毛白杨叶外植体的遗传转化

参考 Horsch 等(1985)建立的叶圆片转化法,建立了一个适合于杨树的遗传转化系统。本文所用的根癌农杆菌菌株含有一个经过改建的 Ti 质粒,这个质粒载有 T-DNA 的4号基因和 CAT 基因。利用毛白杨(Popular tomentosa)的叶外植体经该菌株感染后,培养3-4个星期后,在没有外源激素的 MS_u 培养基上,从叶外植体的边缘处再生出一些不正常的苗。这种苗呈典型的畸胎芽,把这种苗从外植体上切下并转到生根培养基上,在苗的基部生成大量的愈伤组织,但并不形成根,同时在愈伤组织上产生大量的芽。气相色谱法测定内源细胞分裂素的结果表明,转化苗的异戊烯基腺苷的含量是正常苗的14倍。纸电泳结果表明,转化苗均含有特异的胭脂碱。在加有60-100μg/ml 氯霉素的培养基上,转化苗可以正常生长,而未转化的苗在培养约40天后变白死去。用酚类物质水杨酸预处理农杆菌有助于提高转化频率。     

玉米素和IAA对番茄子叶再生的影响

高水平的外源ZT抑制番茄子叶不定芽形成,导致畸形芽大量出现,延缓再生芽的生根过程。在低水平的ZT和高浓度的IAA共同作用下,不定芽的再生速度加快,正常芽比例增加,后期生根过程明显加快。     

乙烯前体ACC对番茄叶外植体分化的影响（简报）

在含有5μmol/L IAA,ZT或BA的MS培养基中,添加1.0μmol/L ACC能显著促进番茄叶外植体分化出芽;而在含有5μmol/L IAA的MS培养基中,添加10μmol/L ACC可直接诱导番茄叶外植体分化出大量的根。番茄叶外植体的乙烯释放速率随培养基中ACC浓度提高而增高,培养第2天时为最高,但与IAA的浓度关系不大。     

添加物影响花生外植体再生及基因转化

取花生上胚轴为外植体,在农杆菌介导的基因载化过程中,在共育时加入一定量的烟草或马铃薯提取物,在共培养基和分化培养基中分别加入0.179mM的还原型谷胱甘肽（GSH）和0.147mM的VC 0.035mM的VE。进行培养,结果表明,添加15%（W/V）的烟草或马铃薯提取物,胚轴生长状况明显好于对照组,分化率较对照组提高10%-23.5%。自由基清除剂能明显减轻外植体褐化现象,促进外植体再生,GUS检测阳性率最高可达10.2%。     

黄瓜子叶外植体龄对子叶生根的影响

本文对无菌培养条件下黄瓜子叶外植体龄对子叶生根能力的影响进行了研究,结果表明,对于分别萌发５天、１０天、１５天、２０天的子叶在ＭＳ基本培养基上,无论有生长系ＮＡＡ,均以５天龄的子叶生根能力最强。附加１．０ｍｇ／ＬＮＡＡ,显著促进子叶生根。对子叶起始过氧化物酶活性测定表明,子叶起始过氧化物酶活性与生根能力之间存在有一定的关系。     

番茄子叶外植体芽的分化过程与乙烯释放的关系

本文对番茄子叶外植体培养过程乙烯的释放和芽的分化进行了研究,结果表明,番茄子叶外植体培养１ｈ后开始释放乙烯,第３小时乙烯释放达到最高峰,随后迅速下降,乙烯释放水平与番茄子叶外植体分化能力呈负相关,外植体芽的分化过程中,乙烯释放水平很低,但在芽形成以前略有增加,乙烯拮抗剂及其生物合成抑制剂促进子叶外植体芽的分化过程,伤害了可能是导致番茄子叶外植体乙烯释放的主要原因。     

枳壳外植体离体再生及农杆菌介导的遗传转化

以枳壳实生苗的上胚轴及茎段为材料,在附加有ＢＡ和ＭＴ培养基上进行培养,上胚轴出芽率普遍高于茎段,ＢＡ为１ｍｇ／Ｌ时,出芽率最高,ＢＡ浓度升高,出芽率随之下降。外植体在培养基上的接种方式,对出芽有一定影响,上胚轴切段 形态学下端垂直插入,出芽率高。     

