Locating of important agronomic genes onto chromosome is helpful for efficient development of new wheat varieties. Wheat chromosome substitution lines between two varieties have been widely used for locating genes bec...Locating of important agronomic genes onto chromosome is helpful for efficient development of new wheat varieties. Wheat chromosome substitution lines between two varieties have been widely used for locating genes because of their distinctive advantages in genetic analysis, compared with the aneuploid genetic materials. Apart from the substituted chromosome, the other chromosomes between the substitution lines and their recipient parent should be identical, which eases the gene locating practice. In this study, a set of chromosome substitution lines with cv. Wichita (WI) as the recipient parent and cv. Cheyenne (CNN) as the donor parent were studied for the composition of high molecular weight glutenin subunits (HMW-GS) as well as a range of agronomic important traits. Results revealed that the substitution lines of WI(CNN5D), WI(CNN6A) and WI(CNN7B) had higher plant heights than the two parents of WI and CNN, and WI(CNN3D) had later maturity than the parents. By sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) analysis, a substitution line WI(CNN5B) was found to contain different HMW-GS patterns from its two parents, in which 1 By9 was replaced by 1 By8 on chromosome 1BL. Simple sequence repeat (SSR) analysis confirmed that the variation on 1BL in WI(CNN5B) was originated from Chinese Spring (CS). It is concluded that chromosomal fragments from bridge material and donor parent were quite often retained in intracultivaral chromosome substitution lines except the substituting chromosomes.展开更多
The regeneration rate of wheat immature embryo varies among genotypes, howbeit many elite agriculture wheat varieties have low regeneration rates. Optimization of tissue culture conditions and attempts of adding signa...The regeneration rate of wheat immature embryo varies among genotypes, howbeit many elite agriculture wheat varieties have low regeneration rates. Optimization of tissue culture conditions and attempts of adding signal molecules are effective ways to increase plant regeneration rate. Inter-culture is one of ways that have not been investigated in plant tissue culture. Moreover, the use of arabinogalactan proteins (AGPs) and hydrogen peroxide (H202) have been reported to increase regeneration rate in a few plant species other than wheat. The current research pioneeringly uses inter-culture of immature embryos of different wheat genotypes, and also investigates impacts of AGP and H2O2 on the induction of embryogenic calli and plant regeneration. As a result, high-frequency regeneration wheat cultivars Kenong 199 (KN 199) and Xinchun 9 (XC9), together with low-frequency regeneration wheat line Chinese Spring (CS), presented striking increase in the induction of embryogenic calli and plant regeneration rate of CS through inter-culture strategy, up to 52.19 and 67.98%, respectively. Adding 50 to 200 mg L-1 AGP or 0.005 to 0.01‰ H2O2 to the callus induction medium, enhanced growth of embryogenic calli and plant regeneration rate in quite a few wheat genotypes. At 50 mg L-1 AGP application level in callus induction medium plant regeneration rates of 8.49,409.06 and 283.16% were achieved for Jimai 22 (JM22), Jingdong 18 (JD18) and Yangmai 18 (YM18), respectively; whereas at 100 mg L-1 AGP level, CS (105.44%), Chuannong 16 (CN16) (80.60%) and Ningchun 4 (NC4) (62.87%) acted the best. Moreover CS (79.05%), JM22 (7.55%), CN16 (101.87%), YM18 (365.56%), Yangmai 20 (YM20) (10.48%), and CB301 (187.40%) were more responsive to 0.005 %o of H2O2, and NC4 (35.37%) obtained the highest shoot regeneration rates at 0.01%o of H2O2. Overall, these two methods, inter-culture and AGP (or H2O2) application, can be further applied to wheat transgenic research.展开更多
