Inheritance and expression of transgenes in T2 and T3 generations of Lotus corniculatus transformed using Agrobacterium tumefaciens

The inheritance and expression of the reporter gene uidA, encoding β-glucuronidase (GUS), was previously analysed in the T₁ generation of 25 independent transformed lines of Lotus corniculatus cv. Leo. In the work reported here, GUS activity in various tissues of seven of these lines was tested in the T₂ generation. Four representative lines were chosen for more detailed study in the T₃ generation. Lines 25 and 38 had multiple, independently segregating transgene inserts; lines 24 and 39 appeared to transmit one segregating transgene insert to their T₁ progeny, although transgene expression was low and was detected in fewer seedlings than expected in line 39. The uidA gene was inherited and expressed in seedlings of T₁, T₂ and T₃ generations of all four lines. In all lines, transgene expression varied between tissues, with more embryos than seedlings having detectable GUS activity. Studies in the T₂ generation showed that use of transgenic plants as female or male parents altered the frequency of expression of the transgene in progeny. By contrast, in the T₃ generation the use of transgenic plants as female or male parents did not effect either frequency of transmission, or expression of the transgene, in any of the four lines. Transgene inheritance was also similar among individual pods within flower heads and between individual flower heads. This revised version was published online in July 2006 with corrections to the Cover Date.     

Evidence of Migration and Endophytic Presence of Agrobacterium tumefaciens in Rose Plants

Agrobacterium tumefaciens was isolated from stem tumors of several rose cultivars showing that the bacterium is the causal agent of aerial galls in rose plants. No differences were observed in the characteristics of the Agrobacterium isolates from crown or aerial galls. Stem inoculation of ten rose cultivars showed that all of them were susceptible to A. tumefaciens but differences in the size of the resulting tumors were observed. The movement of A. tumefaciens in rose plants was demonstrated using two wild type strains and two antibiotic resistant mutants. Three months after inoculation, the inoculated strains were recovered in the roots, crown and below and above the inoculation site but low numbers of pathogenic Agrobacterium cells were isolated. New tumors appeared in 5% of the noninoculated wounds. A. tumefaciens was isolated from the stem at different distances from the tumor in naturally infected plants. In symptomless commercial plants, the isolation from the roots, crown and at different stem levels demonstrated the existence of systemic and latent infections in rose. Direct isolation using a nonselective and selective media with or without a previous enrichment step were efficient methods for isolating tumorigenic Agrobacterium from the different parts of rose plants.     

根癌农杆菌介导的西瓜遗传转化研究进展

介绍了根癌农杆菌介导的西瓜遗传转化的研究现状,综述了西瓜高效再生体系的建立和遗传转化过程中的主要影响因素,包括最佳外植体的选择、最适培养基的制备,以及抗生素、农杆菌菌株类型、菌液浓度、侵染时间、预培养时间、共培养时间和添加乙酰丁香酮等内容,讨论了西瓜遗传转化存在的问题,并对这一体系的发展进行了展望。     

根癌农杆菌介导的CG1-400-RNase基因转化创制锦橙转基因再生植株

【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。     

Communication to the Editor Degradation of Bromoxynil,Ioxynil, Dichlobenil and their Mixtures by Agrobacterium radiobacter 8/4

Degradation of three benzonitrile herbicides, bromoxynil (3,5-dibromo-4-hydroxybenzonitrile), ioxynil (3,5-diiodo-4-hydroxybenzonitrile), dichlobenil (2,6-dichlorobenzonitrile), and their mixtures by the soil micro-organism Agrobacterium radiobacter 8/4 was studied in batch cultures. Bromoxynil was found to be most rapidly degraded, while dichlobenil had the lowest toxicity to our strain. All transformations of studied benzonitriles were performed by the nitrile hydratase which has been shown to act on a broad range of substituted aromatic nitriles. © 1997 SCI.     

