Changes in Physiological and Biochemical Characteristics of Floral Organ Development in a Soybean Cytoplasmic-nuclear Male Sterile Line

[Objectives]This study was conducted to explore the mechanism of soybean cytoplasmic-nuclear male sterility.[Methods]With soybean cytoplasmic-nuclear male sterile line JLCMS9 A and its homotype maintainer line JLCMS9 B as experimental materials,the activity of superoxide dismutase(SOD),peroxidase(POD)and catalase(CAT),malondialdehyde(MDA)content,starch content,soluble protein content,soluble sugar content and free proline content in flower buds,alabastrums and mature flowers were determined,and the contents and changes of auxin(IAA),gibberellin(GA3),isopentenyl adenosine(iPA)and abscisic acid(ABA)at the three stages were analyzed.[Results]The activity of SOD and CAT and the contents of MDA and free proline in the sterile line at the flower bud stage were lower than those of the maintainer line,but the opposite was true at the alabastrum stage and the flowering stage,and their values were higher than those of the maintainer line;the POD activity of the sterile line was significantly lower than that of the maintainer line at the flower bud stage,and the opposite was true at the alabastrum stage and the flowering stage,and its values were higher than those of the maintainer line;and the starch content and soluble sugar content of sterile line 9 A showed an overall upward trend,and were significantly lower than those of the maintainer line 9 B at the alabastrum stage and the flowering stage.During the whole development process of floral organs,the content of IAA in sterile line 9 A showed a trend of first increasing and then decreasing,and the content of iPA increased gradually.The contents of hormones in the sterile line were lower than those in the maintainer line.The ratios of IAA/ABA,IAA/GA3,IAA/iPA and ABA/GA3 in the sterile line were significantly different from those in the maintainer line.It is inferred that the abnormal physiological characteristics of floral organ development are related to the cytoplasmic-nuclear male sterility of soybean.The alabastrum stage may be a critical period for the occurrence of abnormal physiological and biochemical indexes in the floral organs of soybean cytoplasmic-nuclear male sterile lines.[Conclusions]This study provides a theoretical basis for the breeding of fine sterile lines of soybean and the research on the mechanism of sterility.

Preliminary Studies on the Effect of Photoperiod and Temperature on Male Fertility of a Cytoplasmic Male Sterile Line in Wheat

[Objective] The study aimed to reveal the effect of photoperiod and temperature on male fertility of cytoplasmic male sterile line Vtai911289a in wheat and discuss the mechanism of male fertility alteration. [ Method ] The sowing-date tests and designed conditions were conducted during 2003 -2005. [ Result] Fertility of Vtai911289a, could alter under specific photoperiod and temperature conditions. Temperature is one of the main factors influencing male fertility of the male sterile lines. Vtai911289a, showed stable sterility under the condition of the mean of daily temperature at fertility sensitive stage lower than 19℃ and presented partial fertility when the mean of daily temperature at fertility sensitive stage lower than 20 - 22℃. Photoperiod to some extent affects the male fertility of Vtai911289a, long-day condition is helpful for the male fertility of the sterile line. [ Conclusion] The application of photoperiod temperature-sensitive cytoplasmic male sterile line in production has a higher safety than that of temperature sensitive sterile line.

Influence of Genetic Drift of Restoring Gene (Rf) on Seed Purity of Yuetai A, a Honglian-Type Cytoplasmic Male Sterile Line in Hybrid Rice

The seed samples of Yuetai A, a Honglian （HL） type cytoplasmic male sterile （CMS） line in hybrid rice were investigated to assess the seed purity and to analyze the cause of off-type plants by imitating the biological contaminant to Yuetai A in Nanjing, Jiangsu Province and Lingshui, Hainan Province during 2004-2006. The seed impurity of Yuetai A mainly resulted from the genetic drift of restoring fertility gene （Rt） after biological contamination but not from its sterility unstability. All of the ten maintainer lines, five restorer lines and three thermo-sensitive genic male sterile lines used in the study could pollinate Yuetai A and Yuetai B to produce F1 plants, directly or indirectly resulting in Rf-gene drifting into Yuetai A and generating ＇iso-cytoplasm restoring-generations＇. Furthermore, high outcrossing rate and similar heading date of Yuetai A with many varieties used in rice production might easily result in the biological contamination. After removing all plants with Rf-gene mixed in Yuetai A and preventing Rf-gene drifting into Yuetai A, the seed purity of Yuetai A and Yuetai B had been raised to 100%.