Obtaining transgenic plants is a common method for analyzing gene function. Unfortunately, stable genetic transformation is difficult to achieve, especially for plants(e.g., soybean), which are recalcitrant to genet...Obtaining transgenic plants is a common method for analyzing gene function. Unfortunately, stable genetic transformation is difficult to achieve, especially for plants(e.g., soybean), which are recalcitrant to genetic transformation. Transient expression systems, such as Arabidopsis protoplast, Nicotiana leaves, and onion bulb leaves are widely used for gene functional studies. A simple method for obtaining transgenic soybean callus tissues was reported recently. We extend this system with simplified culture conditions to gene functional studies, including promoter analysis, expression and subcellular localization of the target protein, and protein-protein interaction. We also evaluate the plasticity of this system with soybean varieties, different vector constructs, and various Agrobacterium strains. The results indicated that the callus transformation system is efficient and adaptable for gene functional investigation in soybean genotype-, vector-, and Agrobacterium strain-independent modes. We demonstrated an easy set-up and practical homologous strategy for soybean gene functional studies.展开更多
Soil-borne pathogen Phytophthora sojae is an oomycete that causes devastating damage to soybean yield. To mine original resistant genes in soybean is an effective and environmentally-friend approach controlling the di...Soil-borne pathogen Phytophthora sojae is an oomycete that causes devastating damage to soybean yield. To mine original resistant genes in soybean is an effective and environmentally-friend approach controlling the disease. In this study, soybean proteins were extracted from the first trifoliolates infected by predominant P. sojae race 1 and analyzed by twodimensional gel electrophoresis. Nineteen differently-expressed protein spots were detected, and 10 of them were further applied for Matrix-Assisted Laser Desorption/Ionization Time of Flight Mass Spectrometry Assay. One protein containing a dirigent (DIR) domain was identified and belonged to the DIR-b/d family. Therefore, it was named as GmDRR1 (Glycine max Disease Resistance Response 1). Then, GmDRR1 gene was pathologically confirmed to be involved in the resistant to P. sojae in soybean. GmDRR1-GFP (green fluorescent protein) fusion proteins localized in the cell membrane. qRTPCR results showed GmDRR1 gene expressed differently in P. sojae resistant- and susceptible-soybean cultivars. By the promoter analysis, we found a haplotype H8 was existing in most resistant soybean varieties, while a haplotype H77 was existing in most susceptible soybean varieties. The H77 haplotype had seven SNPs (C to A, G to C, C to A, T to A, T to C, T to C, and T to A) and two single nucleotide insertions. The results supported that the expression difference of GmDRR1 genes between P. sojae resistant- and susceptible-soybean cultivars might depend on the GmDRR1 promoter SNPs. The results suggested that GmDRR1 was a dirigent protein involved in soybean resistant to P. sojae and paved a novel way for investigation of the molecular regulatory mechanism of the defense response to P. sojae in soybean.展开更多
基金financially supported by grants in part from the National Natural Science Foundation of China (31371621 and 31271703)
文摘Locating of important agronomic genes onto chromosome is helpful for efficient development of new wheat varieties. Wheat chromosome substitution lines between two varieties have been widely used for locating genes because of their distinctive advantages in genetic analysis, compared with the aneuploid genetic materials. Apart from the substituted chromosome, the other chromosomes between the substitution lines and their recipient parent should be identical, which eases the gene locating practice. In this study, a set of chromosome substitution lines with cv. Wichita (WI) as the recipient parent and cv. Cheyenne (CNN) as the donor parent were studied for the composition of high molecular weight glutenin subunits (HMW-GS) as well as a range of agronomic important traits. Results revealed that the substitution lines of WI(CNN5D), WI(CNN6A) and WI(CNN7B) had higher plant heights than the two parents of WI and CNN, and WI(CNN3D) had later maturity than the parents. By sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) analysis, a substitution line WI(CNN5B) was found to contain different HMW-GS patterns from its two parents, in which 1 By9 was replaced by 1 By8 on chromosome 1BL. Simple sequence repeat (SSR) analysis confirmed that the variation on 1BL in WI(CNN5B) was originated from Chinese Spring (CS). It is concluded that chromosomal fragments from bridge material and donor parent were quite often retained in intracultivaral chromosome substitution lines except the substituting chromosomes.
