Bt水稻杂交育种中转基因的遗传分析

利用PCR、GUS染色和Western印迹杂交技术检测了Bt水稻杂交后代群体,发现在394株GUS阳性株中,共有392株表达Bt蛋白,协同表达株率达99.49%。由此表明,在杂交后代中报告基因Gus和目的基因crylAb紧密连锁遗传与表达。本试验还发现,在BC1、BC1F2和粳粳交F2群体中转基因呈单基因显性遗传,而在籼粳交F2群体中偏离3:1分离。 Abstract:Improved histochemical staining for GUS activity,PCR and Western blotting were used to detect the population of Bt rice crossed to conventional rice varieties.A total of 392 plants expressing Bt toxin protein were found in 394 GUS positive plants.The result demonstrated that cry1Ab gene closely inherited and expressed with reporter gene gus.Therefore,it is possible to develop GUS-assisted-selection to preliminarily identify the Bt gene and study the inheritance of transgenes in (back)cross breeding.Mendelian segragation of reporter gene Gus was observed in F2,BC1 and BC1F2 progenies.Thus indicated that transgenes inherited as a single dominant gene in the progenies of Bt rice crossed to conventional rice varieties.     

水稻空间诱变恢复系杂种优势测定试验初报

利用返回式科学卫星搭载水稻一晚明恢63、外七、湘哥和粳稻北K15等纯系干种子,由于空间环境条件的综合作用,使空间诱变的品种均产生了变异。从诱变后代的突变体中选育出对三系籼型不育系具有恢复能力的强恢复系明恢63、无恢复能力的籼稻外七、湘哥和粳稻北K15。同时选出早熟强恢复系SP40098、具有育性恢复能力的SP40115、SP40037、SP50025等恢复系突变体。结果说明,空间诱变既可改良现有恢复系,又可创造新恢复系。具恢复能力的突变体能遗传重现。 Abstract：Pure seeds of rice varieties including Minghui 63,Wa i qi,Xiangge and Bei K 15（Japanica rice）,were carried into space by retrieve d satellite, and under space condition, mutation took place in all the s eed above mentioned. From these mutants, new restorer lines Minghui 63 with r estoring ability to the sterile line of three-line indica hybrid rice had be en developed, and other early restorers SP40098,SP40115,SP40037 and SP50025 ha ve also been selected from the mutants of Indica rice Waiqi,Xiangge,and Ja panica rice Bei K 15.The results indicated that space mutagenesis can n ot only improve existing restorers,also create new restorers.In addition,thei r restoring ability can be stably inherited.     

Transcriptional silencing and developmental reactivation of cry1Ab gene in transgenic rice

One transgenic rice line lacking CrylAb expression product was screened in the progenies of Agrobacterium-transformed transgenic rice variety Zhong 8215 with a cry1Ab gene under field releasing conditions by using GUS histochemical assay and Western blot. Molecular hybridization results revealed that the crylAb gene was silenced in the transgenic rice variety Zhong 8215 and two copies of ubiquitin promoter were integrated into the rice genome. The silencing of crylAb gene in transgenic rice was found to be due to the methylation of the ubiquitin promoter as revealed by methylation analysis. Meanwhile, different concentrations of demethylation reagent 5-azacytidine combining with different treatment time were employed to treat the silenced transgenic rice seeds. The results indicated that 5-azacytidine could reactivate 8%-30% of the silenced transgenic rice plants and the expression level of the reactivated cry1Ab transgene could reach as high as 0.147% of the total soluble protein. Treatment with low con     

Mapping QTLs for heading synchrony in a doubled haploid population of rice in two environments

