Genetic Analysis of Kafirins and Their Phenotypic Correlations with Feed Quality Traits, In Vitro Digestibility, and Seed Weight in Grain Sorghum

Cereal Chem. 78(4):412–416 Twenty-three entries of grain sorghum (Sorghum bicolor (L.) Moench), including eight inbred lines (five males and three females) and 15 hybrids, were evaluated to determine the proportion of γ, αII, and β-αIkafirins and their association with contents of crude protein, fat, and starch; protein digestibility; in vitro dry matter disappearance; and seed weight. The male lines included three normal-seeded lines (TX2737, TX435, and P954063) and two large-seeded lines (Eastin1 and PL-1). Female lines consisted of three common U.S. seed parent lines (Wheatland, Redlan, and SA3042). The lines and their hybrids were grown under dryland conditions at two locations in Kansas using a randomized complete block design. The effects of genotype, location, and males were significant for all kafirins. Wide variations in composition and general combining ability (GCA) for kafirin content were noted among parent lines and hybrids, with TX2737, Eastin1, and PL1 having the largest GCA values for γ (1.37), αII (1.99), and β-αI (2.57), respectively. Correlations among kafirins ranged from –0.89 to 0, whereas those of kafirins with feed quality traits, digestibility, and seed weight ranged from –0.45 to 0.48. Sorghum is the second most important feed grain in the United States cattle feedlot industry. Apart from being a major source of dietary energy, it is also a major source of dietary protein. The kafirins, storage proteins found in protein bodies, are the most abundant, making up between 70 and 80% of the total endosperm protein in sorghum (Watterson et al 1993; Hamaker et al 1995). Kafirins have been classified as α, β, and γ according to differences in molecular weight, solubility, and structure (Esen 1987; Shull et al 1991). Several studies have investigated the qualitative and quantitative differences and distribution of kafirins in the kernel and endosperm (Shull et al 1991, 1992; Watterson et al 1993; Bean et al 2001). Protein quality is associated with the quantity and composition of protein fractions in the grain, and variation in genotype may affect these parameters (Hamaker et al 1995). Moreover, digestibility is an important attribute of high quality protein. To the extent that kafirins constitute the bulk of total protein, genetic improvement of protein content and quality would require an understanding of the quantitative variation and distribution of kafirins with respect to genotype and their relationship with feed quality traits and digestibility. Recently, sorghum researchers in the United States and Australia have focused plant breeding efforts on developing largeseeded sorghum hybrids with improved yield potential and feed quality. In an analysis of genetically diverse sorghum lines and hybrids, Hicks et al (unpublished data) reported significant variation in seed size and combining ability for feed quality traits and in vitro digestibility. However, the biochemical basis for this variation is not well understood. Reports by Hamaker et al (1995) and Oria et al (2000) suggested that protein structure primarily determines variation in digestibility. However, variation in seed size and other grain characteristics also may affect kafirin composition and, thus, the relationships with feed quality traits and digestibility. Therefore, the objectives of this study were to 1) investigate genetic variation and combining ability for kafirin composition and 2) determine the associations of kafirin composition with major feed quality traits, in vitro digestibility, and seed weight in sorghum cultivars and hybrids. MATERIALS AND METHODS Hicks et al (unpublished data) evaluated 23 sorghum lines and hybrids for feed quality and agronomic characteristics. Given the cost associated with protein analysis, a subset of grain samples from these experiments was evaluated for kafirin composition. Five male and three female parent lines were intercrossed using a Design-II mating scheme to produce 15 hybrids (Comstock and Robinson 1952; Hallauer and Miranda 1988). The males used in this study included three normal-seeded lines (TX2737, TX435, and P954063) and two large-seeded lines (PL-1 and Eastin1). The females in this study were common U.S. seed parent lines (Wheatland, Redlan, and SA3042). The lines and their hybrids (23 entries) were planted and grown under dryland conditions at Kansas State University experiment fields in Ashland and Belleville, KS. The planting dates at the two locations were 27 and 28 May 1999, respectively. The experiment design for the original field trials consisted of a randomized complete block design with four replicates at each location. Entries were grown in single-row plots that were ≈6 m long and 0.76 m apart. Plots were thinned by hand to ensure uniform populations of 129,000 plants ha. Samples of grain from individual plots were cleaned, dried, and ground through a 1-mm screen in a Udy mill. Ground samples were evaluated for contents of crude protein (CP), crude fat (FAT), and crude starch (STA); protein digestibility (PD); and in vitro dry matter disappearance (IVDMD) as described by Hicks et al (unpublished data). In summary, CP was quantified based on total nitrogen measured using the nitrogen combustion method (CN-2000, Leco Corp., St. Joseph, MI). The CP in each sample was estimated from total nitrogen (N × 6.25) (Mosse 1990). The FAT was extracted and quantified using the ether extraction method 920.39 (AOAC 1990). The STA was evaluated similarly, using method 979.10. PD was quantified using a modification of the pepsin digestion procedure described by Mertz et al (1984). Ground flour samples were incubated in the buffered pepsin enzyme solution at 37°C for 2 hr in a shaking water bath. PD (g/kg) was calculated by subtracting undigested protein from total protein and was expressed as a proportion of total protein. The IVDMD was quantified using the procedure developed by Pedersen et al (2000). Ground samples were sealed in ANKOM F57 filter bags and were placed in vessels containing buffer solution and rumen 1 Contribution no. 01-53-J from the Kansas Agricultural Experiment Station. 2 Kansas State University, Department of Agronomy, Manhattan, KS 66502. 3 Kansas State University, Department of Grain Science, Manhattan, KS 66502. 4 USDA-ARS Grain Marketing Production and Research Center, Manhattan, KS 66502. Names are necessary to report factually on available data; however, the USDA neither guarantees nor warrants the standard of the product, and the use of the name by the USDA implies no approval of the product to the exclusion of others that may also be suitable. 5 USDA-REE-ARS-NPA, Wheat, Sorghum, and Forage Research, University of Nebraska, E. Campus, Lincoln, NE 68583. 6 Kansas State University Research and Extension Center-Hays, Hays,KS 67601. 7 Corresponding author. Phone: 785-532-7238. Fax: 785-532-6094. E-mail: mtuinstra@