根癌农杆菌介导的稻瘟病菌近等菌系的构建与初步应用*

 利用根癌农杆菌C58C1介导转化1株致病力极弱的野生型稻瘟病菌菌株CY2,成功地获得3000多个转化子,混合接种于携带25个不同抗病单基因的水稻近等基因系上,获得4株单胞致病突变体;继代培养和PCR检测表明,这些突变体均为T-DNA插入CY2的后代,而非自发突变或外源菌株污染。将4株突变体与实验室已保存的20株致病突变体分别回接水稻近等基因系,推断其携带的致病基因类型,结果表明突变体分别针对13个抗性单基因的18个单基因系发生了突变,其中针对Pi-a,Pik-s,Pi-sh单基因系表现出较高的侵染频率。由于这些突变体均来自同一个菌株,仅致病性存在差异,属于近等菌系,利用其分析出102个水稻品种中的34个品种的抗病基因组成,其中,抗病基因Pik-s,Pi-a,Pi-sh,Pi-19(t)分布频率较高,分别达70.59%,61.76%,52.94%和44.12%,且带有这类基因的糯稻品种多于杂交水稻。     

Building a Better Bean： Prospects for the Next Generation of Transgenic Soybeans

The first transgenic soybeans, with the glyphosate-tolerant trait, were introduced just over a decade ago in 1996. The rapid rate of adoption and global cultivation of transgenic soybeans since then is an unqualified success story. Biotech soybeans are being grown on nearly 60 million hectares in 9 countries and double-digit growth is expected for the near future （James, 2006）. However, while biotechnology-derived soybeans were revolutionizing agriculture, the technology to produce them has advanced only slightly since the first transgenic plants were produced in the late 1980＇s. Soybeans remain a difficult crop to transform and there are only a small number of successful procedures that are routinely applied.     

农杆菌介导大豆遗传转化研究进展及其应用

农杆菌介导是大豆遗传转化的主要手段。近几年来大豆遗传转化方法得到了快速发展,但是仍然存在受体基因型匮乏、转化效率低,再生系统不完善等问题,为了进一步探讨大豆遗传转化的主要问题和策略,本研究归纳了近年来农杆菌介导大豆遗传转化在受体基因型及外植体选择,菌株筛选,标记基因选用和培养基组成等方面所做的改进,以得出有效的解决方法。并且介绍了农杆菌介导大豆遗传转化在作物改良和功能基因组学上的应用。     

Proline and Glutamine Improve in vitro Callus Induction and Subsequent Shooting in Rice

This study was conducted to evaluate the effects of proline and glutamine on in vitro callus induction and subsequent regeneration and to develop a reproducible and highly efficient plant regeneration protocol in four rice genotypes, viz. Pawana, Jaya, Indrayani and Ambemohar. Considerable variation in response to plant growth regulators and amino acid supplements used was observed in all the four genotypes. Medium supplemented with proline and glutamine was shown to be superior to medium without proline and glutamine. The best callusing from mature embryo was observed on Murashige and Skoog(MS) medium supplemented with 2.0 mg/L 2,4-dichlorophenoxyacetic acid(2,4-D), 500 mg/L proline and 500 mg/L glutamine. Shoot induction was higher in the callus obtained from medium supplemented with 500 mg/L proline and 500 mg/L glutamine. The highest shoot regeneration frequency(83.2%) was observed on MS medium with 2.0 mg/L benzylaminopurine, 0.5 mg/L 1-naphthaleneacetic acid, 500 mg/L proline, and 500 mg/L glutamine in the callus obtained from MS medium supplemented with 2.0 mg/L 2,4-D, 500 mg/L proline and 500 mg/L glutamine. Among the four genotypes, Pawana has the highest regeneration efficiency(83.2%), whereas the regeneration efficiency of the rest three rice genotypes was in the range of 32.0% to 72.3%. This optimized regeneration protocol can be efficiently used for Agrobacterium mediated genetic transformation in rice.     

植物非组培遗传转化方法研究的进展

在许多植物中,转化效率低一直是制约分子育种和基因功能研究的障碍。目前,在拟南芥等模式植物的研究中,非组培方法已成为遗传转化的主要手段,这些方法在其它植物中也有广阔的应用前景。本文介绍了几种较为成功的非组培遗传转化方法的应用情况和技术原理,并对下一步研究工作提出了建议。