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1.
当枸杞的愈伤组织转入分化培养基后的第一天,内源ABA含量就大幅度增高,并成为胚性细胞分化和发育过程中的第一个峰值,也是最高峰值。这时也正是胚性细胞的启动分化期,从愈伤组织的切片明显可见胚性细胞的形成,同时SDS-PAGE结果也表明有相应的35KD蛋白质组分产生,而在未分化的愈伤组织中则未见该蛋白质组分。分化培养后的第15天,内源ABA含量达到第二个峰值,这时是球形胚形成时期,而35KD蛋白质组分含量也在此时达到最大值。外源ABA不仅可诱导内源ABA起始含量的升高,其含量峰值位于分化后的第一天和第15天,同时外源ABA可明显地提高体细胞胚发生的频率和质量。由此表明ABA含量的升高与胚性细胞的启动和发育密切相关,同时内源ABA与外源ABA对体细胞胚的发生有相同的促进作用。其作用机理可能是由于ABA直接或间接激活相关基因表达形成特异性胚性蛋白质组分,从而为胚性细胞的发生与发育奠定了分子基础。  相似文献   

2.
体细胞胚发生的生化基础   总被引:19,自引:0,他引:19  
在胚性细胞分化和分裂过程中ATP酶活性和分布的动态变化表明,这些胚性细胞进行着旺盛的主动物质吸收和活跃的新陈代谢过程。在多种植物的体细胞胚发生中过氧化物酶的活性与同工酶的种类都高于对照,而且在大麦中发现过氧化物酶、酯酶和酸性磷酸酶同工酶的结合应用可以作为体细胞胚发生的标志酶。胚性愈伤组织中可溶性蛋白质含量与组分远高于或多于非胚性愈伤组织。大多数材料中都存在45kD-55kD的胚胎发生特异性蛋白质组分。而且在体细胞胚发生中蛋白质和核酸代谢动态呈规律性变化,首先是RNA合成速率增加,继而是蛋白质的迅速合成,并在胚性细胞分化和发育过程中一直保持相对较高水平,其中mRNA种类丰富,不同发育时期mRNA种类不同,因此转译形成多种蛋白质。DNA的代谢相对较稳定,但在胚性细胞系中DNA合成量仍高于非胚性细胞系。加入蛋白质或核酸合成抑制剂,不仅抑制了蛋白质和核酸的合成,同时也抑制了体细胞胚的发生与发育,而且抑制剂加和时间愈早,影响愈严重。由此表明,蛋白质与核酸的合成为体细胞胚的分化和发育奠定了分子基础。  相似文献   

3.
枸杞体细胞胚发生过程中内源多胺代谢动态的研究   总被引:7,自引:0,他引:7  
完全脱分化的枸杞继代愈伤组织在转入分化培养后第1天就开始启动分化,接着体细胞转变为胚性细胞,继而分裂形成多细胞原胚,球形胚和成熟胚等。与此同时,Put含量迅速上升形成第1个峰值,随后有所下降,但到多细胞原胚期Put含量又上升,并达到最高峰,为对照的6倍,Spd只在胚性细胞分化早期出现,Spm仅在体细胞胚发育晚期存在,外源Put不仅可提高体细胞胚发生频率,而且使3种内源多胺含量均有所提高,加入多胺生物合成抑制剂DFMA后,多胺水平下降,体细胞胚发生几乎完全被抑制。补充外源Put后,多胺的生物合成得到部分恢复,DFMA对体细胞胚发生的抑制效应也被部分解除。结果表明,维持一定量的多胺水平是枸杞体细胞胚发生的必要因素。  相似文献   

