植物体细胞胚胎发生机理的研究进展

 体细胞胚胎的诱导是大多数植物组织培养中的关键环节。在愈伤组织分化胚性愈伤组织的过程中,随着细胞功能的转变,细胞内部基因的差异表达,引起了植物体内蛋白质组分、酶组分与活性、激素等各方面的变化,这些变化一直是组织培养研究中的热点。本文对体细胞胚胎的氨基酸变化、标记性蛋白、基因的差异表达、同工酶变化、激素动态和高体细胞胚胎发生率的材料遗传与选育等方面的研究进展作了综述。     

内源激素对黄姜体细胞胚发生的影响

以不同的培养基诱导外植体以建立优良的体细胞胚胎发生系统,并结合其发育过程中内源激素含量的动态变化,进一步探讨黄姜体细胞胚胎发生机理。选择2a生新鲜优良盾叶薯蓣41号,取以腋芽萌发而来的无菌枝条切段,采用MS+BA 1.5 mg/L+NAA 0.5 mg/L、LS+BA1.5mg/L和l/2 MS+NAA 1.0mg/L为诱导培养基,对培养过程中的内源激素的动态变化进行了分析。结果表明,高含量的生长素（IAA）、细胞分裂素（Z、ZR）和赤霉酸（GA3）有利于胚性细胞的形成。可为黄姜愈伤组织及体细胞胚胎形成的调控作用提供理论的参考依据,也可为其它植物的组织培养提供参考。     

绿色棉新彩棉7号体细胞胚胎发生及其植株再生

以绿色棉新彩棉7号的子叶、下胚轴为外植体,MSB(MS培养基附加B5维生素)基本培养基附加不同激素组合,诱导愈伤组织及调控分化,通过体细胞胚胎发生方式获得再生植株.结果表明:0.1 mg· L-1 KT(Kinetin,激动素)+ 0.1 mg·L-12,4-D(2,4-dichlorophenoxyacetic,2,4-二氯苯氧乙酸)为诱导愈伤组织的最适植物激素组合,不同外植体处理出愈率均达到100％,但下胚轴纵切面背向培养基放置培养更有利于诱导愈伤组织形成;分化调控阶段的最佳植物激素组合为0.15 mg·L-1 KT+ 0.3 mg·L-1 IBA(Indole-3-butyric acid,吲哚丁酸),胚性愈伤分化率可达23.33％; MSB中去除NH4NO3同时KNO3加倍,附加0.5 g·L-1Asn(Asparagine,天冬酰胺)和lg· L-1 Gln(Glutamine,谷氨酰胺),胚性愈伤可进一步分化获得体细胞胚,将成熟的子叶胚接种于1/2MS获得完整的再生植株. 本研究通过体细胞胚发生途径获得了新彩棉7号的再生植株,为天然彩色棉基因工程研究奠定了一定基础.     

TDZ在棉花组织培养中的效应研究Ⅰ．TDZ单独使用对棉花组织培养的影响

ＴＤＺ是一种高效低毒棉花脱叶剂，它在棉花组织培养中表现出独特的生理作用。ＴＤＺ的添加促进了外植体的伸长和保绿，有利于外植体脱分化和胚性愈伤组织的诱导。但是ＴＤＺ不利于愈伤组织的保持和体细胞胚的发育，必须把在含ＴＤＺ培养基上诱导产生的愈伤组织转移到不含任何激素的培养基上，才能发育成成熟胚并再生出植株。ＴＤＺ的使用能诱导根的分化。不同外植体、不同基因型以及不同ＴＤＺ浓度之间在棉花组织培养中的表现均有差异。     

植物激素面面观

正植物激素是指植物细胞接受特定环境信号诱导产生的、低浓度时可调节植物生理反应的活性物质。它们在细胞分裂与伸长、组织与器官分化、开花与结实、成熟与衰老、休眠与萌发以及离体组织培养等方面,分别或相互协调地调控植物的生长、发育与分化。这种调节的灵活性和多样性,可通过使用外源激素或人工合成植物生长调节剂的浓度与配比变化,进而改变内源激素水平     

棉花体细胞胚胎发生和植株再生的影响因素

对影响棉花体细胞胚胎发生和植株再生的基因型、外植体及培养基中的碳源、氮源、固化剂、外源激素、pH值等方面的因素进行了综述,并讨论了存在的问题及今后的研究方向。     

棉花优良品种中棉所19高频体细胞胚胎发生细胞系筛选与植株再生

选用我国优良棉花品种中棉所19进行组织培养, 研究了该品种的体细胞胚胎发生和植株再生能力。 中棉所19的体细胞胚胎发生有两种情况： 一是由外植体直接诱导获得胚性愈伤组织和体细胞胚; 二是先诱导获得愈伤组织, 再经继代培养获得胚胎发生。 单独使用ZT可在早期诱导胚胎发生, 在该情况下, 对子叶的诱导能力最强, 胚根     

