Functional roles of root plasticity and its contribution to water uptake and dry matter production of CSSLs with the genetic background of KDML105 under soil moisture fluctuation

Soil moisture fluctuation (SMF) stress due to erratic rainfall in rainfed lowland (RFL) rice ecosystems negatively affect production. Under such condition, root plasticity is one of the key traits that play important roles for plant adaptation. This study aimed to evaluate root plasticity expression and its functional roles in water uptake, dry matter production and yield under SMF using three chromosome segment substitution lines (CSSLs) with major genetic background of KDML105 and a common substituted segment in chromosome 8. The CSSLs showed greater shoot dry matter production than KDML105 under SMF, which was attributed to the maintenance of stomatal conductance resulting in higher grain yield. The root system development based on total root length of the CSSLs were significantly higher than that of KDML105 due to the promoted production of nodal and lateral roots. These results implied that the common substituted segments in chromosome 8 of the 3 CSSLs may be responsible for the expression of their root plasticity under SMF and contributed to the increase in water uptake and consequently dry matter production and yield. These CSSLs could be used as a good source of genetic material for drought resistance breeding programs targeting rainfed lowland condition with fluctuating soil moisture environments and for further genetic studies to elucidate mechanisms underlying root plasticity.     

以广东高州普通野生稻为供体亲本的水稻单片段代换系构建

以栽培稻品种粤香占为受体亲本,利用杂交、回交和微卫星标记辅助选择相结合的方法,构建了以高州普通野生稻为供体亲本的水稻单片段代换系群体。该群体由20个编号的9个单片段代换系构成。这9个单片段代换系分别分布在水稻的第1、2、3、10和11染色体上,代换片段长度为8.1～23.8cM,总长度为152.7cM,平均长度为17.0cM,代换片段对水稻基因组的覆盖总长度为136.1cM,覆盖率为7.5%。构建以高州普通野生稻为供体亲本的水稻单片段代换系,为发掘和利用高州普通野生稻的有利基因提供了理想的试验材料,为进一步开展高州普通野生稻的遗传研究提供了一条新途径。     

Utilizing Chromosome Segment Substitution Lines (CSSLs) for Evaluation of Root Responses to Transient Moisture Stresses in Rice

Abstract

Drought and waterlogging that occur sequentially under field conditions are important abiotic stresses affecting plant growth and development. The ability to maintain the root system development during the contrasting moisture stresses may be one of the key traits for plant adaptation. This study aimed to identify the key root traits that contributed to the above ability by comparatively examining the effects of the two moisture stresses in succession on root system development. The chromosome segment substitution lines (CSSLs) from the crosses between the japonica rice cultivar Nipponbare and indica rice cultivar Kasalath were used for precise comparison of root system development. The rice seedlings were grown by hydroponics under a continuously well-aerated condition for 14 days (non-stressed), a drought condition for 7 days followed by an oxygen (O₂)-deficient (stagnant) condition for 7 days (drought-to-stagnant, D-S), or a stagnant condition for 7 days followed by drought condition for 7 days (stagnant-to-drought, S-D). CSSL43 and 47 did not show any significant differences in growth from Nipponbare under the non-stressed condition, but exhibited greater lateral root production under the stresses. Lateral root production was most closely related to faster seminal root elongation mediated by higher aerenchyma formation in the D-S condition, and to more branching of lateral roots on the seminal root axis in the S-D condition. The D-S condition severely affected lateral root production due to reduced seminal root elongation and aerenchyma formation. These results confirmed the fact that those root traits previously identified using different cultivars greatly contribute to plant adaptation. Oxygen deficiency preceded by drought (D-S) was more stressful to roots than drought preceded by O₂ deficiency (S-D), because drought reduced root aerenchyma formation during the subsequent stagnant condition.     

