木薯分子育种中若干基本科学问题的思考与研究进展

木薯作为全球重要的薯类作物,既是热带地区粮食安全的保障,也是重要的淀粉工业原材料,保障其稳产、高产、优质一直是育种家不变的研究主题．当前,木薯品种选育正处在从杂交育种转向分子育种的发展阶段,深入解析木薯特有的经济性状和生物学特点是利用生物技术进行遗传改良的基础．不同于谷物类作物,木薯光合同化物的转运和库源分配的调控机制必有其独特之处;同时,储藏根的“库容”直接影响其产量．作为热带作物,了解木薯对低温和干旱的响应可为改良其抗逆境能力提供理论依据．不同于其他薯类作物,木薯储藏根特有的“采后生理性变质”问题亟待解决,其发生和调控机制的解析对延长木薯货架期意义重大．随着分子生物学的发展,针对上述各方面的研究日益深入,不仅激发了感兴趣的公众对这些问题的认知和思考,也激励了科研人员不断努力寻找解析相关机制的方法,为最终通过分子育种手段改良木薯提供思路和技术方案,揭开木薯的层层“面纱”,推动木薯分子育种的发展．     

影响根癌农杆菌介导的木薯遗传转化因素分析

采用根癌农杆菌介导的叶盘转化法,以我国南方地区主栽木薯品种—华南8号的胚状体子叶为受体,对影响木薯遗传转化效率的主要因素进行了分析。研究结果表明,在木薯的遗传转化中,选用GV3101作为浸染外植体的农杆菌菌株,将感染时间和共培养时间分别控制在30~45 min和3~4 d、菌液浓度（OD₆₀₀）采用0.45、并添加200 μmol·L^-1的乙酰丁香酮（AS）均可明显提高其转化效率,但若对外植体进行预培养反而会降低其转化效率。利用该体系从453块外植体中共转化获得10株抗性再生植株,经PCR和Southern杂交检测,有8株木薯的基因组中已整合进了外源基因glgC336,转化率为1.77%。     

木薯的组织培养和基因转化

木薯是重要的热带作物之一, 其块根富含淀粉, 是热带、亚热带地区近５ 亿人粮食的主要来源。简单介绍近年来运用生物技术改良木薯的研究进展。     

木薯体细胞胚胎发生及植株再生研究

对木薯体细胞胚胎发生的影响因素进行了优化研究。结果表明,基因型对木薯体细胞胚胎发生影响很大,在供试的六个品种中,“华南 8 号”的体细胞胚胎发生率和产胚量最高,分别为 65% 和19个;侧芽茎尖为最佳外植体,体细胞胚胎发生的最佳培养基为MS +0.5mg/L CuSO4 + 4 mg/L 2,4-D。同时,对木薯体细胞胚再生成完整植株的主要影响因素作了分析,建立了一个高效的植株再生体系。     

Using microsatellites,isozymes and AFLPs to evaluate genetic diversity and redundancy in the cassava core collection and to assess the usefulness of DNA-based markers to maintain germplasm collections

The cassava core collection was selected to represent, with minimum repetitiveness, the potential genetic diversity of the crop. The core (630 accessions) was chosen from the base collection (over 5500 accessions) on the basis of diversity of origin (country and geographic), morphology, isozyme patterns and specific agronomic criteria. To asses the genetic diversity of the core, 521 accessions were typed with four microsatellite loci. Allele diversity and frequency, and size variance of dinucleotide repeats (Rst statistic) were estimated. Microsatellite allele numbers and frequencies varied among countries: Colombia and Brazil had the largest number of different alleles across all loci. Mexico also had a high number, ranking fifth after Peru, Costa Rica and Venezuela (which tied). Unique alleles were present in accessions from Brazil, Colombia, Guatemala, Venezuela and Paraguay. A small number (1.34%) of potential duplicates were identified through isozyme and AFLP profiles. Thus, the present results indicated that traditional markers have been highly effective at selecting unique genotypes for the core. Future selections of cassava germplasm sets can be aided by DNA-based markers to ensure genetically representative, non-redundant samples. This revised version was published online in June 2006 with corrections to the Cover Date.     

木薯雌雄花分化形态结构观察及生理调控研究

本研究以木薯品种新选048为试验材料,观察和测定木薯花分化各时期的形态结构、内源激素及碳氮化合物含量,并开展木薯雌雄花分化的外源激素调控研究,揭示木薯花分化规律,初步探究木薯花分化的生理机制.试验结果表明:木薯从花芽分化开始到完成开花的全过程需要30 d,分化出的花序主要有全雄花序和雌少雄多花序两种类型,木薯性别分化前...     