两种常用激素组合下棉花体细胞胚胎发生过程的组织学观察

 MSB培养基添加两种常用植物激素组合，Indole-3-butyric acid (IBA) + kinetin (KT)和2, 4-dichlorophenoxyacetic acid (2, 4-D) + KT，分别命名为IK和DK处理用来诱导陆地棉体细胞胚胎发生。陆地棉品系YZ1下胚轴切段作为外植体在诱导培养基上培养4周后，继代到分化培养基上促进体细胞胚胎发生。结果表明，两种处理均有体细胞胚胎发生，其中IK处理上外植体嫁接33 d后就可见体细胞胚出现，分化率高达97.6%，大多数外植体表现为体细胞胚较胚性愈伤早出现。DK处理中体细胞胚胎发生慢，体细胞胚都是经过明显的胚性愈伤发育而来，胚性愈伤分化率明显低于IK处理，仅为28.6%。此外，IK处理中的外植体在培养过程中大部分都出现了须根，而DK处理中则没有。两种处理中出现了两种不同的体细胞胚胎发生过程，组织学观察结果表明DK和IK处理中的体细胞胚分别主要起源于初生形成层和皮层。     

大豆组织和细胞培养研究进展

从大豆组织培养的培养基选择、基因型和外植体类型培养差异、器官和胚胎发生再生植株、花药和花粉及 原生的南体培养再生植株、体细胞无性系变异体选择和基因转化等方面综述了大豆作物组织和细胞培养所取得的进展，并对今后的研究提出了建议。     

长江和黄河流域棉区棉花品种体细胞胚胎发生和植株再生比较研究

选用我国长江和黄河流域棉区各10个棉花品种以及珂字201和YZ1共22个基因型,研究和优化棉花体细胞胚胎发生的相关条件参数。在此基础上,比较了两生态区棉花品种的体细胞再生能力。结果表明, 在IBA+KT的激素组合下,多数基因型有较强再生能力;在愈伤组织诱导时期,铵态氮是必需的,而在愈伤组织继代分化时期,一定浓度的硝态氮则能促进分化;没有铁盐不能诱导出愈伤组织,而铁盐浓度为56 mg L^-1时有利于胚分化;长江流域和黄河流域棉花品种的植株再生潜力没有明显差别,但长江流域的品种体细胞胚胎发生所需的时间长。本文首次获得了鄂抗棉3号、5号、鄂棉20、鄂棉23、豫棉9号、豫早73、豫棉12、豫棉1221等8个品种的体细胞胚胎发生和植株再生。     

毛白杨PtAUX1超表达诱导体状胚发生

体状胚的发生由一个生长素信号级联系统控制,其发生过程需要大量的生长素,随后在去除生长素或降低生长素浓度的条件下,体状胚趋于成熟。除植物激素外,环境胁迫也是体状胚发生所必须的,激素和环境胁迫协同诱导细胞的脱分化、再分化和体状胚的发生。目前发现的参与体状胚发生的主要基因有SERK、PKL和BBM等,它们作用于体状胚的不同发育阶段。本研究中,在毛白杨35S::PtAUX1超表达植株中发现叶片表面产生大量体状胚,体状胚分散在叶片主脉及侧脉两侧或聚集在叶片边缘,其中部分体状胚可发育成成熟体状胚。将成熟体状胚取下,置于MS0培养基上进行培养,体状胚发芽并缓慢产生根。研究结果表明,PtAUX1通过建立新的生长素浓度梯度,诱导体细胞转化成胚性细胞,从而使叶片产生出体状胚。除上述基因外,PtAUX1也参与体状胚的发生,这一研究发现现尚未见报道。     

伞形科根茎类药用植物早期抽薹的研究进展

通过简述伞形科根茎类药用植物发生早期抽薹现象的过程,明确伞形科根茎类药用植物发生早期抽薹的原因和影响因素,推测早期抽薹与某些调控开花的基因表达能力有关。另外,认为植物体内激素合成在一定程度上能够调节早期抽薹现象的发生。植物内源激素的合成、代谢、信号传导等过程特定基因发生差异性表达导致植物发生早期抽薹现象。提出可以从植物内源激素分子基因层面及激素作用机理出发,通过改变伞形科根茎类药用植物生长条件或施加外源性激素等人工手段有效抑制早薹率,这将对提高药用产量和品质有重要意义。     