Mapping of Hd-6-2 for Heading Date Using Two Secondary Segregation Populations in Rice

Heading date is one of the most important traits for rice adaption to different cultivation areas and crop seasons. In this study, two single segment substitution lines(SSSLs), W31-41-61-3-11-3-6-7(W31-SSSL) and W32-59-80-2-11-1-10(W32-SSSL) with substituted intervals derived from the donor parents IR66897 B(W31) and IR66167-27-5-1-6(W32), respectively, with Huajingxian 74(HTX74) were found to comprise a gene for extremely late-heading date, and the gene was tentatively designated as Hd-6-2. Two secondary F2 segregating populations were developed by crossing the two heterozygous SSSLs with HJX74 to map Hd-6-2 gene. According to phenotype analysis of the two mapping populations, the late heading date trait was controlled by a major recessive gene. In the segregation population derived from W31-SSSL, Hd-6-2 was mapped on chromosome 6 between PSM677 and RM204 with the genetic distances of 1.3 and 2.7 c M, respectively. In the population of W32-SSSL, the gene for heading date was mapped to the similar region as Hd-6-2 and co-segregated with PSM672. The sequence alignment of Hd3 a in the coding domains and promoter regions of HJX74 and W31-SSSL are completely consistent, whereas there was a great difference between W32-SSSL and HJX74, suggesting that Hd3 a could hardly be the main cause of the heading date variation in W31-SSSL, but it was probably the main reason for the change of heading stage in W32-SSSL.     

利用抗性基因替换的方法将两个拷贝外源基因表达单元整合到枯草芽孢杆菌染色体基因组同一位点

本研究提供了一种新的整合多拷贝外源基因表达单元到枯草芽孢杆菌染色体同一位点的方法。以β-淀粉酶表达单元作为应用实例,本方法包括以下步骤：首先,构建含有一个拷贝β-淀粉酶表达单元的整合质粒pMLK83-CTBA,通过同源双交换获得单拷贝β-淀粉酶表达单元整合到枯草芽孢杆菌1A751染色体α-淀粉酶基因位点的菌株1A751[CTBA]Neo＋;然后,利用抗性基因替换质粒pVK71,将新霉素抗性基因替换为状观霉素抗性基因,得到1A751[CTBA]Neo-Spe＋;最后,整合质粒pMLK83-CTBA再以同源单交换方式整合到1A751[CTBA]Neo-Spe＋染色体,通过新霉素抗性和状观霉素抗性筛选出两个拷贝β-淀粉酶基因整合的重组菌1A751[CTBA2]Neo＋Spe＋。结果显示,利用此方法增加β-淀粉酶基因的拷贝数,能够显著和稳定地提高β-淀粉酶的表达量。     

Genetic characterization of kernel polyphenol oxidases in wheat and related species

Polyphenol oxidase (PPO) activity causes undesirable darkening of raw Asian noodles and other wheat products. In this study we investigate the genetic origins and diversity of wheat kernel PPO. PPO was characterized via activity assays, antigenic staining, and Southern blots in Triticum aestivum, Triticum dicoccoides, Triticum durum, Triticum dicoccum, Triticum monococcum, Triticum urartu, Aegilops speltoides, and Aegilops tauschii. Among these species, PPO activity was well-correlated with antigenic staining intensity toward a wheat kernel-type PPO antibody. High PPO activity was observed in all three T. monococcum accessions (A^m genome), one Ae. speltoides accession, one T. durum accession, and two hexaploid wheat cultivars. Southern blots suggested the presence of two or more kernel-type PPO genes in diploid progenitors of the hexaploid A, B, and D genomes. Whole-kernel PPO activity was evaluated in disomic substitution lines derived from three T. dicoccoides accessions in the background of T. durum ‘Langdon’. PPO activity was primarily associated with chromosome 2A and to a much lower degree with chromosome 2B. DNA sequence comparisons showed that the intron associated with the high PPO allele on chromosome 2AL of hexaploid wheat had 94% nucleotide identity with the homeologous intron found in T. monococcum, a species with high kernel PPO activity. This implies that the ancestral PPO allele on the A genome is one of the high activity, and the low PPO allele found in hexaploid wheat represents a relatively recent genetic alteration. Results confirm the presence of multiple kernel-type PPO genes in the diploid and tetraploid progenitors and relatives of hexaploid wheat. However, it is likely that relatively few of the many kernel-type PPO genes present in wheat contribute substantially to kernel PPO activity. A single genetic locus on homeologous group 2 chromosomes may be the primary cause of high PPO activity in wheat kernels.     

