Effect of Low Temperature Treatment on Agrobacterium-mediated Transformation of Maize Seed Buds

[Objectives]This study was conducted to improve the genetic transformation rate of maize. [Methods]The seeds of maize ＂ Zheng 58＂ as the experimental material were germinated and treated under freezing temperature of-18 ℃ for 30 and 35 min. The EGFP gene was transformed into the growth points of these seed buds by Agrobacterium tumefaciens EHA105. The transformation effect was determined by fluorescence protein detection on transformed buds. [Results]After a certain period of treatment at-18 ℃ following germination and transformation of maize bud growth points with A. tumefaciens,although the survival rate of maize buds was reduced,the percentage of transformed cells significantly increased. [Conclusions]Low temperature treatment can improve the transformation efficiency of A. tumefaciens to the growth point of maize bud.

RNA interference targeting ω-secalin genes differentially affects the processing quality in a wheat T1BL·1RS translocation line

Wheat-rye T1BL·1RS translocation lines are widely used,especially in China,but their processing quality is generally poor.An interfering expression vector targeting theω-secalin genes was constructed with the 1Bx7 seed-specific promoter.Biolistic-mediated genetic transformation of the wheat cultivar KN199 carrying the T1BL·1RS translocation generated 10 transgenic lines.Two representative transgenic lines,8-2 and 13-7,were selected for analysis.Compared with the control,the two transformants showed an up to 4.5-fold decrease in totalω-secalins and various levels of decrease inω-gliadins,γ-gliadins,and low-molecular-weight glutenins.A decrease in high molecular weight(HMW)glutenin 1Bx7 was detected only in 8-2,owing possibly to promoter methylation.Increased levels ofα-gliadins were observed in both transformants,but increased levels of HMW glutenins were observed only in 13-7.Line 13-7 showed increases in gluten index,Zeleny sedimentation value,stabilization time,and maximum resistance.Its bread volume was 849.6 mL,an 11.9%increase over that of the control.Line 8-2 showed decreases in these parameters,but its total cake-making quality score was 88,an 17.3%increase over that of the control.The study demonstrates that the same RNAi construct may produce different effects on wheat processing quality and highlights the influence of the vector promoter in RNA interference.

Effects of Acetylacetone Solution Soaking on Agrobacterium - transformed Maize Seed Buds

[Objectives]The effect of acetosyringone seed soaking on the transformation of maize seed buds was analyzed,so as to improve the genetic transformation efficiency of maize and to provide technical support for transgenic breeding of maize.[Methods]The seeds of the"Zheng 58"maize inbred line were used as experimental materials.When the seeds were germinated,AS was added to the water at concentrations of 70,140,210,and 280μmol/L,respectively,and the seeds germinated without the addition of AS served as the CK.The Agrobacterium-mediated method was used to transform bud growth points of maize seeds,and green fluorescent protein detection was performed on the young shoots transformed with EGFP(enhanced green fluorescent protein)gene.The effect of soaking seeds with acetosyringone solution on the transformation of maize bud growth points by Agrobacterium was studied according to the detection results.[Results]Soaking seeds in acetosyringone solutions for germination had the effect of inhibiting the germination of maize seeds and inhibiting sprout elongation,and the higher the concentration of acetosyringone,the stronger the inhibition.When the concentration of acetosyringone solution was 280μmol/L,the germination rate of seeds was only 36.2%of the CK,while soaking seeds with 70-140μmol/L acetosyringone solution for germination could not only ensure a higher germination rate of maize seeds,but also significantly increased the transformation efficiency of maize bud growth points.When the seeds were soaked with 70μmol/L acetosyringone solution for germination,the positive rate of transformed maize buds was the highest,reaching 32.1%.[Conclusions]When maize bud growth points were used as the receptor of Agrobacterium transformation,soaking seeds with 70-140μmol/L acetosyringone for germination basically did not affect the germination of seeds,and was beneficial to the activation of Agrobacterium,thereby promoting the transformation.

Study on Differentiation of Growing Points in Corn Sprouts into Multiple Stems

[Objectives] This study was conducted to improve the efficiency of genetic transformation using growing points of corn sprouts. [Methods] A mixed liquid of plant hormones 6-BA and KT at a suitable concentration and ratio was added dropwise to the growing points of corn sprouts, followed by culture under appropriate conditions. [Results] Corn sprouts could be induced to form multiple plants, thereby improving the transformation efficiency of growing points in corn sprouts. [Conclusions] This study provides a breeding method using growing points of corn sprouts as transformation receptors for direct plantlet formation without tissue culture by the Agrobacterium transgenic technology.