Enhancement of bovine growth hormone gene expression by increasing the plasmid copy number

Effect of copy number on the expression of bovine growth hormone gene (bGH) was investigated using the copy number mutants such as pKBJ10, pBJ( tet)10, pUBJ10-1, and pUBJ10 plasmids. The cells harboring plasmids below 84 copies/cell did not produced detectable levels of bGH. When the ColE1 replicon was replaced with the mutated ColE1 replicon originated from pUC19 plasmid, the copy number was increased to about 300 copies/cell and bGH production was enhanced by 11.5% (pUBJ10-1) and 12.3% of total cell protein (pUBJ10). A large amount of mRNA caused by increment of copy number would be needed to overcome some inhibitory threshold and might be an important factor for regulating bGH expression.     

多种外源因素对转基因植物抗性影响的研究进展

病毒病对植物的危害在近些年日益严重,培育抗病毒转基因植物是众多防治方法中较为有效的一种。转基因植物在不同机制介导下会产生抗性,尤其是病毒基因介导的抗性试验最为普遍。众多研究证明,抗性受多种因素作用且表现为不同水平。对插入序列来源、转录后hpRNA长度、茎环比和拷贝数等多个因素在近些年的研究进展进行了综述,重点对各自在转基因植物抗性产生中的特点及影响程度进行了分析,并对其他因素进行了展望。     

A small HSP gene is not responsible for diapause and cold tolerance acquisition in Chilo suppressalis

Abstract:  The cDNA sequence of a small heat shock protein ( hsp19.7 ) was cloned and sequenced from the rice stem borer, Chilo suppressalis Walker. The cDNA encoded a protein of 177 amino acids with a calculated molecular weight of 19.7 kDa. The deduced amino acid sequence showed the highest identity of 90% to Bombyx mori hsp19.9 . Expression levels of hsp19.7 were similar between diapausing and non-diapausing larvae. In non-diapausing larvae, but not in diapausing ones, hsp19.7 expression was upregulated by cold acclimation. Involvement of hsp19.7 in larval diapause and cold tolerance in C. suppressalis is discussed.     

An evaluation of models for the effect of plasmid copy number on bacterial growth rate

The rates of growth of recombinant bacteria depend on their plasmid content. This is modelled by expressing the specific growth rate in terms of the number of copies of the plasmid per cell. Three models in common use have been tested with different Escherichia coli strains and one strain of Bacillus stearothermophilus containing different plasmids. While no particular model was decisively better than others for all data, that of Bentley & Quiroga (Biotechnol. Bioeng. 1993, 42: 222–234) was the best for specific growth rates which vary inversely with the plasmid copy number, and a modified form of the model of Satyagal & Agarwal (Biotechnol. Bioeng. 1989, 33: 1135–1144) was the best for growth rates which increase with the copy number. The differences appear to be linked to the plasmid replication mechanisms. Contrary to some claims, no model portrayed the experimentally observed inflection points.     

不同结荚习性夏大豆种质的农艺表现及其与产量的相关分析

以533份不同结荚习性大豆种质为试验材料,研究了不同结荚习性大豆种质在黄淮夏播生态区的农艺性状表现,并对主要农艺性状和产量的相关性进行了分析。结果表明:大豆种质的营养期、株高、有效分枝、单株荚数、倒伏性、株型等性状的平均值随无限-亚有限-有限结荚习性递减,生殖期、单株粒重、百粒重、小区产量等性状平均值的变化趋势则相反。相关分析表明,无限结荚习性种质的产量与株高、单株粒重呈极显著正相关,偏相关系数分别为0.602**、0.566**,与有效分枝、倒伏性呈显著负相关,偏相关系数分别为-0.384*、-0.451*。亚有限结荚习性种质的产量与生殖期、单株粒重呈显著、极显著正相关,偏相关系数分别为0.156*、0.536*,与有效分枝呈极显著负相关,偏相关系数为-0.323**。有限结荚习性种质的产量与单株粒重、株高呈极显著正相关,偏相关系数分别为0.433**、0.262**,与株型、单株荚数呈显著、极显著负相关,偏相关系数分别为-0.149*、-0.198**。结合不同结荚习性品种的生长特点,本研究认为,无限结荚习性品种株高较高且株高与产量呈极显著正相关,适合干旱地区种植;亚有限结荚习性品种生殖期与产量呈正相关,生殖期内生长旺盛需要较多的养分供应;有限结荚习性品种的营养生长持续时间短,株高较矮,吸收光能有限,实现高产主要依赖各性状器官间的平衡。生产中,有限结荚习性品种的营养生长期间既需要充足的肥水促其生长,又要防止旺长。     

F-box基因拷贝数目变异的机制研究:以12种果蝇为例

拷贝数目变异是一种对表型变异和生物进化具有重要意义的基因组结构变异.以前的研究表明不同物种中F-box基因的拷贝数目差异较大.为了深入探索拷贝数目变异的式样和机制.我们以12个果蝇近缘种为研究对象,分析了F-box基因的系统发育关系、进化式样以及它们在染色体上的位置.结果发现,虽然各个物种中F-box基因的拷贝数目差别不大(42-47个),但是仍然存在着很多引起拷贝数目变异的基因获得和丢失事件.这说明表面上变化不大的拷贝数目在一定程度上掩盖了频繁发生的基因获得和丢失事件.通过比较这些基因在染色体上的位置,发现只有在亲缘关系很近的物种之间才能鉴定出有明显微共线性关系的基因组区段.我们还发现,造成F-box基因拷贝数目增加的主要机制是散在重复和串联重复,而反转录转座和新基因的非编码区起源也是两种值得注意的机制.此外,序列变异导致的外显子边界变化以及外显子丢失是引起拷贝数目减少的两种机制.在12种果蝇的最近共同祖先中,F-box基因的拷贝数目与现存物种基本相似,但是基因的获得和丢失事件使得现存物种中的F-box基因在构成上已经有了明显的差别.对数目变异的式样及其与基因功能的关系的研究表明,拷贝数目变异是F-box基因家族"生与死"的进化在基因组层面的系统反映,并有可能为表型变异提供了原始材料.     

