Several Key Factors for Isolating the Glutenin Subunits of Wheat

doi:10.7668/hbnxb.201751710

Abstract

Abstract: The composition of wheat high and low molecular weight glutenin subunits is an important factor affecting the processing quality of wheat. In order to quickly and economically identify the composition of wheat high and low molecular weight glutenin subunits, it is necessary to continuously optimize the methods of extracting and isolating these proteins. Based on the method of extracting and separating wheat high and low molecular weight glutenin subunits proposed by Singh et al., we conducted in-depth research on several key factors like isopropyl alcohol concentration, alkylating agent concentration and extraction steps used in monomeric protein removal, gluten reduction and alkylation. The results showed that there were no difference in the extraction effect of high molecular weight glutenin subunits between different concentrations of isopropanol treatments in the range of 10%-50% during the reduction of glutenin. When the low molecular weight glutenin subunit was extracted with low concentration of isopropanol, the effect was poor. With the increase of isopropanol concentration, the extraction effect gradually increased. When the concentration of isopropanol was increased to 30%, the extraction effect reached the highest, but when the concentration of isopropanol continued to increase to 50%, the effect was no longer improved. Using 30% and 50% isopropanol, the effect of removing the monomeric proteins was the same. When the alkylating agent was directly added to the sample buffer for alkylation, different concentrations(0.6%-1.4%)of alkane treatments had the same alkylation effects for the glutenin subunits, but when the concentration of alkylating agent was 1.4%, the electrophoresis background was heavier. The optimized method was to reduce the isopropanol concentration from 50% to 30% during monomer protein removal and gluten reduction, remove the separate alkylation step, and add 0.6% alkylating agent directly to the sample buffer for alkylation. The optimized method not only reduced the amount of reagents, but also simplified the extraction step and increased the strength of the electrophoresis strips.

Key words: Wheat, Glutenin subunit, SDS-PAGE, Optimization, Isolating method

CLC Number:

S512.1

LI Chunyan, ZHANG Cuimian, CHAI Jianfang, BI Caili, MA Xiuying, ZHAO He, Lü Mengyu, ZHOU Shuo, DONG Fushuang, LIU Yongwei, YANG Fan, WANG Haibo. Several Key Factors for Isolating the Glutenin Subunits of Wheat[J]. ACTA AGRICULTURAE BOREALI-SINICA, 2019, 34(4): 32-36. doi: 10.7668/hbnxb.201751710.