大豆组织培养再生系统的研究进展

大豆因其愈伤组织难以分化、原生质体再生困难等因素,其再生体系一直不够完善。直到20世纪80年代初期,才成功建立了大豆的体细胞胚胎发生和器官发生再生系统。80年代末期原生质体培养获得突破。介绍了大豆器官发生再生系统、体细胞胚胎再生系统和原生质体再生系统的研究进展;对这3种再生系统进行遗传转化的优缺点作了比较;并提出了关于大豆遗传转化再生系统的几点展望。     

Somatic embryogenesis from floral tissue of cassava (Manihot esculenta Crantz)

The aim of this study was to examine the embryogenic potential of floral material of the cassava cultivar MCOL 1505. Macerated immature inflorescences were found to be highly embryogenic, with almost 78% of the explants producing somatic embryos. Somatic embryos were also produced from whole male florets and half florets although at much lower rates. No regeneration was obtained from anther, microspore or floret wall tissue. Somatic embryos derived from immature inflorescences were regenerated via organogenesis and the plants derived from this process were assessed in terms of phenotype and ploidy level. If haploid plants could be produced by this method, this would have significant implications in assisting traditional cassava breeding, as this would allow homozygosity to be reached more rapidly. In a crop such as cassava, which is highly heterozygous in nature, the use of haploids in a breeding programme could considerably shorten the time taken to produce new desirable cultivars. This is the first report on plant regeneration through somatic embryogenesis from floral tissue of cassava. This revised version was published online in July 2006 with corrections to the Cover Date.     

Plant regeneration via organogenesis and somatic embryogenesis in callus cultures derived from mature zygotic embryos of leek (Allium ampeloprasum L.)

Summary A high frequency plant regeneration system via organogenesis and somatic embryogenesis was established with callus cultures derived from mature zygotic embryos of different leek genotypes (Allium ampeloprasum L.). Four different callus types with varying morphogenetic potential were obtained. Relatively high concentrations of the auxin 2,4-dichlorophenoxy-acetic acid reduced callus weight and subsequent shoot regeneration and primordia formation of the callus. Shoot regeneration and primordia formation of the callus decreased after prolonged subculture on media containing 2,4-dichlorophenoxy acetic acid. A callus growth period of six weeks on Murashige and Skoog medium with 0.25–0.5 mg l^-1 2,4-dichlorophenoxy acetic acid showed the highest rate of shoot regeneration after transfer of callus to regeneration medium with 1 mg l^-1 kinetin.Differences between leek genotypes in callus type, callus weight, shoot regeneration and primordia formation were observed. Histological observations showed that plant regeneration took place, both via the pathway of somatic embryogenesis and organogenesis.Abbreviation 2,4-D 2,4-dichlorophenoxy acetic acid - MS Murashige and Skoog (1962) medium     

The HMW Glutenin Subunit and Gliadin Compositions of German-Grown Wheat Varieties and their Relationship with Bread-Making Quality

An in vitro system for the initiation of somatic embryogenesis and plant regeneration of spring- and winter-type rye has been developed. The optimal developmental stage of immature embryos to be used as explants is the late stage of spherical coleoptile when the tissue turns milky in colour and the coleoptile is enlarged but spherical. The type and concentration of auxin in the induction medium is of importance for obtaining high efficiency of somatic embryogenesis. CC-medium high 30 μM Dicamba resulted in well developed somatic embryos in large umbers. Positive effects on the efficiency, intensity and speed of development of somatic embryos have been observed by the addition of coconut water to CC-medium, replacing agar by agarose and culturing the explants in low light intensity. Following a “step by step” optimization, culture conditions have been defined giving rise to a 90—100% efficiency of somatic embryogenesis and a high number of regenerated plants.     

