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采用流式细胞仪分析柑橘愈伤组织的细胞周期 总被引：1，自引：0，他引：1
细胞周期分析可为研究柑橘愈伤组织理化诱变提供依据。试验以暗柳橙、长沙橘、澳洲指橘、纳维来特脐橙等4种培养多年的柑橘愈伤组织为试材,使用流式细胞仪分析其特定时间段的细胞周期,进而找到柑橘愈伤组织理化诱变的最佳时期即分裂最旺盛时期。结果表明,待测4种基因型柑橘愈伤组织在继代2周时处于分裂最旺盛时期,它们的S期时相比例分别为22.4%、21.9%、21.0%和22.9%。同时发现,待测的4种柑橘愈伤组织生长周期长短与再生能力有一定的相关性。极易再生的暗柳橙生长周期最短,易再生的长沙橘、澳洲指橘次之,丧失再生能力的纳维来特脐橙生长周期最长。 相似文献
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以转GFP国庆1号温州蜜柑愈伤组织为试验材料,研究了不同浓度NaCl处理对转GFP国庆1号温州蜜柑愈伤组织增殖量及愈伤组织可溶性蛋白含量、可溶性糖含量、脯氨酸含量、SOD活性、CAT活性的影响。结果表明,随着NaCl浓度的增大,愈伤组织的增殖量、SOD活性、CAT治洼下降;可溶性蛋白含量和脯氨酸含量增加。 相似文献
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以耐寒柑橘子叶为试验材料,研究了不同浓度的2,4-D、NAA、6-BA与KT对柑橘子叶愈伤组织诱导的影响作用。结果表明:2,4-D、NAA、6-BA与KT对柑橘子叶愈伤组织诱导具有重要效应,其综合贡献率分别为86.4%、93.9%、93.9%与94.5%。4种生长调节因子对柑橘子叶愈伤组织诱导的膨大率、膨大速度、出愈率、出愈速度的效应大小分别为2,4-DNAA6-BAKT、KT2,4-DNAA6-BA、2,4-DNAA6-BAKT与2,4-DKTNAA6-BA。综合考虑测量指标效应,对于柑橘子叶愈伤组织诱导的最适生长调节因子组合配比为2,4-D 1.0mg/L+NAA 2.0mg/L+6-BA 0.5mg/L+KT 0.5mg/L。 相似文献
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以11份不同基因型胡萝卜为试材进行游离小孢子培养研究,并对胚状体及愈伤组织形成过程进行细胞学观察。诱导培养92 d后,有5份材料形成肉眼可见的胚状体或愈伤组织。其中70198和80Q54均诱导出胚状体和愈伤组织,胚状体产率分别为每皿10.56和7.00个,愈伤组织产率分别为每皿3.56和8.38个;90234仅形成胚状体,产率为每皿6.75个;70Q78和80Q52仅产生愈伤组织,产率分别为每皿3.00和1.00个。细胞学观察表明胚状体和愈伤组织形成过程具有完全不同的结构特征。发育成胚状体的小孢子细胞壁变薄,膨大伸长,长度可至原来的1.5 ~ 4.0倍,有明显大液泡,初期细胞纵向分裂,成串排列,细胞之间紧密连接,而发育成愈伤组织的小孢子膨大成球状,多次分裂后松散地连接在一起。通过流式细胞仪鉴定70198、70Q78和80Q54获得的137株再生植株倍性,单倍体、二倍体及三倍体所占比例分别为68.6%、29.9%和1.5%。 相似文献
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《果树学报》2013,(6)
【目的】为解决基因型依赖性比较强的苹果品种遗传转化困难等问题,研究不同伤害处理方式下苹果茎尖碎片的再生能力及其对苹果转化率的影响。【方法】以苹果‘姬神’试管苗为试材,运用农杆菌介导法对苹果茎尖和离体叶片进行遗传转化。【结果】将茎尖纵切为4部分时,可以高比率的形成愈伤组织(88.4%)和再生不定芽(59.8%)。在20 mg·L-1的卡那霉素筛选压下,300个茎尖外植体中有50.4%的茎尖碎片形成愈伤组织并获得了19株表达GFP的转基因植株。而在同样的筛选压下有22.6%农杆菌介导的离体叶片形成愈伤组织,但不能获得转基因植株。降低筛选压可以提高植株再生率,但其中包含大量嵌合体,转化效率并没有明显提高。【结论】苹果茎尖做为转化受体基因型依赖性较低,要优于苹果离体叶片。 相似文献
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荔枝体胚发生过程中的细胞学观察 总被引：2，自引：0，他引：2
以荔枝三月红胚性愈伤组织为材料,采用改良的石蜡切片法观察荔枝胚性愈伤组织结构及体胚发生过程,结果显示荔枝的体胚发生经历球形、心形、鱼雷形、子叶形4个过程;荔枝在胚性细胞分裂发育形成体胚之前,出现与周围细胞明显不同的界限,形成细胞间隔离;比较荔枝胚性愈伤组织与非胚性愈伤组织的细胞学特性,发现胚性愈伤组织具有核大、质浓、染色深、细胞排列紧密等特点,与非胚性愈伤组织明显不同。通过系统观察荔枝胚性愈伤组织结构以及胚性细胞分裂、生长方式和体胚发生过程,为胚性愈伤组织的利用提供细胞组织学依据。 相似文献
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R. M. Perez A. M. Galiana L. Navarro 《The Journal of Horticultural Science and Biotechnology》2013,88(6):419-429
SummaryA number of experiments were performed to obtain somatic embryos and embryogenie cell lines from several species and cultivars of citrus by in vitro ovule culture. The source of ovules (pre-anthesis and post-anthesis), incubation conditions (light and darkness) and several modifications of the culture medium were evaluated. All the species and cultivars assayed produced somatic embryos and “pseudobulbi”, but production of friable embryogenie callus was species dependent. The small proliferation of callus and globular bodies recovered from the primary cultures were suitable for recovering embryogenie callus lines by periodical subculturing into fresh medium. After several monthly subcultures discarding embryos and “pseudobulbi”, cell lines virtually devoid of organized structures were obtained from three cultivars of sweet orange, sour orange and Cleopatra mandarin. In all these species, embryogenesis was restored spontaneously and/or by substitution of sucrose by maltose or lactose in the culture medium. The embryos germinated and produced phenotypically normal plants. 相似文献