4.
本文报道石刁柏胚性愈伤组织的可溶性蛋白质含量与组分、过氧化物酶和酯酶的活力及同工酶带均比其体细胞胚的要少。而在体细胞胚胎发生过程中,过氧化物酶和酯酶活力、可溶性蛋白质含量均以球形胚为最低,子叶分化期胚为最高而呈递增趋势;可溶性蛋白质组分以子叶分化期胚、成熟胚为最多,球形胚、香蕉形胚为最少;过氧化物酶同工酶带以梨形胚为最多,子叶分化期胚、成熟胚为最少;酯酶同工酶则以子叶分化期胚为最多,成熟胚为最少。胚性愈伤组织与体细胞胚均有其特异性可溶性蛋白质及同工酶带,可作为体细胞胚胎发生的分子标记。  相似文献   

5.
以宁夏枸杞无菌苗叶片为材料,对体细胞胚发生过程中激素作用、可溶性蛋白质变化及体细胞胚发生频率进行了研究。发现不同激素处理、可溶性蛋白质含量、组分和体细胞胚发生频率均有一定的差异,三者之间存在着相关性。结果如下:(1)MS_1、MS_2和MS_3三种培养基上继代的愈伤组织难以诱导形成体细胞胚,为非胚性愈伤组织;而从MS_1、MS_2和MS_3分别转到MS_0和MS_4培养基上的愈伤组织体细胞胚发生频率高,属于胚性愈伤组织。其中以MS_3转至MS_0后的体细胞胚发生频率最高。(2)可溶性蛋白质SDS-PAGE分析表明:非胚性愈伤组织有特异性蛋白质67kD,胚性愈伤组织有特异性蛋白质35kD和44kD;蛋白质33kD和67kD受2,4-D调控,蛋白质49kD、57kD受6BA调控,而蛋白质37kD、51kD受2,4-D和6BA协同调控。  相似文献   

6.
研究胡萝卜体细胞胚不同发育阶段的细胞组织化学和蛋白质组成变化的结果表明:胚性愈伤组织主要源自维管束周围的细胞.球胚形成前期,淀粉粒和糊粉粒极性分布已很明显.子叶胚期,芽开始分化,有大量糊粉粒累积.在体细胞胚发育过程中,淀粉粒在胚性愈伤组织形成初期和球胚后期、糊粉粒在胚性愈伤组织形成后期和球胚期各有两次累积高峰.  相似文献   

7.
外源腐胺对石刁柏愈伤组织胚性能力的影响   总被引:1,自引:0,他引:1  
石刁柏胚性愈伤组织继代过程中,添加浓度为10 mg·L^-1的外源腐胺能有效地保持愈伤组织的胚胎发生能力,并减少愈伤组织的褐化,但不能提高愈伤组织的体细胞胚的诱导率.腐胺处理过的胚性愈伤组织的内源腐胺含量明显提高.这可能是外源腐胺保持细胞胚性、降低褐化程度的原因.  相似文献   

8.
小麦幼胚在附加2,4-D 2 mg/L+KT0.5 mg/L+LH 300 mg/L+3%蔗糖的MS 培养基上诱导出愈伤组织,继代2—3次后,降低2,4-D 的浓度,该愈伤组织约以60%的频率转变为胚性愈伤组织,继而形成不同发育期的体细胞胚。这些体细胞胚是起源于愈伤组织表层或近表层的单个胚性细胞,体细胞胚经球形胚、梨形胚、盾片胚、成熟胚而成再生植株。在胚性细胞内淀粉粒大最积累,淀粉颗粒大而密集。随着胚性细胞的分裂,淀粉逐渐被消耗,到多细胞原胚期,胚体细胞内淀粉粒趋于消失。球形胚期淀粉粒又开始积累,但到成熟胚期,淀粉积累只限于生长点区域,尔后随着胚体萌发,淀粉含量又趋于减少。这一消长动态过程与小麦合子胚发生中淀粉代谢动态基本一致。  相似文献   

