[Show abstract][Hide abstract] ABSTRACT: Agrobacterium-mediated plant transformation is an extremely complex and evolved process involving genetic determinants of both the bacteria and the host plant cells. However, the mechanism of the determinants remains obscure, especially in some cereal crops such as wheat, which is recalcitrant for Agrobacterium-mediated transformation. In this study, differentially expressed genes (DEGs) and differentially expressed proteins (DEPs) were analyzed in wheat callus cells co-cultured with Agrobacterium by using RNA sequencing (RNA-seq) and two-dimensional electrophoresis (2-DE) in conjunction with mass spectrometry (MS). A set of 4,889 DEGs and 90 DEPs were identified, respectively. Most of them are related to metabolism, chromatin assembly or disassembly and immune defense. After comparative analysis, 24 of the 90 DEPs were detected in RNA-seq and proteomics datasets simultaneously. In addition, real-time RT-PCR experiments were performed to check the differential expression of the 24 genes, and the results were consistent with the RNA-seq data. According to gene ontology (GO) analysis, we found that a big part of these differentially expressed genes were related to the process of stress or immunity response. Several putative determinants and candidate effectors responsive to Agrobacterium mediated transformation of wheat cells were discussed. We speculate that some of these genes are possibly related to Agrobacterium infection. Our results will help to understand the interaction between Agrobacterium and host cells, and may facilitate developing efficient transformation strategies in cereal crops.
PLoS ONE 11/2013; 8(11):e79390. DOI:10.1371/journal.pone.0079390 · 3.23 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Dwarfing germplasms are crucial to the development of wheat (Triticum aestivum L.) varieties with ideal plant height and combinations of agronomic traits. A novel dwarfing wheat line, AS34, was developed in this study by somatic variation. This new wheat germplasm was confirmed by sodium dodecyl sulfate polyacrylamide gel electrophoresis, two-dimensional gel electrophoresis, genomic in situ hybridization, and genomic scanning with 44 simple sequence repeat (SSR) markers, and our analysis showed that AS34 was clearly derived from the somatic variation of Yumai66 (YM66), its tissue culture parent, but not from natural hybridization. Molecular tests using known primers specific to Rht8 and Rht-B1b genes indicated that AS34 and its parent contained Rht8 but not Rht-B1b in their genomes, which was not consistent with previous reports. Similar to YM66, AS34 was sensitive to gibberellic acid, and this characteristic might be induced only by Rht8 or by Rht8 and the new dwarfing genes simultaneously. Genetic analysis revealed that the new dwarfing trait in AS34 was controlled by multiple quantitative genes, and data from the F 2 population yielded no key quantitative trait locus. According to the results of genomic scanning for different SSR markers, some of the new dwarfing genes in AS34 might be located on chromosome 4B. The new dwarfing material had positive effects on the agronomic traits related to grain yield in F-2 populations, particularly the variation range in plant height. This dwarfing resource will be potentially used in wheat breeding for the development of new wheat varieties with available agronomic traits.
[Show abstract][Hide abstract] ABSTRACT: Key message:
Seven kinds of transgenic tobacco plants transformed with combinations of three FBE genes were obtained. The transgenic plants transformed with Ta1-SST + Ta6-SFT genes appeared to have the highest fructan or soluble sugar content and the strongest salt tolerance. Fructan is thought to be one of the important regulators involved in plant tolerance to various abiotic stresses. In this study, wheat-derived genes, Ta1-SST, Ta6-SFT, and Ta1-FFT, encoding fructan biosynthesis enzymes (FBE) were isolated and cloned into vectors modified pBI121 or pZP211. Seven different combinations of the three target genes were transformed into tobacco plants through an Agrobacterium-mediated approach, and transgenic tobacco plants were identified by PCR, ELISA, and Southern blotting. Compared with tobacco plants transformed with other six combinations of the three target genes and with wild-type plants, the transgenic plants transformed with Ta1-SST + Ta6-SFT genes contained the highest fructan and soluble sugar content. All seven types of transgenic tobacco plants displayed a much higher level of tolerance to drought, low temperature, and high salinity compared with the wild type. Differences of drought and low temperature tolerance between the transgenic plants containing a single FBE gene and those harboring two or three FBE genes were not significant, but the salt tolerance level of the transgenic plants with different FBE gene combinations from high to low was: Ta1-SST + Ta6-SFT > Ta1-SST + Ta6-SFT + Ta1-FFT > Ta1-SST + Ta1-FFT > Ta1-SFT + Ta1-FFT > single FBE gene. These results indicated that the tolerances of the transgenic tobacco plants to various abiotic stresses were associated with the transformed target gene combinations and the contents of fructan and soluble sugar contained in the transgenic plants.