Genetic approaches for studying transgene inheritance and genetic recombination in three successive generations of transformed tobacco

Transgene integration into plant genomes is a complex process accompanied by molecular rearrangements. Classic methods that are normally used to study transgenic population genetics are generally inadequate for assessing such integration. Two major characteristics of transgenic populations are that a transgenic genome may harbor many copies of the transgene and that molecular rearrangements can create an unstable transgenic locus. In this work, we examined the segregation of T1, T2 and T3 transgenic tobacco progenies. Since transfer DNA (T-DNA) contains the NptII selectable marker gene that confers resistance to kanamycin, we used this characteristic in developing a method to estimate the number of functional inserts integrated into the genome. This approach was based on calculation of the theoretical segregation ratios in successive generations. Mendelian ratios of 3:1, 15:1 and 63:1 were confirmed for five transformation events whereas six transformation events yielded non-segregating progenies, a finding that raised questions about causal factors. A second approach based on a maximum likelihood method was performed to estimate recombination frequencies between linked inserts. Recombination estimates varied among transformation events and over generations. Some transgenic loci were unstable and evolved continuously to segregate independently in the T3 generation. Recombination and amplification of the transgene and filler DNA yielded additional transformed genotypes.     

实时荧光定量PCR(TaqMan)法测定外源基因的拷贝数

实时荧光定量PCR是近年新兴的一项技术,因其快速、方便、便宜,需要DNA样品量少,无需放射性检测等优点被广泛应用于基因的定量分析。该文就实时荧光定量PCR(TaqMan)技术的发展、基本原理及测定外源基因拷贝数的技术流程做一介绍。     

植物脂肪酸去饱和酶及其编码基因研究进展

脂肪酸去饱和酶是催化脂肪酸链特定位置形成双键和产生不饱和脂肪酸的酶类。植物脂肪酸去饱和酶主要有5种（FAD2、FAD3、FAD6、FAD7和FAD8）,可分为ω-3型（FAD2、FAD6）和ω-6型（FAD3、FAD7、FAD8）两大类。其编码基因（FAD2、FAD3、FAD6、FAD7和FADS）在植物中一般有多个拷贝。同种基因在不同植物中拷贝数不同,同一植物中相同基因的不同拷贝间在序列特征、表达调控和功能等方面也存在显著差异。本文根据国内外对脂肪酸去饱和酶基因及编码产物的研究现状,分别从它们的分类、拷贝数、结构、作用机制、表达调控等方面的研究进展进行了详细的分类阐述。     

Internode length in Pisum. Gene lkd

A new, single gene, recessive internode length mutant in Pisum, lkd, is described. The internodes of lkd plants are ca. 40% shorter than comparable Lkd plants and this difference appears greater in the dark than in the light. The mutant does not appear to be dwarfed due to modified gibberellin (GA) levels, as determined by gas-chromatography-selected ion monitoring (GC-SIM) for GA₁ and GA₂₀. In relative terms, the mutant responds as well as the wild-type to applied GA₁. However due to its initial short stature it does not elongate to the same extent as the wild-type to high doses of GA₁ suggesting that some other factor, unrelated to GA levels or perception is probably limiting growth in this mutant.Author for correspondence     

柚皮素合成关键基因表达水平对目标产物积累水平的量化影响

柚皮素是一种天然黄酮类化合物,具有抗炎、抗氧化、抗病毒、预防动脉粥样硬化等多种药理活性,也是其他黄酮类化合物合成的重要前体,具有重要的应用价值。目前,微生物法生产柚皮素等黄酮类化合物由于代谢通路不平衡等原因导致产量较低,在很大程度上限制了其工业应用。文中以一株产柚皮素的酿酒酵母菌株Y-01为研究对象,利用启动子和拷贝数控制柚皮素合成代谢途径关键酶4-香豆酸:CoA连接酶(4CL)、查尔酮合成酶(CHS)和查尔酮异构酶(CHI)编码基因的表达水平,考察这些基因的表达水平对目标产物积累水平的量化影响。结果表明,柚皮素产量与4CL或CHI编码基因的表达量之间关联性较低,而与chs基因的表达量存在显著的正相关性。通过调控chs基因的表达水平,获得一株高产柚皮素的酿酒酵母工程菌株Y-04,产量较出发菌株Y-01提高了4.1倍。研究结果表明,CHS是柚皮素合成过程的关键调控靶点,合理调控CHS表达可以显著促进酿酒酵母积累柚皮素。相关结果为采用代谢工程强化微生物合成柚皮素等重要黄酮类化合物提供了重要的理论参考。