9.
香雪兰的体细胞胚胎发生可通过两种途径进行,即直接发生与间接发生。在直接发生方式中,体细胞胚直接来源于尚未完全分化的外植体表皮细胞;体细胞胚与母体组织以一种类似胚柄的结构相联系。间接发生方式中,体细胞胚的形成要经过一个愈伤组织阶段。以是否能形成体细胞胚分类,可将愈伤组织分为胚性和非胚性愈伤组织。以间接方式形成的体细胞胚是由胚性愈伤组织中的一种决定细胞发育来的。这种体细胞胚不具有类似胚柄的结构,而与母体组织共同形成一个复合体。体细胞胚具有自己独立的维管束系统,在脱离母体组织后能够独立发育成株。  相似文献   

10.
影响水稻幼穗培养体细胞胚胎发生因素的研究   总被引:6,自引:0,他引:6       下载免费PDF全文
用水稻幼穗为外植体进行组织培养,研究了影响其体细胞胚胎发生的有关因素,建立了高频率体细胞胚胎发生的培养程序。结果表明不同基因型之间体细胞胚胎发生率的差异达到61.2%;生长素2,4-D对诱导水稻体细胞胚起重要的调控作用,细胞分裂素BA有一定的协同促进作用。干燥处理可提高愈伤组织体细胞胚胎发生率,愈伤组织含水量在60%~80%范围的培养效果较好;外源DNA溶液浸泡发育早期的心状体细胞胚.其进一步发育时异常胚的频率显著增加.成苗率降低。  相似文献   

11.
以普通狗牙根[Cynodon dacylon(L.)Pers.cv.'Suncitv']颖果为外植体,以MS为基本培养基,外加浓度在2.0~6.0mg/L的2,4-D,能高频率地诱导出高质量的胚性愈伤组织,其中以4.0 mg/L为最佳.胚性愈伤组织最佳继代及分化的培养方法为:用MS 2,4-D 4.0mg/L继代1~2次,然后转入1/2 MS 2,4-D 2.0 mg/L中继代1~2次,再在无激素的1/2MS中光照培养10 d,最后在MS 6-BA 3.0 mg/L中诱导分化,分化成苗率达31.7%.经电镜观察发现,胚性愈伤组织结构紧密,细胞较小,内容物丰富,而非胚性愈伤组织结构疏松,细胞巨大,内含一大液泡,几无细胞器.  相似文献   

12.
Vitis vinifera L. cv. ‘Manicure Finger’ is one of the major table grape varieties in China. To provide a strong foundation for genetic transformation with potential for crop improvement, we undertook plant regeneration via somatic embryogenesis. Anthers and gynoecia were harvested from immature flowers and used as explants to induce embryogenic calli. Explants cultured in MS1 medium (based on Murashige and Skoog basal salts), supplemented with 4.5-μM 2,4-dichlorophenoxyacetic acid (2,4-D) and 4.4-μM 6-benzylaminopurine (6-BA) showed the highest rates of embryogenic callus induction (3.7%?±?1.3% for anthers and 4.8%?±?2.5% for gynoecia). After several months, somatic embryos were produced from embryogenic calli cultured in plant growth regulator-free MS2 medium (with reduced sucrose). Somatic embryos (SE) at the cotyledonary stage were isolated and cultured on three different media (MS2, MS3, or B) for conversion into plantlets, the efficiency of which ranged from 63.9%?±?4.8% to 83.9%?±?8.4%. After 1 mo of in vitro culture, 80% of plants with at least six leaves were successfully transplanted into soil. SE was repeatedly induced from previously induced somatic embryos for up to 1.5 yr. Using embryogenic calli as starting material, suspension cultures containing embryogenic cell aggregates were also established in liquid MS medium supplemented with 4.5-μM 2,4-D. The embryogenic cell aggregates continued to proliferate without differentiating for successive subculture cycles. After transfer to 2,4-D-free liquid medium for 4 wk, an average of 63.7%?±?9.0% mature SEs were produced per 20 mL of liquid medium. More than 40% of somatic embryos at cotyledonary stage, derived from the suspension cultures, successfully germinated into plants using solid medium.  相似文献   