[Show abstract][Hide abstract] ABSTRACT: With 4 figures and 2 tablesAbstractThe bifunctional α-amylase/subtilisin inhibitor (BASI) encoded by the Isa gene on barley chromosome 2HL could inhibit wheat α-amylase activity, thereby reducing sprouting and improving the quality of wheat. Regenerated plants were derived from immature embryo culture of hybrids of common wheat varieties ‘Zhengmai 9023’, ‘CB037’ and ‘Zhongmai 16’ with the wheat–barley 2H alien substitution lines 2H(2A) and 2H(2B) after callus induction, subculture and differentiation. The presence of barley 2H chromatin was detected in regenerated plants (SC1) and their selfed progeny (SC2–5) using homoeologous group 2 SSR markers from barley and further identified in selected SC4 and SC5 lines using genomic in situ hybridization with barley genomic DNA as probe and CS genomic DNA for blocking. The Isa gene from the identified SC4 and SC5 lines was also amplified using Isa-specific primers. We identified wheat–barley 2H chromosome translocation lines carring the Isa gene.
[Show abstract][Hide abstract] ABSTRACT: Barley yellow dwarf virus (BYDV) may cause a serious disease affecting wheat worldwide. True resistance to BYDV is not naturally found in wheat. BYDV resistance genes are found in more than 10 wild relative species belonging to the genera of Thinopyrum, Agropyron, Elymus, Leymus, Roegneria, and Psathyrostachy. Through wide crosses combining with cell culture, use of ph mutants, or irradiation, 3 BYDV resistance genes in Th. intermedium, including Bdv2, Bdv3 and Bdv4, were introgressed into common wheat background. Various wheat-Th. intermedium addition and substitution, translocation lines with BYDV-resistance were developed and characterized, such as 7D-7Ai#1 (bearing Bdv2), 7B-7Ai#1, 7D-7E (bearing Bdv3), and 2D-2Ai-2 (bearing Bdv4) translocations. Three wheat varieties with BYDV resistance from Th. intermedium were developed and released in Australia and China, respectively. In addition, wheat-Agropyron cristatum translocation lines, wheat-Ag. pulcherrimum addition and substitution lines, and a wheat-Leymus multicaulis addition line (line24) with different resistance genes were developed. Cytological analysis, morphological markers, biochemical markers, and molecular markers associated with the alien chromatin carrying BYDV resistance genes were identified and applied to determine the presence of alien, chromosomes or segments, size of alien chromosome segments, and compositions of the alien chromosomes. Furthermore, some resistance-related genes, such as RGA, P450, HSP70, protein kinases, centrin, and transducin, were identified, which expressed specifically in the resistance translocation lines with Bdv2. These studies lay the foundations for developing resistant wheat cultivars and unraveling the resistance mechanism against BYDV.
Journal of Genetics and Genomics 09/2009; 36(9):567-73. DOI:10.1016/S1673-8527(08)60148-4 · 3.59 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The wheat line H960642 is a homozygous wheat-Thinopyrum intermedium translocation line with resistance to BYDV by genomicin situ hybridization (GISH) and RFLP analysis. The genomic DNA ofTh. intermedium was used as a probe, and common wheat genomic DNA as a blocking in GISH experiment. The results showed that the chromosome
segments ofTh. intermedium were transferred to the distal end of a pair of wheat chromosomes. RFLP analysis indicated that the translocation line H960642
is a T7DS-7DL-7XL translocation by using 8 probes mapped on the homoeologous group 7 in wheat. The translocation breakpoint
is located between Xpsr680 and Xpsr965 about 90–99 cM from the centromere. The RFLP markers psr680 and psr687 were closely
linked with the BYDV resistance gene. The gene is located on the distal end of 7XL around Xpsr680 and Xpsr687.
Science in China Series C Life Sciences 12/1999; 42(6):663-668. DOI:10.1007/BF02881585 · 1.61 Impact Factor