Development and Application of SCAR Markers for Discriminating Cytoplasmic Male Sterile Lines from Their Cognate Maintainer Lines in Indica Rice

The DNAfragments about 1 600 bp were amplified using random amplified polymorphism DNA （RAPD） primer OPAl2 with the templates of mitochondrial DNA of Zhenshan 97A and Zhenshan 97B, and were sequenced. The nucleotide sequences and lengths of the fragments from Zhenshan 97A and Zhenshan 97B showed no difference. The precise length of the fragment was 1 588 bp. Sequence characterized amplification region （SCAR） primers were then developed to discriminate the cytoplasmic male sterile （CMS） lines and their maintainer lines. A specific 1 588 bp fragment could be amplified with SCAR primers, CHI19F2/CHI19R2 and CHI20F3/CHI23R3, in the mitochondrial DNA of Zhenshan 97A, but not Zhenshan 97B. Furthermore, the specific fragment could be also amplified from the total DNA from green leaf tissues of Zhenshan 97A with SCAR primers, but not Zhenshan 97B. With the corresponding primers, the specific fragment could also be amplified from the total DNA of green leaves of other two CMS lines with wild abortive type cytoplasm （CMS-WA）, namely Zhenpin A and Tianfeng A, but not in their maintainer lines. Moreover, using total DNA as template, each of the four pairs of SCAR primers could also be used to amplify the 1 588 bp fragment in CMS-ID （Indonesia paddy type） line 11-32A but not in 11-32B, and the specific fragment was amplified from the DNA of both F1 and F2 seedlings of Shanyou 63. The results of detecting the genetic purity of a man-made mixture of the seeds of Zhenshan 97A using CHI20F3/CHI23R3 were completely consistent with the phenotypes. Taken together, these results indicated that the specific 1 588 bp-fragment amplified by CHI20F3/CHI23R3 was the unique amplification products of CMS mitochondrial DNA, and could be used to distinguish CMS-WA and CMS-ID lines from their corresponding maintainer lines at the seedling stage.

Differences in Nuclear DNA Between Male-Sterile and Male-Fertile Lines of Sorghum bicolor

Cytoplasmic male sterility(cms)is determined by nuclear-cytoplasmic interactions. Up to now, most studies are focused on the comparison of cytoplasmic DNAs of male-sterile lines and male-fertile lines, and analysis of nuclear DNA has not been documented yet. In order to find out the possible difference in nuclear genome of male-sterile line A1 Tx623 and corresponding male-fertile line Tx623 of sorghum, random amplified polymorphic DNA(RAPD)approach was used to analyze their cytoplasmic and nuclear genomes. Total DNAs of them were amplified at first to screen primers, which were able to generate reproducible bands specific to male-sterile line or male-fertile line. Then the selected primers were used to amplify their mitochon-drial DNA(mtDNA)and chloroplast DNA(cpDNA). The origins of all the polymorphic fragments were analyzed. After ruling out those amplified from cytoplasmic DNA, seventeen polymorphic fragments were determined to be amplified from nuclear DNA. These fragments originated from nuclear DNA indicate that differences in sequence exist between the nuclear DNA of male-sterile line and male-fertile line of sorghum, which do not agree with the traditional standpoint that they have identical nucleus.

Genetic Mechanism of Dominant Earliness in Kefeng A,a New Rice Cytoplasmic Male Sterile Line

Kefeng A is an early maturing indica cytoplasmic male sterile （CMS） line of rice. Combinations derived from Kefeng A and late maturing indica restorer lines showed dominant earliness to various extents. To understand the genetic basis of dominant earliness, the genotype of photoperiod-sensitive genes in Kefeng A was analyzed using a complete set of heading time near isogenic lines （NILs） EGO to EG7, ER, LR, T65, T65E^b, T65E^bm, T65m, NIL（Hd1） and NIL（Hd4）. Results indicated that Kefeng A contained two dominant photoperiod-sensitive alleles E1 and Se-1^U on E1 and Se-1 loci, respectively, and the genotype of photoperiod-sensitivity genes for heading time in Kefeng A was E1E1e2e2E3E3Se-1^USe-1UEf-1Ef-1. Based on the detected heading time genotype, in combination with the heading time of Kefeng A and the early maturing phenomenon in its derived F1 hybrids, it is speculated that Kefeng A might carry a dominant inhibitor gene Su-E7 for the dominant photoperiod- sensitive gene E1, and a recessive inhibitor gene i-Se-1 for another dominant photoperiod-sensitive gene Se-1. The reason why F1 hybrids from Kefeng A exhibited early maturing was hereby analyzed and the breeding value of dominant earliness related genes in Kefeng A was discussed.