番茄ACC合酶反义基因对河套蜜瓜的转化

河套蜜瓜（CucumismeloLcvHetau）的子叶经预培养。芽诱导和生根培养,获得再生小植株,诱导率达58％。取带有番茄ACC合酶反义基因的双元载体pMQ6／JM109与农杆菌（Agrobacteriumtumefaciens）LBA4404经三亲融合后,与在MS0上萌发5d、并在MS＋1mg／LNAA培养基上预培养3d的子叶共培养48h,然后转入含50mg／L卡那霉素的MS＋6mg／LZT的芽诱导培养基中,1l个月后诱导生芽,待芽长1．5－2cm时转入生根培养基中,1－2周后可诱导产生大量的根,形成完整的转基因小植株。经PCR和分子杂交检测证明,目的基因已整合入河套蜜瓜的基因组中。     

Ri质粒转化番茄的初步研究

作者利用具Ri质粒的发根农杆菌(Agrobacterium rhizogenes)PRi 15834,PRiA4和PRi2659,对番茄不同品种的不同外植体进行转化试验。在感染部位诱导出毛状根和再生植株,经检测毛状根及再生植株均含有农杆碱和甘露碱。比较不同菌种诱导毛状根的频率和在不同番茄品种间差异。结果表明:发根农杆菌PRi15834和PRiA4感染番茄诱导毛状根的频率比PRi2569诱导频率高。供试番茄品种中强力米寿和粉白砂,感染效果最好。感染方式以叶盘法和胚轴的直接注射法比较适宜。     

利用GUS基因瞬时表达对大豆子叶节和胚尖转化方法的比较及优化

建立了一个综合利用CaMV 35S：：GUS基因的瞬时表达水平、表达位点和表达率来评价大豆转化效率的检测体系,并利用该体系对分别以栽培大豆（Glycine max L．）“科丰6号”、“合丰35”、“中黄13”和“垦农18”的子叶节和胚尖为外植体的农杆菌介导的2种不同转化方法进行了比较和优化。实验结果表明,不同的转化方法对大豆的品种要求不同,“科丰6号”和“合丰35”2个品种适合子叶节转化法．而“中黄13”更适于胚尖转化法。大豆萌发培养基和共培养基对转化频率的影响存在交互作用,低6-BA的萌发培养基和高6～BA的共培养基配合、或者高6-BA的萌发培养基和低6-BA的共培养基配合使用适合“科丰6号”子叶节转化法,而“中黄13”胚尖转化法要求萌发培养基和共培养基中的6-BA浓度都较高。另外,在子叶节转化系统中,6d苗龄的子叶节共培养3dGUS基因的瞬时表达率较高,而在胚尖转化系统中大豆的萌发时间对GUS基因的瞬时表达率并没有显著的影响,胚尖与农杆菌共培养较宜的时间为5d。     

根癌农杆菌介导转化番茄的影响因素

综述影响根癌农杆菌介导番茄转化效率的因素,包括根癌农杆菌菌株类型、Vir基因的活化、选择标记基因、植物基因型、外植体类型、培养基中是否附加植物激素和抑菌抗生素、菌液浓度、侵染时间长短,是否预培养和共培养天数等;同时不同的培养方式也是影响番茄转化效率的主要因素,包括液体培养法、农杆菌介导的floral-dip转化法、超声波辅助农杆菌介导法、农杆菌介导与基因枪轰击结合法等.     

Transformation of Cucumis melo cv. Hetau with Tomato Antisense ACC Synthase Gene

An efficient in vitro plant regeneration system of Cucumis melo L. cv. Hetau was established. Regenerated plantlets were obtained from cotyledons after preculture, shoot inducing culture and root inducing culture. A high regeneration rate was achieved up to 58%. Cucumis melo was transformed with the antisense tomato ACC synthase gene in binary vector pMQ6 via Agrobacterium tumefaciens mediated gene transfer. Kanamycin resistant plantlets were obtained on MS medium with 6 mg/L zeatin, 50 mg/L kanamycin and 650 mg/L cefotaximine. PCR and molecular hybridization analysis showed that tomato ACC synthase antisense cDNA was integreted into the genome of C. melo.     