基金financially supported in part by the National Key Project for Tansgenic Study, Ministry of Agriculture of China(2011ZX08010-004)
文摘The regeneration rate of wheat immature embryo varies among genotypes, howbeit many elite agriculture wheat varieties have low regeneration rates. Optimization of tissue culture conditions and attempts of adding signal molecules are effective ways to increase plant regeneration rate. Inter-culture is one of ways that have not been investigated in plant tissue culture. Moreover, the use of arabinogalactan proteins (AGPs) and hydrogen peroxide (H202) have been reported to increase regeneration rate in a few plant species other than wheat. The current research pioneeringly uses inter-culture of immature embryos of different wheat genotypes, and also investigates impacts of AGP and H2O2 on the induction of embryogenic calli and plant regeneration. As a result, high-frequency regeneration wheat cultivars Kenong 199 (KN 199) and Xinchun 9 (XC9), together with low-frequency regeneration wheat line Chinese Spring (CS), presented striking increase in the induction of embryogenic calli and plant regeneration rate of CS through inter-culture strategy, up to 52.19 and 67.98%, respectively. Adding 50 to 200 mg L-1 AGP or 0.005 to 0.01‰ H2O2 to the callus induction medium, enhanced growth of embryogenic calli and plant regeneration rate in quite a few wheat genotypes. At 50 mg L-1 AGP application level in callus induction medium plant regeneration rates of 8.49,409.06 and 283.16% were achieved for Jimai 22 (JM22), Jingdong 18 (JD18) and Yangmai 18 (YM18), respectively; whereas at 100 mg L-1 AGP level, CS (105.44%), Chuannong 16 (CN16) (80.60%) and Ningchun 4 (NC4) (62.87%) acted the best. Moreover CS (79.05%), JM22 (7.55%), CN16 (101.87%), YM18 (365.56%), Yangmai 20 (YM20) (10.48%), and CB301 (187.40%) were more responsive to 0.005 %o of H2O2, and NC4 (35.37%) obtained the highest shoot regeneration rates at 0.01%o of H2O2. Overall, these two methods, inter-culture and AGP (or H2O2) application, can be further applied to wheat transgenic research.
基金supported by the Transgenic Programs,China(2014ZX0800930B and 2016ZX08009-001)the National Natural Science Found of China(31371703)
文摘Obtaining transgenic plants is a common method for analyzing gene function. Unfortunately, stable genetic transformation is difficult to achieve, especially for plants(e.g., soybean), which are recalcitrant to genetic transformation. Transient expression systems, such as Arabidopsis protoplast, Nicotiana leaves, and onion bulb leaves are widely used for gene functional studies. A simple method for obtaining transgenic soybean callus tissues was reported recently. We extend this system with simplified culture conditions to gene functional studies, including promoter analysis, expression and subcellular localization of the target protein, and protein-protein interaction. We also evaluate the plasticity of this system with soybean varieties, different vector constructs, and various Agrobacterium strains. The results indicated that the callus transformation system is efficient and adaptable for gene functional investigation in soybean genotype-, vector-, and Agrobacterium strain-independent modes. We demonstrated an easy set-up and practical homologous strategy for soybean gene functional studies.
基金financially supported by the Academic Skeleton Support Plan of Department of Education of Heilongjiang Province,China (1254G011)the National Natural Science Foundation of China (31271747,31471516,31400074,31401465,31501332)+3 种基金the National High-Tech R&D Program of China (the 863 Program,2013AA102602)the Research Fund for the Doctoral Program of Higher Education of China (20122325120015)the Academic Backbone Project of Northeast Agricultural University,China (15XG02)the Talented Young Project of Northeast Agricultural University,China (518062)
文摘Soil-borne pathogen Phytophthora sojae is an oomycete that causes devastating damage to soybean yield. To mine original resistant genes in soybean is an effective and environmentally-friend approach controlling the disease. In this study, soybean proteins were extracted from the first trifoliolates infected by predominant P. sojae race 1 and analyzed by twodimensional gel electrophoresis. Nineteen differently-expressed protein spots were detected, and 10 of them were further applied for Matrix-Assisted Laser Desorption/Ionization Time of Flight Mass Spectrometry Assay. One protein containing a dirigent (DIR) domain was identified and belonged to the DIR-b/d family. Therefore, it was named as GmDRR1 (Glycine max Disease Resistance Response 1). Then, GmDRR1 gene was pathologically confirmed to be involved in the resistant to P. sojae in soybean. GmDRR1-GFP (green fluorescent protein) fusion proteins localized in the cell membrane. qRTPCR results showed GmDRR1 gene expressed differently in P. sojae resistant- and susceptible-soybean cultivars. By the promoter analysis, we found a haplotype H8 was existing in most resistant soybean varieties, while a haplotype H77 was existing in most susceptible soybean varieties. The H77 haplotype had seven SNPs (C to A, G to C, C to A, T to A, T to C, T to C, and T to A) and two single nucleotide insertions. The results supported that the expression difference of GmDRR1 genes between P. sojae resistant- and susceptible-soybean cultivars might depend on the GmDRR1 promoter SNPs. The results suggested that GmDRR1 was a dirigent protein involved in soybean resistant to P. sojae and paved a novel way for investigation of the molecular regulatory mechanism of the defense response to P. sojae in soybean.