13.
14.
Somatic embryogenesis (SE) was induced in female flower buds from mature Schisandra chinensis cultivar ‘Hongzhenzhu’. Somatic embryo structures were induced at a low frequency from unopened female flower buds and excised unopened on Murashige and Skoog (MS) agar medium containing 4.0 mg l−1 2,4-dichlorophenoxyacetic acid (2,4-D). Friable embryogenic calli were induced from somatic embryo structures after three to four subcultures on initiation medium. The frequencies of mature somatic embryo germination and plantlet conversion were low, but increased in the presence of gibberellic acid (GA3). Some germinated somatic embryos could form friable embryogenic calli on medium without plant growth regulators (PGRs). The germination and conversion frequencies of somatic embryos from embryogenic calli induced using PGR-free medium were higher than for somatic embryos from embryogenic calli induced on medium containing 2,4-D. Most somatic embryos from 2,4-D-induced embryogenic calli had trumpet-shaped embryos, and most somatic embryos from PGR-free medium–induced embryogenic calli had two or three cotyledons. Histological observation indicated that two- and three-cotyledon embryos had defined shoot primordia, but most of the trumpet-shaped embryos yielded plantlets that lacked or had poorly developed meristem tissue. Cytological and random amplification of polymorphic DNA (RAPD) analyses indicated no evidence of genetic variation in the plantlets of somatic embryo origin.  相似文献   

15.
石防风试管苗的根经2,4-D诱导可形成具有发生体细胞胚潜能的愈伤组织,用愈伤组织制备悬浮细胞。细胞及组织学的观察表明,体细胞胚发生经历了单细胞、丝状体、细胞团、愈伤组织及胚性细胞团的出现及类胚体的各个发育阶段。丝状体可以经过不同的分裂途径发育为细胞团。愈伤组织表面或者内部的某些细胞演变为胚性细胞,它们不断分裂形成了体细胞胚,一个愈伤组织可形成一个或几个体细胞胚。  相似文献   

16.
Compact, friable and embryogenic calli were initiated from immature inflorescences and young leaf bases of one week old seedlings of Paspalum scrobiculatum cultured on MS medium supplemented with 2,4-D. A stable, embryogenic suspension culture was initiated from these calli and maintained in a liquid version of the same MS medium. Embryogenic calli and somatic embryos were obtained by plating suspension culture cells onto semi-solid medium containing 2,4-D. Complete, normal plantlets developed on 2,4-D free medium at a high frequency from somatic embryos. NAA and BAP in the medium promoted plant development.Abbreviations 2,4-D 2,4-dichlorophenoxyacetic acid - NAA 1-naphthaleneacetic acid - BAP 6-benzylaminopurine - ABA Abscisic acid - MS Murashige and Skoog (1962) - CM Coconut milk  相似文献   

17.
The present research aimed to establish conditions for synchronized plantlet regeneration from rice callus based on a quantitative analysis of the relationship between intracellular 2,4-dichlorophenoxyacetic acid (2,4-D) concentration and shoot regeneration rate. To prepare the rice calli with different intracellular 2,4-D concentrations prior to regeneration, callus precultures were carried out in medium containing 4 mg/l 2,4-D and in 2,4-D-free medium for predetermined periods. As the critical intracellular 2,4-D concentration of the calli precultured in 2,4-D-free medium was too low to analyze precisely by conventional analytical methods, it was estimated using a kinetic model which described the behavior of 2,4-D by taking its uptake, metabolism and/or inactivation rates during the callus preculture into consideration. An experimental relationship between intracellular 2,4-D concentration and regeneration rate of rice calli revealed that the intracellular 2,4-D concentration should be controlled as low as 2.6×10–2μg/g fresh weight to reach the same synchronization in shoot regeneration as seen with rice seed germination. This condition was realized by feeding sugar into the 2,4-D-free medium after 4 days preculture when the carbon source was exhausted. Received: 29 June 1998 / Revision received: 24 September 1998 / Accepted: 27 October 1998  相似文献   

