小油桐外植体的KN1基因遗传转化及其超量表达的转基因植株

本研究采用农杆菌介导法将KN1基因遗传转化小油桐,并获得了转基因植株。在研究中分析了农杆菌菌液菌液的浓度、侵染时间和外植体的大小对遗传转化效率的影响以及KN1基因超量表达对转基因植株再生的影响。研究结果表明:以苗龄15d左右的小油桐无菌苗子叶为外植体,农杆菌菌液浓度OD600为0.6～0.8时,侵染8min,外植体大小为(0.8×0.8)～(1.0×1.0)mm时,遗传转化效果最好;对抗性芽及再生植株进行GUS及PCR检测结果表明,KN1基因已经整合到小油桐植物基因组中。KN1基因的超量表达可提高小油桐再生芽分化,影响转化芽及植株的外观形态及叶片的表型,包括芽及植株矮小,茎杆粗壮;叶片缩小,边缘分裂,对称性丧失,无子叶柄等。     

转KN1毛果杨细胞分裂素、赤霉素含量及其相关形态结构

为了研究KN1基因超量表达对木本植物生长发育的影响,本研究测定了转KN1基因毛果杨细胞分裂素类激素及赤霉素的含量,并进行表型及解剖结构观察。结果表明,转基因毛果杨植株细胞分裂素类激素ZR、dhZR和iPA的含量分别为14.14ng/g、18.29ng/g和12.43ng/g,明显高于对照的9.44ng/g、12.47ng/g和7.60ng/g,而赤霉素GAs含量为9.61ng/g,低于对照的12.14ng/g,从而引起其植株矮化、分枝增加、叶片增厚、叶色变深、表皮细胞增大、栅栏和海绵组织发达等现象。本研究将为探索KN1基因对木本植物生长发育的影响提供依据。     

ipt基因促进矮牵牛遗传转化效率及热激启动子驱动基因删除

本研究主要探讨ipt基因对矮牵牛遗传转化不定芽诱导影响及拟南芥热激启动子hsp18.2驱动重组酶基因flp的热诱导外源基因删除表达载体在矮牵牛中的基因删除效果.本研究中将植物表达载体pBin-hsp18.2:flp-35S:ipt及对照载体pBin-hsp18.2:flp遗传转化矮牵牛,以获得转基因植株,分析比较不定芽的诱导和转基因植株进行的热激基因删除.研究结果表明,ipt基因可促进矮牵牛遗传转化过程中不定芽的诱导,其不定芽诱导率为21.5%,显著高于对照的8.7%.在44℃,6 h,热激6次的条件下,转基因矮牵牛植株表型恢复正常,经 GUS蛋白表达分析及PCR、RT-PCR检测,证明外源基因已经被删除.转基因矮牵牛基因删除效率最高可达43.8%.本研究为ipt基因在一些遗传转化困难植物转基因中的应用奠定了基础.     

农杆菌GV3101中反式玉米素合成基因促进烟草再生北大核心CSCD

胭脂碱型农杆菌GV3101已经被广泛用于植物遗传转化研究。已有的研究结果证明,农杆菌GV3101株系含有的反式玉米素合成(trans-zeatin synthesizing,tzs)基因编码产物会影响烟草细胞器的形态及细胞的生理状态。然而,有关tzs基因对遗传转化过程外植体再生的影响研究却少有报道。本文在前期研究工作的基础上,以2种烟草、4个农杆菌株系为组培实验材料,验证了胭脂碱型农杆菌tzs基因产物的生理活性。结果表明:以外源添加生长调节物质的外植体为阳性对照,在不添加任何生长调节剂的培养基上,与GV3101菌株共培养的烟草外植体能分化再生,并发育成完整植株;外植体再生与GV3101携带的质粒种类无关;外植体与农杆菌GV3101培养液共培养24 h,烟草再生效果较好;与GV3101株系共培养24 h,将外植体烟草叶片匀浆,经亲和柱分离纯化后,检测出烟草外植体叶片中高达0.78 ng/g FW-1的反式玉米素含量。菌落PCR扩增结果证实,农杆菌GV3101株系有tzs基因序列。以上结果表明,农杆菌GV3101株系内的tzs基因的表达产物有生理学活性,能够促进烟草外植体再生,调节细胞生长。     