染色体单片段代换系的构建及应用于QTL精细定位

染色体片段代换系(Chromosome Segment Substitution Lines,CSSLs),又称为代换系(Substitution lines,SLs)或导入系(Introgression lines,ILs)。只含一个代换片段的代换系,即单片段代换系(Single Segment Substitution Lines,SSSLs)是理想的代换系。单片段代换系只有代换片段与受体亲本不同,其它遗传背景与受体亲本完全一致,对代换区段中的QTL进行分析时遗传背景干扰很小,有利于QTL的分析,不少学者利用单片段代换系材料对许多QTL进行了鉴定和精细定位,并克隆了一些重要性状的QTL。本文介绍了染色体单片段代换系构建的原理和利用微卫星标记(SSR）进行单片段代换系鉴定的方法,讨论了代换系构建过程中值得注意的问题,并对用染色体单片段代换系进行QTL精细定位做了展望。     

利用染色体片段置换系定位水稻芽期耐冷性QTL

以籼稻品种9311为受体、粳稻品种日本晴为供体构建的95个染色体片段置换系为材料,在5℃低温条件下进行芽期耐冷性鉴定。结果表明,6个置换系低温处理后的成苗率与受体亲本9311有一定差异,其耐冷性略强于9311。利用代换作图法共鉴定出4个与芽期耐冷性相关的QTL,分别位于水稻第5和第7染色体上。其中qCTB-5-1、qCTB-5-2和qCTB-5-3分别被定位在第5染色体RM267与RM1237、RM2422与RM6054及RM3321与RM1054之间遗传距离分别为21.3cM、27.4cM和12.7cM的置换片段上;qCTB-7被定位在第7染色体RM11-RM2752区间遗传距离为6.8cM的置换片段上。     

Assessment of Genetic Diversity of Drought Tolerant and Susceptible Rice Genotypes Using Microsatellite Markers

The introgression of wild chromosomal segments into popular rice varieties is one of the potential approaches for developing varieties for drought stress condition. Sixteen genotypes, including nine indica, two tropical japonica and five chromosome segments substitution lines(CSSLs) with different levels of tolerance/susceptibility to drought stress, were selected for diversity study. Sixty-three microsatellite markers were utilized for assessing genetic diversity. A total of 95 alleles were amplified, and out of them, 60 were polymorphic. Six unique alleles, amplified by the microsatellite loci RM276, RM472,RM488, RM537, RM541 and RM28089, were identified in six genotypes, namely FR13A, Brahamanakhi,RUF44, Swarna-sub1, Brahamanakhi and Satyabhama. The highest genetic similarity was found among CSSLs. Polymorphism information content(PIC) value varied from 0 to 1.00 with an average of 0.66 per locus. Twenty-eight microsatellites were found to be polymorphic, which could be used in marker-assisted selection programme. All the sixteen genotypes were grouped into two major clusters at genetic similarity of 0.64. In the cluster I, five CSSLs identified as diverse genotypes had wild ancestor segments responsible for drought tolerance, and hence they could be utilized as potential donors. The popular Indian varieties, Swarna-sub1 and IR64-sub1, could be used as recurrent parents in the future breeding program for developing varieties for abiotic stresses such as submergence and drought.     

盐胁迫对小麦代换系光合性状的影响及染色体效应研究

为了选育小麦耐盐基因型,以中国春-Synthetic 6x小麦染色体代换系及其亲本为材料,设置对照(0mmol·L~(-1) NaCl)和盐胁迫(150mmol·L~(-1) NaCl)2个处理,通过测定不同处理条件下代换系幼苗比叶重、叶绿素和类胡萝卜素含量及光合速率的变化,探讨盐胁迫对小麦代换系幼苗光合性状的影响,并对其调控相关特性的基因进行染色体定位。结果表明,盐胁迫导致多数小麦代换系叶绿素和类胡萝卜素含量、比叶重、光合速率降低;盐胁迫条件下,Synthetic 6x的2D染色体上可能存在诱导比叶重升高的基因,1A、5A、7A、2B、3B、5B、6B和6D染色体上可能存在诱导叶绿素含量增加的基因,1A、4A、7A和5B染色体上可能存在诱导类胡萝卜素含量增加的基因,1A和7D染色体上可能存在诱导幼苗光合速率增强的基因,即Synthetic 6x的1A染色体上可能存在调控光合特性的关键基因。