Recent advances in genetic transformation of forage and turf grasses

Summary Forage and turf grasses are critical to sustainable agriculture and contribute extensively to the world economy. Tremendous progress has been made in genetic transformation of forage and turf grasses in the past decade. The rapid advancement of cellular and molecular biology and transgenic technology provides novel methods to accelerate and complement conventional breeding efforts. This review summarizes the latest developments in genetic transformation methods and the applications of molecular techniques for the improvement of forage and turf grasses.     

中国肉牛分子与基因修饰育种研究进展

随着世界肉牛产业科技的快速发展,我国肉牛产业的整体水平得到明显提高并取得丰硕成果。肉牛育种技术实现了由常规杂交育种向分子标记辅助育种、全基因组选择育种和基因组修饰育种的技术跨越,揭示出大量与生长发育、肉质品质、繁殖与疾病等相关的候选基因与分子标记,并逐步应用于肉牛育种实践。与生长发育性状相关的基因或分子标记主要集中在生长激素基因、生肌调节因子家族、肌肉生长抑制因子和胰岛素样生长因子等;参与肉质形成的基因主要集中在脂肪酸运输与沉积相关信号通路、钙蛋白酶信号通路、生肌调节因子家族与肌肉生长抑制因子等;繁殖性状相关基因或分子标记主要集中在GnRH-FSHR-LH、生长分化因子9、催乳素受体和FoxO1等;抗病相关基因主要有MHC基因家族、TOLL样受体4基因等。目前,利用精准基因编辑技术已培育出促生长发育与提高肉品质的肉牛育种新材料。本文总结了近年来我国在肉牛分子与基因组修饰育种领域取得的研究进展,以期为我国肉牛遗传育种技术研究提供参考和借鉴。     

Large-scale genome-wide association study,using historical data,identifies conserved genetic architecture of cyanogenic glucoside content in cassava (Manihot esculenta Crantz) root

Manihot esculenta (cassava) is a root crop originating from South America that is a major staple in the tropics, including in marginal environments. This study focused on South American and African germplasm and investigated the genetic architecture of hydrogen cyanide (HCN), a major component of root quality. HCN, representing total cyanogenic glucosides, is a plant defense component against herbivory but is also toxic for human consumption. We genotyped 3354 landraces and modern breeding lines originating from 26 Brazilian states and 1389 individuals were phenotypically characterized across multi-year trials for HCN. All plant material was subjected to high-density genotyping using genotyping by sequencing. We performed genome-wide association mapping to characterize the genetic architecture and gene mapping of HCN. Field experiments revealed strong broad- and narrow-sense trait heritability (0.82 and 0.41, respectively). Two major loci were identified, encoding for an ATPase and a MATE protein, and contributing up to 7 and 30% of the HCN concentration in roots, respectively. We developed diagnostic markers for breeding applications, validated trait architecture consistency in African germplasm and investigated further evidence for the domestication of sweet and bitter cassava. Fine genomic characterization revealed: (i) the major role played by vacuolar transporters in regulating HCN content; (ii) the co-domestication of sweet and bitter cassava major alleles are dependent upon geographical zone; and (iii) the major loci allele for high HCN in M. esculenta Crantz seems to originate from its ancestor, M. esculenta subsp. flabellifolia. Taken together, these findings expand our insights into cyanogenic glucosides in cassava roots and its glycosylated derivatives in plants.     

木薯离体培养与快速繁殖

选取3个木薯品种的腋芽作为外植体,分别接种于MS基本培养基上进行离体培养及快速繁殖。结果表明, 6-BA不同浓度对木薯品种不定芽诱导效果不一,在MS+6-BA 0.5~1.0mg/L中,3个品种均可诱导产生幼芽;在MS+6-BA 3.0mg/L中,3个品种的幼芽诱导率高达100%;MS+6-BA 3.0mg/L+NAA 0.1mg/L对继代效果较好;MS+NAA 0.1mg/L对生根效果最佳。     