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【目的】为了快速有效地利用基因工程的方法获得柑橘无核新种质,【方法】以我国特色多核柑橘优良品种锦橙实生苗上胚轴切段为外植体,采用根癌农杆菌介导法进行能导致种子败育基因CG1-400-RNase的转化;为快速有效地筛选出转化子,在实生苗上胚轴切段转化再生过程中,根据不同发育阶段的组织或器官对抗生素的敏感程度不同采用不同的选择压。【结果】结果表明,在抗性芽再生过程中卡那霉素质量浓度设定为50 mg.L-1,获得的362个抗性芽转入卡那霉素质量浓度为100 mg.L-1的伸长培养基中进行伸长培养后,进行早期PCR检测,获得28个阳性芽;经过不定芽诱导生根或试管嫁接,获得22株完整植株。【结论】再生植株经PCR和Southern杂交检测,获得2株目的基因以单拷贝的形式插入锦橙基因组的转基因植株,为最终获得具有无核性状且可稳定遗传的柑橘新种质奠定了基础。 相似文献
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以仙客来（Cyclamen persicum Mill.）开花植株的新生叶片为外植体，诱导筛选胚性愈伤组织，并使其进一步发育成体细胞胚。组织切片观察结果表明：胚性愈伤组织由胚性和非胚性细胞组成，胚性细胞多以胚性细胞团的形式存在，胚性细胞团起源于诱导的胚性决定细胞。体细胞胚起源于单个胚性细胞，经多细胞原胚、球形胚、心形胚和鱼雷胚等时期发育成完整植株。在体细胞胚的发生发育过程中，淀粉粒出现4次积累高峰，分别为胚性细胞、球形胚、早期鱼雷胚和成熟胚发育成完整的小植株时期，淀粉代谢与体细胞胚发生、发育及小植株的形态建成密切相关。 相似文献
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This study describes a successful method of somatic embryogenesis and genetic transformation using immature cotyledons of Prunus mume. Immature cotyledons from four different developmental stages of eight different P. mume cultivars were used for the experiments to optimize somatic embryogenesis and genetic transformation protocols. Somatic embryogenesis was induced when the explants were cultured on somatic embryo inducing medium consisting of MS basic medium supplemented with 1 μM 2,4-dichlorophenoxyacetic acid (2,4-D) and 1 μM 6-benzyladenine (BA). They were cultured for 30 days and then transferred to somatic embryo propagation medium containing 0.1 μM α-naphthaleneacetic acid (NAA) and 5 μM BA. It appeared that the developmental stage of the immature cotyledons used as explants was the most important factor for somatic embryogenesis; higher frequencies of somatic embryogenesis were observed when the immature cotyledons were less than 5 mm in length regardless of cultivars. For genetic transformation, the immature cotyledons were inoculated with Agrobacterium tumefaciens EHA101 harbouring a binary plasmid vector with neomycin phosphotransferase II and an intron-interrupted β-glucuronidase gene under the control of cauliflower mosaic virus 35S promoter, and three transgenic plant lines were obtained from inoculated “Sirakaga” immature cotyledons. Transgenic somatic embryos and shoots were selected using 25 mg l−1 kanamycin. Integration of transgenes in the genome of GUS-positive putative transgenic shoots was confirmed by PCR and Southern blot analyses. 相似文献
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Yongrui He Shanchun Chen Aihong Peng Xiuping Zou Lanzheng Xu Tiangang Lei Xiaofeng Liu Lixiao Yao 《Scientia Horticulturae》2011
Transgenic plants of Jincheng orange (Citrus sinensis Osbeck) and Newhall navel orange (C. sinensis Osbeck) containing antibacterial peptide genes Shiva A and Cecropin B were successfully obtained by a novel Agrobacterium tumefaciens-mediated transformation of the mature axillary buds. PCR and Southern blot analysis of the transgenic plants verified that the Shiva A and Cecropin B genes were integrated into the citrus genome. The transgenic plants began to blossom and bear fruit in the 2nd year after grafting on trifoliate orange (Poncirus trifoliata Raf) rootstock in greenhouse. Water-soluble extracts from transgenic citrus leaves exhibit in vitro suppressive effects on