Simultaneous heading of plants within the same rice variety, also refer to heading synchrony, is an important factor that affects simultaneous ripening of the variety. Understanding of the genetic basis of heading synchrony may contribute to molecular breeding of rice with simultaneous heading and ripening. In the present study, a doubled haploid （DH） population, derived from a cross between Chunjiang 06 and TN1 was used to analyze quantitative trait locus （QTL） for heading synchrony related traits, i.e., early heading date （EHD）, late heading date （LHD）, heading asynchrony （HAS）, and tiller number （PN）. A total of 19 QTLs for four traits distributed on nine chromosomes were detected in two environments. One QTL, qHAS-8 for HAS, explained 27.7% of the phenotypic variation, co-located with the QTLs for EHD and LHD, but it was only significant under long-day conditions in Hangzhou, China. The other three QTLs, qHAS-6, qHAS-9, and qHAS-10, were identified under short-day conditions in Hainan, China, each of which explained about 11% of the phenotypic variation. Two of them, qHAS-6 and qHAS-9, were co-located with the QTLs for EHD and LHD. Two QTLs, qPN-4 and qPN-5 for PN, were detected in Hangzhou, and qPN-5 was also detected in Hainan. However, none of them was co-located with QTLs for EHD, LHD, and HAS, suggesting that PN and HAS were controlled by different genetic factors. The results of this study can be useful in marker assisted breeding for improvement of heading synchrony.     

Additive and additive × additive interaction make important contributions to spikelets per panicle in rice near isogenic (Oryza sativa L.) lines

Epistasis plays an important role in the genetic basis of rice yield traits. Taking interactions into account in breeding programs will help the development of high-yielding rice varieties. In this study, three sets of near isogenic lines (NILs) targeting three QTLs for spikelets per panicle (SPP), namely qSPP1, qSPP2 and qSPP7, which share the same Zhenshan 97 genetic background, were used to produce an F 2 population in which the three QTLs segregated simultaneously. The genotypes of the individual F 2 plants at the three QTLs were replaced with three markers that are closely linked to the corresponding QTLs. These QTLs were validated in the F 2 and F 3 popula- tions at the single marker level. qSPP7 exhibited major pleiotropic effects on SPP, plant height and heading date. Multifactor analysis of variance was performed for the F 2 population and its progeny. Additive × additive interaction between qSPP2 and qSPP7 had significant effects on SPP in both the F 2 population and its progeny. Both additive and additive × additive interactions could explain about 73% of the total SPP phenotypic variance. The SPP performance of 27 three-locus combinations was ranked and favorable combinations were rec- ommended for rice breeding in different ecosystems.     

The breeding of two polyploid rice lines with the characteristic of polyploid meiosis stability

Polyploidization is a basic feature of plant evolution. Nearly all of the main food, cotton and oil crops are polyploid. When ploidy levels increase, yields double; this phenomenon suggested a new strategy of rice breeding that utilizes wide crosses and polyploidization dual advantages to breed super rice. Because low seed set rates in polyploid rice usually makes it difficult to breed, the selection of Ph-liked gene lines was emphasized. After progenies of indica-japonica were identified and selected, two poly- ploid lines, PMeS-1 and PMeS-2 with Polyploid Meiosis Stability (PMeS) genes were bred. The proce- dure included seven steps: selecting parents, crossing or multiple crossing, back-crossing, doubling chromosomes, identifying the polyploid, and choosing plants with high seed set rates that can breed themselves into stable lines. The characteristics of PMeS were determined by observing meiotic be- haviors and by cross-identification of seed sets. PMeS-1 and PMeS-2, (japonica rice), have several characteristics different from other polyploid rice lines, including a higher rate of seed set (more than 65%, increasing to more than 70% in their F1 offspring); and stable meiotic behaviors (pairing with bi- valents and quarivalents nearly without over-quarivalent in prophase, nearly without lagging chromo- somes in metaphase and without micronuclei in anaphase and telophase). The latter was obviously different from control polyploid line Dure-4X, which displayed abnormal meiotic behaviors including a higher rate of multivalents, univalents and trivalents in prophase, lagging chromosomes in metaphase and micronuclei in anaphase and telophase. There were also three differences of the breeding method between PMeS lines and normal diploid lines: chromosomes doubling, polyploidism identifying and higher seed set testing. The selection of PMeS lines is the first step in polyploid rice breeding; their use will advance the progress of polyploid rice breeding, which will in turn offer a new way to breed super rice.     