植物生长调节物质IP－1号对木薯产量及其生物性状的影响

１９９０和１９９１年在木薯（ＭａｎｉｈｏｔｅｓｃｕｌｅｎｔａＣｒａｎｔｚ）生长期以植物生长调节物质ＩＰ－１号０,２０,３０和４０ｐｐｍ进行叶面喷洒,结果表明：３０ｐｐｍ处理可使木薯块根产量平均增加５４．４４％,块根淀粉含量平均提高２０．８１％。单株最大薯重提高３１．５５％,块根数增加２１．１７％,块根长度增长１７．６２％,地上部鲜重增加３４．３６％,植株高度增加４．３６％,植株收获期保留青叶数增加１９．４２％,主茎直径增加６．２６％,块根直径增加２．５８％,叶片的叶绿素和蛋白质含量分别提高５．５７％和２５．９６％,叶片光合作用强度提高１５．８６％,而对主茎高度、主茎节数没有明显影响。     

Comparison of NAA and 2,4-D induced somatic embryogenesis in Cassava

NAA and 2,4-D were compared for their ability to induce somatic embryogenesis in cassava (Manihot esculenta Crantz). In all seven cultivars tested, only 2,4-D had the capacity to induce primary somatic embryos from leaf explants, however, both NAA and 2,4-D were capable of inducing secondary somatic embryos. More secondary somatic embryos were formed in NAA than in 2,4-D medium. Furthermore, the maturation period for secondary somatic embryos was shorter in NAA medium than in 2,4-D medium. In some cultivars, repeated subculture of secondary somatic embryos in NAA medium resulted in a gradual shift from somatic embryogenesis to adventitious root formation. This shift could be stopped and reversed by subculture of the material in 2,4-D medium. In NAA medium the most secondary somatic embryos were formed when they were subcultured every 15 days whereas in 2,4-D a 20 day subculture interval was optimal. Subculture of secondary somatic embryos at a high inoculum density (>1.5 g jar⁻¹) in NAA medium did not result in the formation of secondary somatic embryos, whereas in 2,4-D it lead to the formation of globular secondary somatic embryos. With 2,4-D the newly induced secondary somatic embryos were connected vertically to the explant and with NAA medium horizontally. For all cultivars tested, desiccation stimulated normal germination of NAA-induced somatic embryos. However, the desiccated, secondary somatic embryos required a medium supplemented with BA for high frequency germination. The concentration of BA needed for high frequency germination was higher when the desiccated secondary somatic embryos were cultured in light instead of dark. In only one cultivar desiccation enhanced germination of 2,4-D induced secondary somatic embryos and in three other cultivars it stimulated only root formation. This revised version was published online in June 2006 with corrections to the Cover Date.     

PIG-mediated cassava transformation using positive and negative selection

 In order to develop new selection systems for production of transgenic cassava (Manihot esculenta Crantz), two different selection regimes were assessed for their efficiency on regeneration of transgenic cassava plants: positive selection using mannose and negative selection using hygromycin. Explants from somatic cotyledons and embryogenic suspensions were used as target tissues in the transformation experiments and bombarded using the particle inflow gun. Different culture and selection strategies were assessed to optimise the selection protocols. For the first time transgenic plants could be obtained using positive, and in the case of embryogenic suspensions, hygromycin-based negative selection. The stably transformed nature of the regenerated cassava plant lines and the expression of the transgenes were verified with PCR, RT-PCR, Southern and northern analyses. A rooting test for transgenic plants on a medium supplemented with mannose was developed to further improve the efficacy of the positive selection system. Our results demonstrate that it is possible to obtain transgenic cassava plants using non-antibiotic positive selection. Received: 21 February 2000 / Revision received: 2 May 2000 / Accepted: 5 May 2000     

遗传标记及其在家禽遗传育种中应用的研究进展

概述了现代分子生物学所发展的各类遗传标记的发展和现状及其在家禽遗传育种中的应用,同时也探讨了数量与质量性状基因定位在动物遗传育种中的应用。     

The molecular grouping of Fusarium isolates on cassava in Nigeria

The Fusarium isolates molecularly grouped in this experiment were isolated from the roots of three cassava genotypes (TMS 30572, TMS 4(2)1425 and TME-1) harvested in Ibadan derived savanna, Sabongidda-Ora humid forest and the Onne humid forest regions of Nigeria in 2004 and 2005. Isolates were previously identified using conventional morphological characters at the Plant Pathology Laboratory of the International Institute of Tropical Agriculture, Ibadan. Molecular grouping was done using amplified fragment length polymorphism at the Fusarium Laboratory of the Kansas State University, Kansas, USA. The molecular groupings agreed with the conventional morphological identification. Ten distinct amplified fragment length polymorphism (AFLP) groups of Fusarium were distinguishable, each group probably a distinct species, and many of them might represent previously undescribed Fusarium species. The two largest of the AFLP groups correspond to F. oxysporum and F. solani species complex. F. solani is an important root rot pathogen of cassava Nigeria.     