Plant tissue culture of Secale: A review

Summary Plant tissue culture of rye employs different parts of plant bodies originating from various stages of rye ontogenesis. For the culture initiation mainly diploid and after that tetraploid forms were used, however, haploids and triploids were subject of investigation. The development of plant regeneration via somatic embryogenesis or organogenesis required the using of numerous basal media with various plant growth hormones. Haploidization appeared to be the most difficult problem and only interspecies hybridization helped to overcome this problem. Wide sexual intergenera and interspecies hybridization in cereals require the development of proembryo and immature embryo culture system. Using the nurse culture basing on immature endosperm resulted in getting fully formed rye plants. There is no progress in somatic cell genetic manipulation of rye because of lack of plant regeneration system in mesophyll green leaf and suspension protoplast cultures.Abbreviations 2, 4, 5, Cl₃POP... 2, 4, 5-Cl₃-phenoxypropionic acid, dicamba...-3, 6-dichloroasinic acid - GA₃... gibberellic acid - BA... benzyladenine - IBA... 3-indolebutric acid - IAA... indoleacetic acid - NAA... 1-naphtylacetic acid - 2, 4-D... 2, 4-dichlorophenoxyacetic acid - 2, 4, 5-T... 2, 4, 5-trichloroacetic acid - p-CPA... parachlorophenoxyacetic acid - CM... coconut milk - CW... deproteined coconut milk - Kin Kinetin     

Micropropagation of mother plants of lucerne (Medicago sativa L.) for somatic embryogenesis

Summary A reliable and standard method was established for micropropagation of the A70-34 selected genotype of lecerne (Medicago sativa L., genotype A70-34), aimed at reducing contamination problems, seasonal and phenological influences on regeneration and phytosanitary problems of the mother plants, while maintaining the regenerative potential for somatic embryogenesis of the plant tissues. Mother plants were routinely maintained for several subcultures and somatic embryogenesis was regularly obtained from the subcultured explants. Proliferation, rooting and embryogenetic ability of plants cultured for 30 days was greater than those cultured for 20 days. The regenerative potential of tissues from different organs and of triturated and intact whole plants was also tested. Petioles were confirmed as the best source for embryogenesis as far as efficiency and repeatability were concerned, even though regeneration from other explant types was also achieved. Production of somatic embryos through mechanical trituration of the in vitro cultured plants was obtained; the regeneration ability of the triturated plants was greater than that observed in the intact plants.     

花生体细胞胚诱导及植株再生研究

为了建立花生体细胞胚诱导及植株再生体系,为花生分子育种提供技术支撑,以花生品种‘桂花30’和‘桂花771’为材料,采用预培养3天的种子胚小叶、下胚轴、子叶节为外植体,在添加外源激素的MS培养基中使体细胞脱分化形成体细胞胚,再分化成植株。结果表明,在所设定2,4-D浓度（3,5,10,15,20 mg/L）范围内,胚小叶最容易诱导形成体细胞胚,2,4-D的适宜浓度为10 mg/L,经过约30天培养,可产生大量体细胞胚,‘桂花30’和‘桂花771’的平均诱导率分别为55.37%和36.72%。平均每个外植体产胚量分别为5.68个和4.27个。将诱导形成的体细胞胚转接到6-BA浓度由5 mg/L逐渐降低到1.5 mg/L的MS培养基中,体细胞胚萌发再生成无根小苗,正常植株再生率‘桂花30’为32.6%,‘桂花771’为23.5%。将无根苗转接到生根培养基中可获得完整植株。花生是较难诱导体细胞胚形成的作物之一,筛选合适的基因型、外植体和激素浓度是获得较高体细胞胚发生率和植株再生率的关键技术。     

Improving in vitro induction of autopolyploidy in grapevine seedless cultivars

The efficiency of in vitro polyploidization depends on several variables associated to the plant, the antimicrotubule agent and the interactions between them. In the present work, we have used response-surface methodology to determine the best operating conditions for plant recovery in polyploidization assays for shoot apices and somatic embryos of two seedless grape cultivars, employing colchicine and oryzalin. Explant type, tubulin-interfering compound and concentration were the critical factors determining plant recovery. Linear reduction in viable plant regeneration via organogenesis and somatic embryogenesis was obtained by increasing oryzalin concentrations and treatment time, whereas the effects of colchicine were better described by a quadratic design for both explants types. The conditions promoting higher rates of plant recovery were used in chromosome doubling experiments with oryzalin and colchicine for shoot apices and somatic embryos of ‘Crimson seedless’ and ‘BRS Clara’. The established protocols allowed the recovery of non-chimerical autotetraploid plants at rates higher than 30 % for both cultivars. Stomata size parameters statistically correlate to the ploidy level of the regenerants and were effective for preliminary polyploidy screening. Autotetraploid lines of seedless grapes were incorporated into the Vitis germplasm bank for further agronomical evaluations. To our knowledge, this is the first report of in vitro oryzalin induced polyploidization of grapevine and of the use of mathematical modeling to optimize chromosome doubling in plants.