Agrobacterium-mediated transformation of the CrDAT gene and selection of transgenic periwinkle lines with a high vincristine accumulation

Catharanthus roseus contains vincristine and vinblastine, which are outstanding drugs for cancer. In the biosynthetic pathways of terpenoid indole alkaloids (TIAs) in C. roseus, deacetylvindoline 4-O-acetyltransferase (DAT) is a key enzyme that catalyses the last reaction of vindoline biosynthesis to form vinblastine and vincristine. In this study, the CrDAT transgene was transferred into the periwinkle by Agrobacterium-mediated transformation and generated transgenic periwinkle lines with an increase in vincristine accumulation. The C. roseus DAT gene was introduced into C. roseus plants and it was confirmed that CrDAT was successfully transferred into the genome of periwinkle plants and efficiently translated to synthesise recombinant DAT protein. Four transgenic periwinkle lines in T1 generation, T1-1, T1-3, T1-6, and T1-7, expressed recombinant DAT protein with the total protein content in the range of 2.86 μg.mg^?1 to 5.12 μg.mg^?1. Moreover, the vincristine contents of four transgenic lines increased by 1.63?2.48-fold compared to non-transgenic plants, ranging from 6.91 µg.g^?1 (fresh weight) to 10.53 µg.g^?1 (fresh weight). The T1-1 line had the highest vincristine content. Hence, the overexpression of the recombinant DAT protein can improve the vincristine accumulation of transgenic C. roseus plants.

Abbreviation: CrDAT - Catharanthus roseus Deacetylvindoline-4-O-Acetyl Transferase; D4H - Deacetoxyvindoline 4-hydroxylase; ELISA - Enzyme-Linked Immunosorbent Assay Monoterpene indole alkaloid; T0, T1 - Generations of transgenic plants; TIAs - Terpenoid indole alkaloids; WT- The wild-type tobacco plants (non transgenic plant); 35S - Cauliflower mosaic virus 35S promoter     

转基因甘蔗BtG-2的T-DNA侧翼序列分析及其转化事件特异性检测

转基因甘蔗BtG-2是利用农杆菌介导法把Cry1Ac-2A-gna融合抗虫基因导入‘新台糖22号’的转基因甘蔗株系,具有良好的抗虫特性和农艺性状。为了明确转基因甘蔗BtG-2的分子特征及其检测方法,推进其生物安全性评价工作,以BtG-2的T2代为研究材料,利用Southern杂交检测外源基因在转基因甘蔗基因组内的拷贝数;利用染色体步移技术分离外源基因在甘蔗基因组中插入位点的侧翼序列,并建立了该转化体高效灵敏的特异性PCR检测方法。结果表明：Southern杂交检测证明外源T-DNA以单拷贝方式插入BtG-2株系;经过3次的热不对称巢式PCR扩增,获得外源基因T-DNA左边侧翼序列984 bp和右边侧翼序列705 bp;以这2个序列和相应的T-DNA的左右端序列分别设计3对检测引物对,建立了BtG-2株系的转化事件特异性PCR检测方法,扩增效率最高的引物对LS011/LA451和RS160/RA588分别扩增到440 bp和428 bp的特异片段。其中T-DNA左侧设计的LS011/LA451检测引物对扩增的灵敏度高、特异性强,能够在甘蔗BtG-2基因组DNA相对含量为0.1%的模板中检测出转基因目的成分,相当于9个单倍体基因组拷贝数。本研究完成了转基因株系BtG-2的分子特征及其转化事件特异性检测,为该转基因甘蔗及其衍生产品的检测和身份识别提供技术依据。     

规模化畜牧养殖对生态环境的破坏及防治

近年,畜牧养殖业蓬勃发展,过去的区域化散养已经基本不存在,养殖结构逐渐向集约化、一体化、规模化过渡,养殖规模也逐渐增大。养殖副产品肉、蛋、奶等极大满足了人们的生产生活,养殖中产生的尿液、粪污对生态环境造成极大影响。为促进规模化养殖实现绿色健康发展,该文分析规模化畜牧养殖对生态环境的破坏及防治措施。     

畜牧养殖污染现状及综合治理对策

畜牧养殖业污染问题是畜牧养殖生产过程中的常见问题,对于养殖业也有一定的影响,直接影响畜牧产品质量及动物疫病防治工作。该文针对畜牧养殖业污染现状进行分析研究,阐述畜牧养殖污染问题现状,并综合新疆地区的畜牧养殖业发展状态,提出畜牧养殖污染问题综合治理的有效对策。     