Introduction of a rice blight resistance gene,Xa21, into five Chinese rice varieties through anAgrobacterium-mediated system

A cloned gene, Xa21 was transferred into five widely-used Chinese rice varieties through an Agrobacterium-mediated system, and over 110 independent transgenic lines were obtained. PCR and Southern analysis of transgenic plants revealed the integration of the whole Xa21 gene into the host genomes. The integrated Xa21 gene was stably inherited, and segregated in a 3 : 1 ratio in the selfed T1 generation when one copy of the gene was integrated in the transfor-mants. Inoculation tests displayed that transgenic T0 plants and Xa21 PCR-positive T1 plants were highly resistant to bacterial blight disease. The selected Xa21 homozygous resistant transgenic lines with desirable qualities may be propagated as new varieties or utilized in hybrid rice breeding.     

一种紫色水稻的遗传及其在光敏不育系育种中应用的研究

研究表明,本院获得的一种紫色水稻的植株色遗传受控于C、 A、Pl3个独立基因座位上显性基因的互补作用,另有一独立的显性基因对Pl基因的表达起抑制作用。由于该抑制基因在籼稻中的高频率存在,因而,紫稻与一般绿稻品种杂交F1多表现为绿株。紫稻光敏不育(株)系的不育性表达和配合力表现,均可达到与普通光敏不育系相似的水平。本文还讨论了选育籼型紫稻光敏不育系设想的可行性。 Abstract:The inheritance of a purple rice in crosses to green rices was investigated.A group of dominant and interactive genes,C,A and Pl,was found to control the expression of the trait and the other independent inhibitor I-Pl-1 to inhibit the effect of the gene Pl.Because of the wide existence of the gene I-Pl-1 in indica rice,most of F1 plants of the crosses between purple rice and green cultivars were green.The primary study indicated that,for the purple photoperiod-sensitive genic male-sterile lines,the degrees of the sterility and its stability in the sterile stage,and of the fertility in the fertile stage,and the combining ability levels of the purple rice were as high as the degrees and levels for the green rice.A tentative idea on breeding purple photoperiod-sensitive male-sterile lines of indica rice was suggested and its feasibility and advantage was discussed.     

Diversity and stability study on rice mutants induced in space environment

To further study the characteristics of changes on the molecular level of rice mutants induced in space environment, we analyzed proteins in leaves and seeds of four rice mutants （two high-tillering and two low-tillering） in the 8th and 9th generations after a 15-day spaceflight, and compared with their ground controls by two-dimentional polyacrylamide gel electrophoresis and reverse phase liquid chromatography （RPLC）. In addition, the albumin, globulin, prolamine, glutelin, and amylose of the mutant seeds were analyzed by RPLC and ultra-violet spectrometry. The results showed that the low-abundance proteins of leaves in the peak tillering stage are more likely to he induced compared with their corresponding controls. The albumin, globulin, and prolamine of the mutant seeds revealed changes when compared with their controls, and the characteristics of changes in different mu- tants were stably inherited in the 8th and 9th generations, suggesting that they can be used as biomarkers to identity the mutants induced by spaceflight. Moreover, two proteins （SSP9111 and SSP6302） were found to be expressed with high intensity （two-fold change） in different mutants, which were both correlated with photosystem according to mass spectrometry and database searching.     

Potential Autonomous Selfing in Gesneria citrina （Gesneriaceae）, a Specialized Hummingbird Pollinated Species with Variable Expression of Herkogamy

Species with mixed mating systems often demonstrate variable expression of breeding system characteristics and thus represent the opportunity to understand the factors and mechanisms that promote both outcrossed and selfed seed production. Here, we investigate variation in levels of herkogamy （variation in stigma-anther separation distance） in a Puerto Rican population of hummingbird pollinated Gesneria citrina Urban. There is significant variation in herkogamy levels among individuals of this species and stigma-anther separation is negatively associated with the ability to set fruits and seeds in the absence of pollinators. The variation in levels of herkogamy may represent a mechanism to ensure the production of some self-fertilized progeny in the absence of hummingbird pollinators. We also describe a novel breeding system in G. citrina, where stamens elongate over time to reach stigma height, but stamen elongation is accelerated by pollination. These results suggest that once the flowers are pollinated, stamen elongation may favor increased pollen removal and siring success, while the reduction in stigma-anther distance no longer imposes the risk of interference between male and female functions. We discuss our findings of breeding system variation in the context of pollination system evolution in an island setting （Antillean islands）.