农杆菌GV3101中反式玉米素合成基因促进烟草再生

胭脂碱型农杆菌GV3101已经被广泛用于植物遗传转化研究。已有的研究结果证明,农杆菌GV3101株系含有的反式玉米素合成 (trans-zeatin synthesizing,tzs)基因编码产物会影响烟草细胞器的形态及细胞的生理状态。然而,有关tzs基因对遗传转化过程外植体再生的影响研究却少有报道。本文在前期研究工作的基础上,以2种烟草、4个农杆菌株系为组培实验材料,验证了胭脂碱型农杆菌tzs基因产物的生理活性。结果表明：以外源添加生长调节物质的外植体为阳性对照,在不添加任何生长调节剂的培养基上,与GV3101菌株共培养的烟草外植体能分化再生,并发育成完整植株;外植体再生与GV3101携带的质粒种类无关;外植体与农杆菌GV3101培养液共培养24 h,烟草再生效果较好;与GV3101株系共培养24 h,将外植体烟草叶片匀浆,经亲和柱分离纯化后,检测出烟草外植体叶片中高达0.78 ng/g FW-1的反式玉米素含量。菌落PCR扩增结果证实,农杆菌GV3101株系有tzs基因序列。以上结果表明,农杆菌GV3101株系内的tzs基因的表达产物有生理学活性,能够促进烟草外植体再生,调节细胞生长。     

超量表达IPT和KN1基因对转基因烟草生长发育的影响

为了比较异戊烯转移酶基因IPT和玉米homeobox基因KNI超量表达对植物生长发育的影响,将35S：：IPT和35S：：KNI分别导入烟草。观察发现,过量表达IPT和KNI基因对植株再生频率、形态、生长周期和顶端优势等方面的影响均相似,但在生根和开花结籽方面,35S：：IPT转基因植物所受影响更显著。细胞分裂素含量检测表明,35S：：IPT转基因植物叶片中细胞分裂素的含量高于35S：：KNI转基因植物。     

大豆诱导型启动子驱动类受体蛋白激酶GmSARK转基因植物分析

植物LRR型类受体蛋白激酶在植物生命活动中发挥着重要作用。前期研究发现, 大豆(Glycine max)LRR型类受体蛋白激酶基因GmSARK可能参与调控大豆叶片的衰老过程。利用CaMV 35S启动子驱动组成型过表达GmSARK基因可导致转基因植株出现致死表型, 据此构建了可诱导型启动子GVG驱动GmSARK基因过表达的双元表达载体, 转化野生型拟南芥(Arabidopsis thaliana)并获得了多株转基因植株。研究结果表明, 外源施加诱导物地塞米松可引起GmSARK基因在转基因植株中过表达, 并导致转基因植株出现叶片变黄下卷和生长受抑制等表型; 外源细胞分裂素处理可以抑制GmSARK的表达, 但是不能逆转GmSARK过表达所引起的上述变化。     

大豆诱导型启动子驱动类受体蛋白激酶GmSARK转基因植物分析

植物LRR型类受体蛋白激酶在植物生命活动中发挥着重要作用。前期研究发现,大豆（Glycine max）LRR型类受体蛋白激酶基因GmSARK可能参与调控大豆叶片的衰老过程。利用CaMV35S启动子驱动组成型过表达GmSARK基因可导致转基因植株出现致死表型,据此构建了可诱导型启动子GVG驱动GmSARK基因过表达的双元表达载体,转化野生型拟南芥（Arabidopsis thaliana）并获得了多株转基因植株。研究结果表明,外源施加诱导物地塞米松可引起GmSARK基因在转基因植株中过表达,并导致转基因植株出现叶片变黄下卷和生长受抑制等表型;外源细胞分裂素处理可以抑制GmSARK的表达,但是不能逆转GmSARK过表达所引起的上述变化。     