水稻OsUBA基因的表达及其在促进种子萌发和开花中的功能

自噬是将功能异常或不需要的胞内组分降解的细胞学过程,广泛参与真核生物的生长发育过程、对营养缺乏的响应及生物/非生物胁迫反应。NBR1 (Next to BRCA1 gene 1, NBR1)是在植物中发现的最重要的自噬受体,但有关植物NBR1类自噬受体的研究较少,水稻中此类蛋白的研究还是空白。本文通过RT-PCR方法,从水稻日本晴幼苗的cDNA中克隆到一个含有泛素相关结构域(Ubiquitinassociated,UBA)的基因,将其命名为OsUBA。OsUBA的开放阅读框长2538 bp,编码845个氨基酸残基。OsUBA属于水稻中的NBR1类蛋白。OsUBA的启动子区有多个与光、逆境胁迫及激素反应相关的元件; OsUBA基因在水稻花药、正在萌发的种子以及根中的表达量较高,在茎和叶中也有表达; 200μmol L~(–1) ABA处理显著抑制OsUBA的表达,100μmol L~(–1) GA处理后OsUBA的表达略有升高。对OsUBA过表达水稻株系的研究表明,转基因水稻种子的萌发比野生型更快, ABA (3μmol L~(–1))处理显著抑制OsUBA过表达水稻株系种子的萌发, GA (100μmol L~(–1))处理对OsUBA过表达水稻株系种子的萌发略有促进;OsUBA过表达水稻株系的开花时间较野生型明显提前。这些结果表明,水稻NBR1蛋白基因OsUBA的表达和功能可能与对开花时间和种子萌发的调控以及生物/非生物胁迫反应有关。     

动物细胞因子研究进展

细胞因子是免疫活性细胞和其它细胞分泌的具有多种生物活性的多肽或蛋白质 ,这些细胞因子是重要的信息传递物质 ,特别在免疫、炎症和造血系统等生物学反应过程中具有重要的作用。近几年 ,细胞因子的研究已成为生物学研究中最活跃的领域之一。新的细胞因子不断被发现 ,大量的细胞因子正在或将要应用于医学临床。细胞因子的研究带动了整个免疫学的发展。其应用展现出广阔的前景。文章简要介绍了动物细胞因子的作用及其应用前景 ,包括细胞因子与免疫抑制的关系、细胞因子在免疫增强剂和免疫治疗剂、构建新型基因工程苗以及细胞因子对于蛋白质、脂肪和葡萄糖代谢的调控作用等方面的主要研究进展     

玛沁县畜牧业现状及可持续发展对策

通过对玛沁县畜牧业发展现状的分析，总结多年来的经验，提出了畜牧业可持续发展的策略。     

Genotype evaluation,muscle histopathology and ultrastructure in a Duchenne muscular dystrophy animal model

AIM: To evaluate the genotype , muscle histopathology and ultrastructure in dko mice. METHODS: Dystrophin/Utrophin-deficient double knockouts (dko) mice were obtained from university of Oxford, UK. Genotype of filial generation of heterozygote was evaluated by PCR-SSP. HE staining and fluorescent immunohistochemistry by SABC-Cy3 were used to detect striated muscle of dko mouse, and the muscle ultrastructure was observed by transmission electron microscope(TEM). RESULTS: In 112 filial generation mice, there were 28 mdx (25.0%), 26 dko (23.2%) and 58 heterozygote (51.8%), which coincided with the law of Mendelian inheritance. HE staining showed that the myocytes were not very uniform, there were phenomenon of round outline, centrally nucleated fibers, widening interspace, inflammatory cell infiltration and connective tissue proliferation in dko mice. There were no any immunofluorescent expression of dystrophin and utrophin in sarcolemma in dko mice. TEM showed sarcolemma breakage, separation and edema, and loose myofibril texture, inflammatory cell infiltration and connective tissue proliferation in dko mice. CONCLUSION: PCR-SSP is a very quick and accurate way for genotype evaluation of filial generation. The pathophysiology of dko mouse was very similar to Duchenne muscular dystrophy (DMD), and dko mouse is an ideal animal model for study of DMD clinical therapy.     

镉毒性研究进展

本文综述了动物生产领域对镉的研究结果。这些内容包括 :(1)镉的化学特性 ;(2)镉在自然界的分布 ;(3)镉的生物学作用 ;(4)镉在动物体内的吸收和排泄 ;(5)镉对动物的危害 ;(6)镉与其它物质之间的关系 ;(7)镉的需要量、中毒量及饲料中镉的最大允许量。     

共轭亚油酸(CLA)对动物营养效应研究进展

本文对共轭亚油酸对动物的免疫作用、抗癌及防止动脉粥样硬化作用、抗氧化作用等进行了综述 ,重点论述了共轭亚油酸的营养再分配作用 ,并对有待继续研究的问题进行了探讨。