Incorporation of Resistance Gene into Tomato using Binary Vector System

ＩｎｃｏｒｐｏｒａｔｉｏｎｏｆＲｅｓｉｓｔａｎｃｅＧｅｎｅｉｎｔｏＴｏｍａｔｏｕｓｉｎｇＢｉｎａｒｙＶｅｃｔｏｒ　ＳｙｓｔｅｍＺＨＡＯＳｈｕ－ｈｕｉ（赵淑慧）;ＸＵＥＧｕｏ－ｘｉｏｎｇ（薛国雄）（ＩｎｓｔｉｔｕｔｅｏｆＤｅｖｅｌｏｐｍｅｎｔａｌＢｉｏ...     

Plant Regeneration from Cotyledon Protoplasts of Tomato (Lycopersicon esculentum L.)

otyledon protoplasts of tomato (Lycopersicon esculentum L.) isolated from 2–3 week grown seedlings were cultured in MS liquid medium (2,4-D 1, 6-BA 0.1 mg/l) and fresh medium added subsequently. After 6 weeks culture, the cell clusters were transferred to semisolid medium (the additive same as in liquid medium, agar 0.3%). When the calli grew to 0.5 cm in diameter, transfer them to MS medium (6-BA 2, IAA 0.2 mg/l) for differentiation. The regenerated plants were obtained. After comparing different culture methods, tomato protoplasts grew better in double layers than in agar plate and hanging drops.     

根癌农杆菌介导AtNHX1基因转化番茄的研究

构建AtNHX1基因植物表达载体,通过农杆菌介导法将其转入番茄。探讨了外植体类型、农杆菌菌株和不同筛选标记对芽诱导分化的影响。对抗性植株进行PCR检测,获得15株转基因植株。对转基因番茄T1代进行80mmol/LNaHCO胁迫处理,转基因植株的相对生长量高于对照植株,显示AtNHX1基因的导入提高了番茄对碱性盐的耐受性。     

影响一品红再生系统建立的因素

本试验以一品红“旗帜”和“福星”两个品种的茎段和叶片为试验材料,研究了影响建立一品红再生系统的因素。筛选出一品红初代培养的适宜培养基。     

真菌遗传转化系统的研究进展

真菌遗传转化是功能基因研究的重要方法,就真菌遗传转化系统的最新研究进展进行了综述,主要包括真菌遗传转化的方法、各种方法的优缺点、选择标记的种类、标记基因的分离方法以及转化系统的应用等。     

农杆菌介导的番茄遗传转化的研究进展

番茄(Lycopersicon esculentum)传转化体系对其功能基因的研究和基因工程育种有重要影响,对农杆菌(Agrobacterium tumefaciens)介导的番茄遗传转化的研究进展进行了综述,主要包括影响番茄遗传转化效率的几个因素,如番茄的基因型、外植体状态、预培养和侵染过程、分化培养基中的激素和抗生...     

In Planta Transformation of Tomato

In this study, in planta transformation of tomato (Solanum lycopersicum L.), using fruit injection and floral dip, is reported. Agrobacterium tumefaciens strain EHA 105 containing one of three constructs, i.e., pROKIIAP1GUSint (carrying the Apetala 1 [AP1] gene), pROKIILFYGUSint (carrying the LEAFY [LFY] gene), or p35SGUSint (carrying the β-glucuronidase [GUS] gene), was used for plant transformation. For fruit injection transformation, no significant effects (p > 0.05) of the construct used were observed. The highest frequency of transformation was obtained following 48-h incubation of tomato fruit with bacterial cells harboring either one of the three constructs; transformation frequencies of 17%, 19%, and 21% for AP1, LFY, and GUS gene constructs, respectively, were obtained. When fruit maturity was evaluated in fruit injection experiments, mature red fruit resulted in higher frequency of transformants than immature green fruit with 40%, 35%, and 42% for AP1, LFY, and GUS gene constructs, respectively. For floral dip transformation, a higher number of transformants was obtained when the GUS gene construct was used instead of either the AP1 or LFY gene construct, thus suggesting a possible inhibitory effect of the flowering genes used. When flowers were transformed prior to rather than following pollination, they yielded a higher transformation frequency, 12% for the LFY construct and 23% for the GUS construct (p < 0.05), although no transformant was obtained with the AP1 gene construct. All putative GUS-positive transformants were analyzed using polymerase chain reaction and confirmed for the presence of the transgene. Compared to control plants, transgenic plants carrying either the AP1 or LFY transgene flowered earlier and showed several different morphological characters.