根癌农杆菌介导转化Lesquerella fendleri L．的研究

以Lesquerella fendleri L.叶片和叶柄为外植体,研究了根癌农杆菌介导的β-葡萄糖醛酸苷酶基因(β-glucuronidase,Gus)的遗传转化技术,将再生植株叶片进行PCR分子鉴定和Gus组织染色,证实外源基因已稳定整合到植物基因组中,并得到表达。     

根癌农杆菌介导的CMO基因转化马铃薯的研究

通过根癌农杆菌介导法将强组成型表达启动子CaMV 35S和逆境诱导表达启动子rd29A 驱动的菠菜胆碱单加氧酶(CMO)基因导入马铃薯栽培品种夏波蒂(Shepody)和费乌瑞它( Favorita)中,获得了抗卡那霉素再生植株,对转化植株进行PCR检测初步表明CMO基因已整 合到转基因马铃薯基因组中.并对光照强度对马铃薯试管薯遗传转化的影响进行了研究,结果表明在诱导芽分化的过程中,2 000 lx的光照强度有利于试管薯薄片直接再生出抗性芽.     

Investigation of the mechanism of temperature sensitivity of the electroreceptors of ampullae of lorenzini

In experiments on Black Sea skates (Raja clavata), the potential of the receptor epithelium of the ampullae of Lorenzini and spike activity of single nerve fibers connected to them were investigated during electrical and temperature stimulation. Usually the potential within the canal was between 0 and –2 mV, and the input resistance of the ampulla 250–400 k. Heating of the region of the receptor epithelium was accompanied by a negative wave of potential, an increase in input resistance, and inhibition of spike activity. With worsening of the animal's condition the transepithelial potential became positive (up to +10 mV) but the input resistance of the ampulla during stimulation with a positive current was nonlinear in some cases: a regenerative spike of positive polarity appeared in the channel. During heating, the spike response was sometimes reversed in sign. It is suggested that fluctuations of the transepithelial potential and spike responses to temperature stimulation reflect changes in the potential difference on the basal membrane of the receptor cells, which is described by a relationship of the Nernst's or Goldman's equation type.I. P. Pavlov Institute of Physiology, Academy of Sciences of the USSR, Leningrad. I. M. Sechenov, Institute of Evolutionary Physiology and Biochemistry, Academy of Sciences of the USSR, Leningrad. Pacific Institute of Oceanology, Far Eastern Scientific Center, Academy of Sciences of the USSR, Vladivostok. Translated from Neirofiziologiya, Vol. 12, No. 1, pp. 67–74, January–February, 1980.     

Determination of stoichiometry of radioiodination of proteins

A sensitive method for the detection of small quantities of hydrophobic antioxidant free radical scavengers such as butylatedhydroxytoluene (BHT) and butylatedhydroxyanisole (BHA) in aqueous samples is described. The procedure involves extraction of the hydrophobic free radical scavenger into an organic solvent phase, followed by the subsequent reaction of an aliquot of this extract with the stable cation radical tris(p-bromophenyl)amminium hexachloroantimonate (TBACA). In experiments with BHT and BHA, the loss of TBACA absorbance at 730 nm was found to be linearly proportional to the amount of antioxidant added, with quantities of BHT as small as 200 pmol being easily detectable. In aqueous suspensions of dimyristoylphosphatidylcholine vesicles, assays of the aqueous BHT concentration showed that BHT partitioned strongly into the membrane phase, achieving very high BHT/phospholipid ratios. For a given concentration of BHT, partitioning into the membrane phase was greater in large, multilamellar liposomes than in either small, single-walled vesicles or in purified rat brain synaptic vesicle membranes. Direct assay of BHT and BHA in phospholipid membranes, however, was complicated by a nonspecific interaction between TBACA and the phospholipid.