The Selection of Transgenic Recipients from New Elite Wheat Cultivars and Study on Its Plant Regeneration System

In the protocol of wheat transformation, to use elite wheat cultivars as exogenous gene recipients can speed up the process of commercial field applications of transgenic wheat. However, it is necessary to screen wheat cultivars with good tissue culture response (TCR) continuously from plenty of elite wheat cultivars released for wheat transformation, and it is also important to find a plant regeneration system that is suitable for these cultivars. So, the TCR of mature and immature embryos of six wheat cultivars Chuannong 11 (CN11), Chuannong12 (CN12), Chuannong17 (CN17), Chuannong18 (CN18), Chuannong19 (CN19), and Chuannong21 (CN21), which possess superior agronomic traits, were investigated by using a good TCR wheat cultivar Bobwhite as control. The results indicated that only the immature and mature embryos of CN12, CN17, and CN18 exhibited good TCR compared with Bobwhite. No significant differences were observed between embryos of Bobwhite and of the three cultivars in TCR. Mature embryo-derived calli of CN12 were used as explants for transformation by particle bombardment of SAMDC gene. Seven transformants were obtained and the efficiency was 2.3%. This research supplies three new elite recipient cultivars for wheat transformation. The wheat plant regeneration system used in this research is different from those successful ones reported previously and it could be a reference for other wheat genotypes. Furthermore, Bobwhite and the three wheat cultivars were proved to be 1RS/1BL translocation, by methods of A-PAGE, C- banding, and genomic in situ hybridization (GISH). These results imply that probably there is some relationship between 1RS/1BL translocation and TCR of wheat embryos. So this research gives us a hint that we should pay more attention to the 1RS/1BL translocations when we screen the wheat cultivars with good TCR and also that the mechanism of the effect of 1RS/ 1BL translocation on TCR is worthy of being investigated.

Alteration of Terminal Heterochromatin and Chromosome Rearrangements in Derivatives of Wheat-Rye Hybrids

小麦黑麦增加和替换线和他们的自我子孙在 telomeric heterochromatin 和着丝点揭示了变化。而且，一个有丝分裂地不稳定的 dicentric 染色体和稳定的 multicentric 染色体在一根中国春天帝国的黑麦 3R 增加线的子孙被观察。一个不稳定的 multicentric 染色体在一根 6R/6D 替换线的子孙被发现。包括终端 heterochromatin 的运动和消失的终端 heterochromatin 的激烈的变化发生在小麦黑麦增加线 3R，和 5RS ditelosomic 增加线的子孙。高度稳定的 minichromosomes 在一根 monosomic 4R 增加线，一根 ditelosomic 5RS 增加线和一根 6R/6D 替换线的子孙被观察。有或没有为 telomeric DNA (TTTAGGG ) n 的 FISH 信号， Minichromosomes 源于一根 monosomic 4R 增加线对下一代稳定、能递送。结果显示着丝点和终端 heterochromatin 能深刻地在小麦黑麦混血儿衍生物被改变。

Identification and Physical Mapping of New PCR-Based Markers Specific for the Long Arm of Rye(Secale cereale L.) Chromosome 6

To effectively use elite genes on the long arm of rye chromosome 6(the 6RL arm) in wheat breeding programs,precise and fast identification of 6RL chromatin in wheat backgrounds is necessary.PCR-based 6RL-specific markers can facilitate the detection of elite genes on 6RL in wheat breeding.However,only a limited number of 6RL-specific markers have been developed.In the present study.300 new PCR-based 6RL-specific markers were identified using specific length amplified fragment sequencing(SLAF-seq) technology,and were further physically mapped to four regions on the 6RL arm using 6R and 6RL deletion lines.Interestingly,127 of the 300 markers were physically localized to a region from the site between 2.3 and 2.5 to the telomere,the same region where the powdery mildew resistance gene was mapped.In addition,95 of the 300 markers exhibit polymorphisms,which can be used to investigate the diversity of rye 6RL arms.The markers developed in this study can be used to identify given segments of 6RL in wheat backgrounds and accelerate the utilization of elite genes on 6RL in wheat breeding.

Structural and biochemical mechanism responsible for the stay-green phenotype in common wheat

In order to increase the supply of assimilated carbon to grain, a new stay-green wheat cultivar, Chuannong17 (CN17), with delayed leaf senescence, carrying wheat-rye 1RS.1BL translo- cated chromosome was developed. CN17 exhibited distinct differences in net photosynthetic rate (Pn), chlorophyll (Chl) content, malondialdehyde (MDA) content, activity of both superoxide dismutase (SOD) and catalase (CAT) during the grain filling stage, and flag leaf senescence compared with the control. The new cultivar maintained longer and higher photosyn- thetic competence compared with the control, and this aspect correlated with the difference in chloro-plast development. Moreover, the stay-green pheno-type of CN17 was also observed under natural growth conditions. Consequently, the coordination of the physiological, biochemical, and structural aspects in the stay-green cultivar produced higher seed weights and per-plant yield compared with the control cultivar.

Analysis of DNA methylation variation in wheat genetic background after alien chromatin introduction based on methylation-sensitive amplification polymorphism

During the process of alien germplasm introduced into wheat genome by chromosome engineering, extensive genetic variations of genome structure and gene expression in recipient could be induced. In this study, we performed GISH (genome in situ hybridization) and AFLP (amplified fragment length po-lymorphism) on wheat-rye chromosome translocation lines and their parents to detect the identity in genomic structure of different translocation lines. The results showed that the genome primary struc-ture variations were not obviously detected in different translocation lines except the same 1RS chromosome translocation. Methylation sensitive amplification polymorphism (MSAP) analyses on genomic DNA showed that the ratios of fully-methylated sites were significantly increased in translo-cation lines (CN12, 20.15%; CN17, 20.91%; CN18, 22.42%), but the ratios of hemimethylated sites were significantly lowered (CN12, 21.41%; CN17, 23.43%; CN18, 22.42%), whereas 16.37% were fully-me-thylated and 25.44% were hemimethylated in case of their wheat parent. Twenty-nine classes of me-thylation patterns were identified in a comparative assay of cytosine methylation patterns between wheat-rye translocation lines and their wheat parent, including 13 hypermethylation patterns (33.74%), 9 demethylation patterns (22.76%) and 7 uncertain patterns (4.07%). In further sequence analysis, the alterations of methylation pattern affected both repetitive DNA sequences, such as retrotransposons and tandem repetitive sequences, and low-copy DNA.