Maxie A- a novel cytoplasm male sterile line

The pollen abortion of Maxie A and Zhengshan97 A belong to stainless type with Ksolu-tion.The maiority of Maxie A abortive Pollenstook cireular shape and few was staining abortion

Analysis of Heterosis, Combining Ability and Heritability of Cadmium Content in Brown Rice of Three-line Indica Hybrid Rice

Five cytoplasmic male sterile （CMS） lines were used as parents in an incomplete diallet cross and six restorer lines of rice design （Nc II design）. Thirty hybrid combinations with the same growth period were selected as experimental ma- terials to investigate the heterosis, combined ability and heredity of Cd content in brown rice of indica hybrid rice. According to the results, Cd content in brown rice showed a significantly negative heterosis; the general combining ability and specific combination ability of Cd content in CMS and restorer lines both reached extremely significant level （P〈0.01）, indicating that both genetic improvement of parents and e- valuation of combinations are important to the breeding of hybrid combinations with low accumulation of Cd; the broad-sense heritabitity and narrow-sense heritability of Cd content were both relatively high with slight differences, which respectively reached 97.73% and 80.10%, indicating that Cd content in brown rice mainly de- pends on the additive action of genes; in addition, parent improvement showed bet- ter effect on the selection of early generation.

辣椒雄性不育分子研究进展

为对辣椒雄性不育分子研究取得的进展有更直观、系统、深入的了解,梳理已鉴定的辣椒核雄性不育(NMS)基因及部分NMS基因分子标记开发,重点阐述近年来核雄性不育基因精细定位及候选基因探究工作;同时回顾辣椒胞质雄性不育(CMS)相关的线粒体不育基因及其关联分子标记和CMS恢复基因分子标记开发,重点概述近年来恢复基因精细定位研究取得的进展。研究指出,精细定位NMS基因,明确候选基因并开发功能标记是辣椒NMS研究的重点,既能为探明辣椒NMS机理奠定基础又可为育种提供高效利用的分子标记。而CMS相较于NMS更为复杂,表现在遗传机制多样且更易受环境影响等方面,因此,从对CMS育性恢复基因的精细定位到基因克隆以及功能研究并探明CMS分子机制还有很长的一段路要走。

Sequence analysis of the gene correlated with cytoplasmic male sterility (CMS) in rape-seed (Brassica napus) Polima and Shaan 2A

orf224 is a CMS-related mitochondrial gene discovered in Polima cytoplasm. Shaan 2A CMS line is the parent of the first rapeseed hybrid cultivar Qinyou No. 2 that has been grown in many regions of China. In this work, genomic DNA of Polima CMS line and Shaan 2A CMS line were used as templates, two primers of specific oligonucleo-tides at 5’ and 3’ ends were used, PCR was performed, the amplification fragments were cloned into pGEM-T Easy vectors and DNA sequences were determined. The CMS-associated gene, orf224-1 present in Shaan 2A CMS line, has a sequence highly homologous to the orf224 of the Polima CMS line, except for one nucleotidc at position +398. There were only one base (AAC→AGC) and one amino acid (Asn→Ser) differences between the two. The homologies of the two sequences in nucleotide and amino acid were 99.9% and 99.6%, respectively. It is concluded that orf224 in Polima CMS line and orf224-1 of Shaan 2A CMS line are the allele at the same locus in mitochondria.