Xanthomonas axonopodis pv. citri, suggesting that the expressed products of Shiva A and Cecropin B in citrus retain their native antibacterial activities. Artificial inoculation in greenhouse and open field further indicates significantly increased resistance of transgenic plants to X. axonopodis pv. citri when compared with non-transgenic lines. No significant difference was found in the content of total soluble solids, total acidity, reduced sugar content and other fruit characteristics between transgenic and non-transgenic plants. In this present study, 11 transgenic lines were obtained from 40 transgenic lines, showing enhanced resistance to citrus canker disease. 相似文献
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《Scientia Horticulturae》2005,105(1):117-126
The objectives of the present work were to study the embryogenic competence of floral tissues of Feijoa sellowiana and to investigate the influence of plant growth regulators on somatic embryo induction and development in order to establish a somatic embryogenesis protocol starting from somatic tissues. Petals, stamens and ovaries of floral buds were cultivated onto LPm basal medium supplemented with different levels of 2,4-D, Picloram, 2-iP, Kin and BAP. The highest embryogenic callus induction was obtained with Picloram (10 μM) and Kin (1 μM). Rates of embryogenic calluses induction in stamens and petals were significantly affected by PGRs. Embryogenic calluses were transferred to the same medium, supplemented with gradually reduced levels of PGRs-free medium. After 60 days in suspension cultures with 2,4-D (1 μM) and 2-iP (1 μM) calluses were transferred to PGR-free medium. After 30 days it was observed the development of globular somatic embryos on the surface of 18% of friable calluses previously induced with Picloram (10 μM) and Kin (1 μM). Only embryogenic calluses derived from stamens gave rise to this morphogenetic pattern.Torpedo and cotyledonary somatic embryos transferred to PGR-free culture medium were converted to complete plantlets. This is the first report of somatic embryogenesis in this species starting from somatic tissues. 相似文献
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选用‘椪柑’、‘锦橙’、‘桃叶橙’、‘华农本地早’花后7周幼果胚珠进行培养.其胚状体发生的百分率分别是:48.5%、21.2%、19.8%、6.4%;胚性愈伤组织则为:28.2%、13.5%、15.8%、5.8%.各品种胚状体和胚性愈伤组织发生的百分率明显不同.与品种胚数多少相一致.不同激素和有机物对柑桔胚珠培养、胚状体的发生及胚性愈伤组织形成产生不同的影响.多单胚的‘华农本地早’仅从基本培养基中产生胚状体和胚性愈伤组织.胚珠培养中,胚状体发生的方式,根据来源不同分为三种:第一种是由胚珠(或已黄的球形胚上)长出白色的胚性愈伤组织,继而产生胚状体:第二种是在已分化的幼苗下胚轴或根部产生胚性愈伤组织和胚状体;第三种是在畸形胚上直接不断的分化出胚状体. 相似文献
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为了利用Cecropin B（CB）抗菌肽基因提高柑橘对溃疡病的抗性,人工合成了两个含有信
号肽的Cecropin B 抗菌肽基因PR1aCB 和AATCB。洋葱表皮瞬时表达分析表明,与非分泌型CB 抗菌肽
相比,PR1aCB 和AATCB 抗菌肽在细胞间隙中优势积累。进一步构建了CaMV 35S 调控 PR1aCB 和AATCB
基因的植物表达载体,转化塔罗科血橙（Citrus sinensis Osbeck）上胚轴,获得转基因植株。GUS 组织化
学染色、PCR 和Southern blot 分析表明外源基因已成功整合入柑橘基因组。Real-time PCR 定量分析表明,
抗菌肽基因在转基因植株中成功表达。柑橘叶片离体抗性分析表明,转PR1aCB 和AATCB 基因植株的抗
性显著强于非转基因植株,其抗性水平与高抗品种‘南丰蜜橘’（C. succosa Hort. Ex Tan）和‘四季橘’
（C. madurensis）相当。 相似文献
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不同类型柑橘中黄脉病毒衣壳蛋白基因的保守性及复制水平 总被引：1，自引：0，他引：1
为研究不同柑橘品种对柑橘黄脉病毒(Citrus yellow vein clearing virus,CYVCV)变异的影响,将CYVCV毒株CQ01嫁接接种于4类共35个柑橘品种,对不同植株中CYVCV的衣壳蛋白(CP)基因进行分析,结果发现仅有少数位点发生了变异。运用RT-qPCR技术检测不同柑橘品种中CYVCV的相对含量,发现甜橙类品种中CYVCV的相对含量最高,墨西哥来檬中CYVCV的相对含量最低。植株中CYVCV的含量与其症状不存在明显的关联。 相似文献