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References

[1] Luo C, GriffinW B, Brandlard G, McNei D L.Comparison of low-and high molecular-weight wheat glutenin allele effects on flour quality[J]. Theoretical and Applied Genetics, 2001, 102(6-7):1088-1098. doi:10.1007/s001220000433.
[2] Carrillo J M, Rousset M, Qualset C O, Kasarda D D. Use of recombinant inbred lines of wheat for study of associations of high-molecular-weight glutenin subunit alleles to quantitative traits. 1.
Gain yield and quality prediction tests[J]. Theoretical and Applied Genetics, 1990, 79(3):321-330. doi:10.1007/BF01186074.
[3] 赵和,卢少源,李宗智.小麦高分子量麦谷蛋白亚基遗传变异及其与品质和其它农艺性状关系的研究[J].作物学报,1994,20(1):67-75.
Zao H,Lu S Y,Li Z Z. Studies on inheritance and variation of HMW glutenin subunits and their correlation with quality and other agronomic characters in wheat[J].Agronomica Sinica, 1994,20(1):67-75.
[4] 刘会云,刘畅,王坤杨,杜丽璞,王轲,佘茂云,叶兴国. 小麦高分子量麦谷蛋白亚基鉴定及其品质效应研究进展[J]. 植物遗传资源学报,2016,17(4):701-709. doi:10.13430/j.cnki.jpgr.2016.04.015.
Lu H Y, Liu C, Wang K Y, Du L P, Wang K, Yu M Y, Ye X G. Review on the identification and bread-making quality of high molecular weight glutenin subunits in wheat[J]. Journal of Plant Genetic Resources, 2016,17(4):701-709.
[5] Gupta R B, Bekes F, Wrigley C W. Prediction of physical dough properties from glutenin subunit composition in bread wheats correlation studies[J]. Cereal Chemistry, 1991, 68(4):328-333.
[6] 马勇. 小麦低分子量麦谷蛋白亚基研究进展[J]. 农业科技通讯,2017(9):9-10.
M Y. Research progress of wheat low molecular weight glutenin subunits[J]. Agricultural Science and Technology Communication, 2017(9):9-10.
[7] 宋维富,杨雪峰,宋庆杰,张春利,辛文利,肖志敏. 低分子量麦谷蛋白亚基种类与强筋小麦品质关系[J]. 黑龙江农业科学,2018(2):133-135. doi:10.11942/j.issn1002-2767.2018.02.0133.
Sng W F, Yang X F, Song Q J, Zhang C L, Xin W L, Xiao Z M. Relationship between the low-molecular-weight glutenin subunits and quality of strong gluten wheat[J]. Heilongjiang Agricultural Sciences, 2018(2):133-135.
[8] Tanaka H, Shimizu R, Tsujimoto H. Genetical analysis of contribution of low-molecular-weight glutenin subunits to dough strength in common wheat(Triticum aestivum L.)[J]. Euphytica, 2005, 141(1-2):157-162. doi:10.1007/s10681-005-6714-6.
[9] 刘丽, 周阳, 何中虎, 王德森, 张艳, Pena R J. 高、低分子量麦谷蛋白亚基等位变异对小麦加工品质性状的影响[J]. 中国农业科学, 2004, 37(1):8-14.
Lu L, Zhou Y, He Z H, Wang D S, Zhang Y, Pena R J. Effect of allelic variation in HMW and LMW glutenin subunits on the processing quality in common wheat[J]. Scientia Agricultura Sinica, 2004, 37(1):8-14.
[10] Singh N K, Shepherd K W, Cornish G B. A simplified SDS-PAGE procedure for separating LMW subunits of glutenin[J]. Journal of Cereal Science, 1991, 14(3):203-208. doi:10.1016/s07ss-5210(09)80039-8.
[11] 孙辉,李保云,朱金宝,刘广田, 张树臻.小麦高、低分子量谷蛋白亚基单向一步分离法新探[J]. 中国农业大学学报,1998,3(S4):19-23.
Sn H, Li B Y, Zhu J B, Liu G T, Zhang S Z. A simple SDS-PAGE procedure for separation of high and low molecular weight glutenin subunits[J]. Journal of Chinese Agricultural University, 1998,3(S4):19-23.
[12] Fu B X, Kovacs M I P. Rapid single-step procedure for isolating total glutenin proteins of wheat flour[J]. Journal of Cereal Science , 1999, 29(2):113-116.doi:10.1006/jcrs.1998.0225.
[13] 纪军, 刘冬成, 王静,李俊明, 张爱民. 一种小麦高、低分子量麦谷蛋白亚基的提取方法[J]. 遗传,2008, 30(1):123-126. doi:10.3724/SP.J.1005.2008.00123.
J J, Liu D C, Wang J, Li J M, Zhang A M. A method of extraction and separation of wheat gluten[J]. Hereditas, 2008, 30(1):123-126.
[14] 覃建兵,任燕萍,祝长青. 小麦低分子量麦谷蛋白亚基分离条件优化[J]. 生物技术, 2010, 20(4):37-39. doi:10.3969/j.issn.1004-311X.2010.04.122.
Tn J B, Ren Y P, Zhu C Q. Separation of Low-Molecular-Weight glutenin subunits in wheat[J]. Biotechnology, 2010, 20(4):37-39.
[15] 吴芳, 董惠, 韩兆雪,潘志芬, 邓光兵, 余懋群. 中国小麦品种高分子量谷蛋白亚基和低分子量谷蛋白亚基组成分析[J]. 麦类作物学报,2006, 26(3):82-86.doi:10.7606/j.issn.1009-1041.2006.03.110.
W F, Dong H, Han Z X, Pan Z F, Deng G B, Yu M Q. Genetic diversity of HMW and LMW glutenin subunits in chinese hexaploid wheats[J]. Journal of Triticeae Crops, 2006, 26(3):82-86.
[16] 张平平,马鸿翔,姚金保, 周淼平, 张鹏. 高分子量谷蛋白单亚基缺失对软质小麦宁麦9号加工品质的影响[J].作物学报,2016,42(5):633-640. doi:10.3724/SP.J.1006.2016.00633.
Zang P P, Ma H X, Yao J B, Zhou M P, Zhang P. Effect of HMW-GS deletion on processing quality of soft wheat Ningmai 9[J]. Acta Agronomica Sinica, 2016,42(5):633-640.
[17] Ibba M I, Kiszonas A M, Guzmán C, Morris C F. Definition of the low molecular weight glutenin subunit gene family members in a set of standard bread wheat(Triticum aestivum L.)varieties[J], Journal of Cereal Science, 2017, 74:263-271. doi:10.1016/j.jcs.2017.02.015.
[18] Lee J Y, Beom H R, Altenbach S B, Lim S H, Kim Y T, Kang C S, Yoon U H, Gupta R, Kim S T, Ahn S N, Kim Y M. Comprehensive identification of LMW-GS genes and their protein products in a common wheat variety[J]. Functional & Integrative Genomics, 2016, 16(3):269-279. doi:10.1007/s10142-016-0482-3.
[19] Beom H R, Kim J S, Jang Y R, Lim S H, Kim C K, Lee C K, Lee J Y. Proteomic analysis of low-molecular-weight glutenin subunits and relationship with their genes in a common wheat variety[J]. 3 Biotech, 2018, 8(1):56. doi:10.1007/s13205-017-1081-2.
[20] Fu B X, Sapirstein H. Procedure for isolating monomeric proteins and polymeric glutenin of wheat flour[J]. Cereal Chemistry, 1996, 73(1):143-152.

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