高粱线粒体雄性不育基因KASP标记开发

为缩短育种年限、提高育种效率及创制新种质,本研究以96份高粱不育系、保持系和恢复系为材料,通过线粒体重测序技术、表型数据与SNP位点的卡方检验,发掘与育性相关的SNP位点,并开发KASP分子标记位点。结果鉴定出与育性相关的SNP位点434个,筛选出101个位点进行卡方检验,SNP位点χ^(2)=9.574,p=0.002,其变异位点以颠换为主,整体Tv/Ts为1.55。区别出含有不育基因的材料49个,可育基因型材料47个。筛选出的SNP位点共开发出19对高质量的KASP引物,利用这些标记可以准确地区分高粱不育系和保持系,并能够鉴定出恢复系中是否含有粒线体雄性不育基因。将这些SNP位点开发成分子标记将在育种过程中节约大量的人工和时间成本,并提高准确度。

花椰菜异源胞质雄性不育系C50-2选育研究

用具有 Ogura胞质雄性不育基因的花椰菜雄性不育材料 NYAcms为母本 ,不同花椰菜品种为父本 ,通过连续 6代回交选育 ,育成异源胞质雄性不育系 C5 0 - 2及其相应的保持系 5 0 11- 2。C5 0 - 2不育系具有生长发育正常、抗病、农艺性状优、配合力好等特点 ,通过组合配制试验 ,初步筛选出 2个优良组合在生产上示范推广。

玻里马胞质大白菜雄性不育系CMS3411-7温度敏感特性的研究

采用光照培养箱恒温、变温和田间自然变温 ,研究大白菜玻里马胞质雄性不育系CMS 3411 7温度敏感的特性。结果表明 :在 3～ 15℃内CMS 3411 7均有育性转换 ,其中昼 /夜温度 8/ 4～ 14 / 10℃ (即日平均温度 6～ 12℃ )是有效转换的温度范围。在恒温 6℃条件下 ,经过 3～ 5d处理时CMS 3411 7的育性转换值 (不育级别 )可达到 2级 ,该温度下随着处理时间延长 (3～ 9d)不育性转换程度逐渐加重。在 9℃恒温下处理 6d较处理 3d和处理 9d时的转换程度都大。在 14 / 10℃下转换程度与处理时间无关。在 17/ 13℃下处理时间延长反而抑制不育性转换。虽然不同处理下CMS 3411 7不育性转换开始和转换结束的时间差异较大 ,但显著转换发生的时间一般稳定在温度处理后 10～ 2 7d ,表现相对集中。就育性转换的启动来看 ,经过 3d处理后 8/ 4～ 17/ 13℃下均发生了育性转换 ,说明

大白菜新型细胞质雄性不育系6w-9605A的育性鉴定和花药败育的细胞学观察

采用石蜡切片技术,研究了大白菜(Brassica campestris L. ssp. pekinensis)细胞质雄性不育系6w-9605A及其保持系6w-9605B的花药发育过程的细胞形态学特征,确定不育系花药败育时期及方式,并对不育系6w-9605A进行花器官观察和育性鉴定。结果表明:保持系6w-9605B花药发育正常;不育系6w-9605A花药发育受阻于孢原分化时期,占总败育花药的66.7%,不形成花粉囊和花粉粒,属于无花粉囊型败育;另外33.3%的败育花药可形成花粉囊,小孢子均受阻于单核靠边期或者二胞期,败育特点为绒毡层细胞异常肥大,挤压小孢子,导致小孢子和绒毡层解体;6w-9605A的不育性稳定、彻底,不育株率和不育度均为100%。

水稻CMS-WA和CMS-Y育性恢复基因Rf3和Rf4的等位性分化

为了检测水稻第1染色体上恢复基因Rf3和第10染色体上恢复基因Rf4的等位性分化,利用8个携带Rf3基因座位,16个携带Rf4基因座位的染色体单片段代换系(chromosome single segmentsubstitution line,SSSL)和华粳籼74(HJX74)为父本,野败型(WA)细胞质雄性不育系珍汕97A(ZsA)和Y型(Y)细胞质雄性不育系Y华农A(HnA)为母本杂交,采用分子标记辅助选择的方法,鉴定携带基因型Rf3rf3/Rf4rf4的F1杂种株并对其花粉和小穗育性进行考察。结果表明:(1)24个SSSLs和HJX74对于2个不育系的恢复力存在着显著的不同,携带Rf3基因座位的SSSLs恢复力均低于携带Rf4基因座位的SSSLs,并且低于对照品种HJX74;SSSL S6对于WA-CMS的花粉小穗育性分别为7.2%和15.5%,对于Y-CMS的花粉小穗育性分别为1.3%和12.4%,表现出最弱的恢复力;SSSLs S14-S18和SSSL S20对这2种不育系的平均花粉和小穗育性分别高于70%和85%,表现出较强的恢复力。(2)在恢复基因Rf3和Rf4基因座分别鉴定出3个和4个等位基因,由弱到强依次命名为Rf3-1、Rf3-2、Rf3-3和Rf4-1、Rf4-2、Rf4-3、、Rf4-4,受体亲本HJX74的基因型为Rf3Rf3/Rf4Rf4,其Rf3的恢复性强于Rf4恢复性。(3)在HJX74的遗传背景下,WA型不育细胞质的可恢复性好于Y型不育细胞质。