RAPD分子标记及其在作物遗传育种中的应用

ＲＡＰＤ分子标记是一种新型的分了遗传标记,其原理是采用１０个碱基的随机引物,以基因组ＤＮＡ为模板进行ＰＣＲ随机扩增。其优越性在于其方法简单;分析中的花费少,用时短;分析所需的ＤＮＡ用量也不多等。本文着重介绍以电泳技术和ＰＣＲ扩增技术为核心的分子标记以及在农作物遗传育种中的应用。     

Development of waxy cassava with different Biological and physico-chemical characteristics of starches for industrial applications

The quality of cassava starch, an important trait in cassava breeding programs, determines its applications in various industries. For example, development of waxy (having a low level of amylose) cassava is in demand. Amylose is synthesized by granule-bound starch synthase I (GBSSI) in plants, and therefore, down-regulation of GBSSI expression in cassava might lead to reduced amylose content. We produced 63 transgenic cassava plant lines that express hair-pin dsRNAs homologous to the cassava GBSSI conserved region under the control of the vascular-specific promoter p54/1.0 from cassava (p54/1.0::GBSSI-RNAi) or cauliflower mosaic virus (CaMV) 35S (35S::GBSSI-RNAi). After the screening storage roots and starch granules from field-grown plants with iodine staining, the waxy phenotype was discovered: p54/1.0::GBSSI-RNAi line A8 and 35S::GBSSI-RNAi lines B9, B10, and B23. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed that there was no detectable GBSSI protein in the starch granules of plants with the waxy phenotype. Further, the amylose content of transgenic starches was significantly reduced (<5%) compared with the level in starch granules from the wild-type (about 25%). The inner structure of the waxy starch granules differed from that of the untransformed ones, as revealed by transmission electron microscopy analysis as well as morphological changes in the iodine-starch complex. Endothermic enthalpy was reduced in waxy cassava starches, according to differential scanning calorimeter analysis. Except B9, all waxy starches displayed the A-type X-ray diffraction pattern. Amylogram patterns of the waxy cassava starches were analyzed using a rapid viscosity analyzer and found to have increased values for clarity, peak viscosity, gel breakdown, and swelling index. Setback, consistency, and solubility were notably reduced. Therefore, waxy cassava with novel starch in its storage roots was produced using the biotechnological approach, promoting its industrial utilization.     

柑桔遗传转化技术的研究进展

柑桔是当今世界种植面积最大的果树。遗传转化技术的发展为柑桔育种提供了一条全新的途径。该文就柑桔遗传转化的研究进展 ,包括外源DNA的直接转化与农杆菌介导的转化 ,作一简要的综述 ,并对当前研究中存在的问题及今后的研究方向作了进一步的探讨。     

中国水稻遗传育种历程与展望

我国的水稻育种经历了矮化育种、杂种优势利用和绿色超级稻培育3次飞跃,其间伴随矮化育种(第一次绿色革命)、三系杂交稻培育、二系杂交稻培育、亚种间杂种优势利用、理想株型育种和绿色超级稻培育等6个重要历程。育种目标从唯产量是举到高抗、优质和高产并重,育种理念从高产优质逐步提升为“少投入,多产出,保护环境”。水稻功能基因组研究为第二次绿色革命准备了大量的有重要利用价值的基因,水稻育种正迈向设计育种的新时代。基因组选择技术和转基因技术将为培育“少打农药,少施化肥,节水抗旱,优质高产” 绿色超级稻保驾护航。本文对我国水稻遗传育种的发展历程进行了概括,指出了各种育种方法和育种技术的优缺点,系统介绍了水稻细胞质雄性不育和光温敏雄性核不育以及籼粳杂种不育的分子机制的研究进展,综述了水稻株型、穗型、粒形和养分高效利用相关的重要功能基因,阐明了产量与开花期联动的关系,凸显了我国水稻基础研究在国际上的重要地位。特别指出,近年来,我国水稻生产方式发生了或正在发生巨大变革,育种理念也要与时俱进。未来,杂交育种技术要与现代育种技术紧密结合,选育水稻品种不仅要满足市场需求,而且更要具备绿色健康的特点,同时还要适应新耕作制度和新耕作方法。