to be the same in CI3927 and Mutant 1508. Munck (5) reported that the major effect of the high lysine gene in CI3947 was associated with increased synthesis of the lysine rich albumin/globulin fraction. He also reported that the albumin fraction contained a higher proportion of lysine than is usually found in normal varieties. On the other hand, Ingverson et al. (4) found that in Mutant 1508 there was a 65% increase in the albumin/globulin fraction with a concomitant decrease in the lysine poor prolamin fraction. The proportion of lysine in the albumin/globulin fraction remained unchanged, while the glutelin and especially the prolamin fractions were much richer in lysine.
This report provides genetic evidence that the two genes for increased lysine content in barley are independent and non-allelic.
Reciprocal crosses were made between Hiproly and Mutant 1508. No seeds were obtained when Hiproly was used as the female parent. Therefore, all results presented here are from the cross Mutant 1508/CI3947. Approximately 20 F1 seeds were obtained from crossing two different Mutant 1508 heads. Six of these were planted in the greenhouse to produce F2 seed (seed from F1 plants). This F2 seed was space planted in the field. A total of 210 single F2 plants were individually harvested and threshed. Bulked F1 and F2 seed, and seed from 20 individual F2 plants, selected at random, were analyzed for protein and lysine content. All protein determinations were performed in duplicate by the Kjeldahl technique (N x 6.25) (1). Lysine determinations were made on a Technicon TSM automatic amino acid analyzer. Duplicate analyses were performed on the F1 and F2 seed. Single analyses were performed on the F3 seed.
Acrylamide electrophoresis was conducted according to Davis (2).
CI3947 carries the gene markers hulless and two-row. In addition, it has short rachilla hairs. Mutant 1508 is hulled, two-row and has long rachilla hairs. The two lines also differ in two prominent esterase bands. The progeny of the cross showed segregation for hulled vs. hulless seeds and for long vs. short rachilla hairs. They were homozygous for two-row. Ten individual F2 seedlings were examined for their esterase banding patterns. Both parental and heterozygous patterns were observed. No extraneous esterase bands were seen. These data clearly indicate that the observed progeny were from crossed and not from selfed seed.
The bulked F1 seed showed values of 2.81 and 2.98 g lysine/100 g protein in analysis of two different seed batches. Similar analysis of the bulked F2 seed gave the values 4.39 and 4.66 g lysine/100 g protein. The lysine values of F3 seed ranged from 2.78 to 4.75 g lysine/100 g protein. The data will be published elsewhere.
In analyzing the data we considered two alternatives, i.e., (1) that the two genes are allelic and (2) that they are independent. If the two genes are allelic, all the F1, F2,and F3 seed would be high in lysine content. The variability would not be expected to be greater than that between the two parents. If the two genes are independent, the F1 seed would be low in lysine content. The F2 seed, due to segregation, would be a mixture of high and low seed with a calculated average of 4.3 g lysine/100 g protein. The F3 seed would show segregation for low and high lysine values. The values should range from low values of normal barley to high values greater than either parent.
The results fit all criteria for two independent genes except that no F3 lines higher in lysine than both parents were found. This, however, is not unusual since only 20 F3 lines were analyzed and only one out of 16 would be expected to be homozygous for both high lysine genes. Thus, the double homozygote was probably missed.
References:
1. American Association of Cereal Chemists, Cereal laboratory methods, The Association, St. Paul, Minnesota 7 (1962).
2. Davis, B. J. 1964. Disc Electrophoresis II. Method and application to human serum proteins. Ann. N.Y. Acad. Sci. 121: 404-427.
3. Doll, H. 1973. Inheritance of the high-lysine character of a barley mutant. Hereditas 74: 293-294.
4. Ingverson, J., B. Køie, and H. Doll, 1973. Induced seed protein mutant of barley. Experientia 29: 1151-1152.
5. Munck, L. 1972. Barley Seed Proteins. In "Symposium: Seed Proteins" (G. E. Inglett, ed.), pp. 144-164. Avi Pub. Co., Inc., Westport, Conneticut.
6. Munck, L., K. E. Karlsson, and A. Hagberg, 1971. Selection and characterization of a high protein, high lysine variety from the World Barley Collection. In "Barley Genetics II, Second International Barley Genetics Symposium", Washington State University Press, Pullman, Washington.
7. Munck, L., K. E. Karlsson, A. Hagberg, and B. 0. Effam, 1970. Gene for improved nutritional value in barley seed protein. Science 168: 985-987.