籼稻体细胞克隆雄性不育系54257/162-5及其保持系

本文报道了籼稻雄性不育系54257/162-5的基本特性,其母本54257及父本162-5均为离体培养起源的体细胞无性系,分别来自 IR_(54)及IR_(52)。 不育系54257/162-5已回交8代,不育株率100％,自交不结实。花粉败育方面,部分个体表现为典败,部分个体表现为染败,或典败与染败兼而有之。试验表明,不育系54257/162-5个体间这种花粉败育的“分离”并非父本不纯所致,乃该不育系固有的特点。 以114恢、测64、IR_(24)、3550、测222及丰桂6号等6个野败型不育系之恢复系与野败不育系及 54257/162-5测交,恢复程度有较大差异。只有3个(114恢、测64及测222)能使54257/162-5的育性恢复;2个(IR_(24)及3550)只能达到半恢复;有1个(丰桂6号)不仅不能恢复,而且保持了其雄性不育。这表明,不育系54257/162-5与野败不育系在细胞质上可能既有相似的一面,也有相异的一面。 讨论了保持系体细胞克隆162-5的基因型,认为它是由恢复系IR_(52)经离体培养直接变为保持系。这在体细胞无性系变异中尚属首例。

不同核背景对小麦V-CMS coxⅢ基因转录本编辑的影响

RNA编辑现象普遍存在于高等植物线粒体中,RNA编辑的不充分或偏离与植物细胞质雄性不育具有相关性。以V型小麦细胞质雄性不育恢复基因的近等基因系为材料,研究了不育系、保持系、恢复系和杂种F1 coxⅢ基因转录本编辑位点的差异,发现coxⅢ基因转录本共有14个编辑位点,其中有12个引起了氨基酸种类的变化。通过对不育系、保持系、恢复系和杂种F1线粒体RNA编辑频率的比较,发现引入恢复基因后,不育胞质中coxⅢ基因转录本的编辑频率明显提高,说明coxⅢ基因转录本的RNA编辑与细胞质雄性不育具有一定相关性。

玉米CMS-S恢复基因Rf_3近等基因系的蛋白质分析

【目的】Rf3是玉米CMS-S型不育系两个以上恢复基因中的标准基因,对其进行分子生物学研究意义很大。【方法】利用自主创建的Rf3近等基因系不育群体(Ps)和可育恢复系群体(Pf)为试材,采用改进的双向(二维)聚丙烯酰胺凝胶电泳(2D-PAGE)技术,对苗期、成株期叶片细胞总蛋白进行分析。【结果】分离到在Pf遗传背景下分子量为25.2kD,等电点为5.0的特异蛋白质RRPⅡ,优化了2D-PAGE技术。【结论】该特异蛋白质点的出现与消失可能与玉米CMS-S的育性恢复有一定关系,对其纯化分离将为Rf3基因的克隆奠定基础和进一步了解玉米质核互作不育表达模式的分子机制。结合本研究,文章还讨论了采用2D-PAGE技术在玉米分子生物学研究中的优化以及基因组学与蛋白组学研究的衔接。

甘蓝型油菜改良型不育系RGCMS-1141A的选育

利用作者提出的“植物细胞核 +细胞质雄性不育”的理论 ,对在生产上大面积推广的甘蓝型油菜波里马细胞质雄性不育优质杂交种“华杂 4号”的亲本不育系 1141A进行遗传改良 ,选育出遗传背景基本保持不变的油菜细胞核 +细胞质雄性不育系RGCMS 1141A及其保持系RGCMS