Development of Marker-free TaGlu-Ax1 Transgenic Rice Harboring a Wheat High-molecular-weight Glutenin Subunit (HMW-GS) Protein

Abstract

High-molecular-weight glutenin subunits (HMW-GSs) are extremely important determinants of the functional properties of wheat dough. Transgenic rice plants containing a wheat TaGlu-Ax1 gene encoding a HMG-GS were produced from the Korean wheat cultivar 'Jokyeong' and used to enhance the bread-making quality of rice dough using the Agrobacterium-mediated co-transformation method. Two expression cassettes with separate DNA fragments containing only TaGlu-Ax1 and hygromycin phosphotransferase II (HPTII) resistance genes were introduced separately into the Agrobacterium tumefaciens EHA105 strain for co-infection. Rice calli were infected with each EHA105 strain harboring TaGlu-Ax1 or HPTII at a 3:1 ratio of TaGlu-Ax1 and HPTII. Among 210 hygromycin-resistant T0 plants, 20 transgenic lines harboring both the TaGlu-Ax1 and HPTII genes in the rice genome were obtained. The integration of the TaGlu-Ax1 gene into the rice genome was reconfirmed by Southern blot analysis. The transcripts and proteins of the wheat TaGlu-Ax1 were stably expressed in rice T1 seeds. Finally, the marker-free plants harboring only the TaGlu-Ax1 gene were successfully screened in the T1 generation. There were no morphological differences between the wild-type and marker-free transgenic plants. The quality of only one HMW-GS (TaGlu-Ax1) was unsuitable for bread making using transgenic rice dough. Greater numbers and combinations of HMW and LMW-GSs and gliadins of wheat are required to further improve the processing qualities of rice dough. TaGlu-Ax1 marker-free transgenic plants could provide good materials to make transgenic rice with improved bread-making qualities.

Molecular characterization and expression of α-gliadin genes from wheat cultivar Dacke in Bg 250 rice variety

“…15 In the past years, glutenin genes (LMW &HMW) have been successfully expressed in E. coli 16 as well as transferred to different wheat [17][18][19][20] and rice cultivars. [21][22][23][24][25][26] Gliadin has been expressed in E. coli. 27 However, reports on the transgenic expression of gliadin in plants are limited.…”

Overexpressing wheat low-molecular-weight glutenin subunits in rice (Oryza sativa L. japonica cv. Koami) seeds

“…Moreover, the bread-making qualities of rice were improved by adding wheat gluten to rice dough (Sivaramakrishnan et al 2004;Shin 2009). In addition, many molecular breeding studies have been undertaken to improve the bread-making qualities of rice through genetic engineering, including the production of soybean β-conglycinin subunits in rice seeds to improve nutritional and processing properties (Motoyama et al 2009) and the expression of wheat HMW-GSs such as 1Ax1, 1Bx7, 1Dx5, and 1Dy10 in rice seeds to improve dough-making qualities (Jeong et al 2016;Park et al 2014;Oszvald et al 2007Oszvald et al , 2013Jo et al 2017). However, improving the poor suitability of rice flour for various food products remains an important challenge in countries where rice is used as a staple crop.…”

Cellular Localization of Wheat High Molecular Weight Glutenin Subunits in Transgenic Rice Grain

“…In addition, the over-expression of wheat 1Dx5 and/or 1Dy10 HMW-GSs improves the extensibility and elasticity of dough from wheat [ 26 , 27 , 28 ] and rye [ 29 ]. In efforts that were aimed at improving rice dough quality, preliminary attempts have been made to express wheat SSPs, including 1Ax1 [ 30 ], 1Bx7 [ 31 ], 1Dx5 [ 32 , 33 ], and 1Dy10 [ 34 ] in rice seeds, although to date, only 1Dx5 has been shown to confer a positive effect on rice dough properties. The objective of this study was to engineer wheat HMW-GS into rice seeds.…”