18.
Callus cultures were initiated from mature excised caryopses of bahiagrass (Paspalum notatum Flugge) on Murashige & Skoog medium supplemented with 20 gl–1 sucrose and 2 mg l–1 2,4-D. Excised mature caryopses readily germinated and callus developed at the base of coleoptiles. There was considerable variation in the amount of non-embryogenic callus among the cultures. Most of the explants produced non-embryogenic translucent callus consisting of thin-walled cells and unorganized tissue. Some of these calli gave rise only to roots. Other explants formed embryogenic calli which were distinguished morphologically as white, globular and friable. Somatic embryos developed and germinated precociously when embryogenic calli were transferred to a 2,4-D-free medium. Somatic embryogenesis was confirmed by histological sections and scanning electron microscopy. Of the 300 cultures, 35 were embryogenic but only 10 produced plants that were successfully grown to maturity.  相似文献   

19.
We developed a new protocol for highly efficient somatic embryogenesis and plantlet conversion of Schisandra chinensis. Friable embryogenic callus was induced from cotyledonary leaves and hypocotyls of germinated zygotic embryos on Murashige and Skoog (MS) agar medium containing 2,4-dichlorophenoxyacetic acid (2,4-D). Preculture of zygotic embryos on 2,4-D-containing medium increased embryogenic callus induction efficiency. The highest embryogenic callus induction frequency of 56.7% was obtained from shoot apical meristem-containing hypocotyl explants from 1-week-old germinated embryos on MS medium containing 4.0 mg l−1 2,4-D. Embryogenic callus proliferation, somatic embryo (SE) formation, and subsequent plantlet conversion occurred under optimal culture conditions. The effects of MS medium strength, sucrose, gibberellic acid (GA3), and 6-benzyladenine (BA) on SE formation and plantlet conversion were evaluated. Low MS medium strength (1/4 to 1/2) was necessary for SE formation, and the optimal sucrose concentration was 2.0%. Supplementing medium with GA3 negatively impacted SE formation and subsequent development. BA significantly increased the number of SEs and the plantlet conversion capacity. One-third-strength MS medium with 1.0% sucrose and 0.5 mg l−1 BA produced the highest number of SEs (309 embryos from 9 mg embryogenic callus) and the highest frequency of plantlet conversion from germinated SEs (52.6%). When transplanted to soil, 90% of the regenerated plants developed into normal plants.  相似文献   

20.
The embryogenic calli (EC) were obtained from hypocotyl explants of groundnut (Arachis hypogaea L.) cultured on Murashige and Skoog (MS) medium supplemented with different concentrations of 2,4-dichlorophenoxyacetic acid (2,4-D) in combination with 0.5 mg dm−3 6-benzylaminopurine (BAP). The EC were exposed to γ-radiation (10–50 Gy) or treated with 1–5 mM of ethyl methane sulphonate (EMS) or sodium azide (SA). The mutated EC were subcultured on embryo induction medium containing 20 mg dm−3 2,4-D. Somatic embryos (SE) developed from these calli were transferred to MS medium supplemented with BAP (2.0 mg dm−3) and 0.5 mg dm−3 2,4-D for maturation. The well-developed embryos were cultured on germination medium consisting of MS salts with 2.0 mg dm−3 BAP and 0.25 mg dm−3 naphthaleneacetic acid (NAA). Well-developed plantlets were transferred for hardening and hardened plants produced normal flowers and set viable seeds. The fresh mass of the EC, mean number of SE per explant and regeneration percentage were higher at lower concentrations of mutagens (up to 30 Gy/3 mM). Some abnormalities in regenerated plants were observed, especially variations in leaf shape.  相似文献   

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