Oct4,Sox2,Nanog和c-Myc基因在蚯蚓发育早期的表达

通过已知物种的干细胞多能性基因核酸序列,利用同源序列法,成功克隆得到赤子爱胜蚓(Eisenia foetida)的Oct4,Nanog,Sox2和c-Myc 4个基因,并通过实时荧光定量PCR分析了上述4个基因在赤子爱胜蚓茧孵化期间的表达情况。结果表明:赤子爱胜蚓茧在孵化期0~21 d,4个基因呈相似的规律,表达量均逐步升高,在21 d达到峰值;在幼虫期Oct4,Nanog和c-Myc 3个基因的表达量高于孵化期。结果表明,上述基因在赤子爱胜蚓早期发育过程中表达模式具有一定的相似性。     

利用分子标记辅助选择技术培育抗虫水稻品种的研究

Bt基因是从苏云金芽孢杆菌中克隆出的针对鳞翅目害虫的抗虫基因,经过基因工程方法人工改造已经获得Cry1Ab/Ac、Cry1C等抗虫基因。以云南主栽水稻品种楚粳28为受体,以携带Bt抗虫基因的"华恢1号"、"RJ-5"和"T1C-19"分别作为供体,利用分子标记辅助选择(MAS)技术进行回交育种,选育成携带Bt抗虫基因水稻新品系。结合农艺性状表现,获得了云抗虫稻1号、云抗虫稻2号、云抗虫稻3号Bt蛋白高表达的水稻抗虫品系。这些抗虫品系为云南省抗虫水稻的遗传改良和生产应用提供了基因资源和应用基础。     

有机物料接种蚯蚓对设施菠菜产量及品质的影响

为解决农村有机固体废弃物产能过剩及设施菜地生产力下降的问题,本研究拟建立"有机物料还田+蚯蚓"的生态农业生产方式,实现有机物料资源化利用,并提高蔬菜产量和品质。通过2个田间试验,研究不同种蚯蚓(赤子爱胜蚓和威廉腔环毛蚓)和有机物料种类及施用方式对菠菜产量与品质的影响。结果表明:腐熟牛粪的施用方式对菠菜产量和品质无显著影响,在施用腐熟牛粪条件下,接种赤子爱胜蚓和威廉腔环毛蚓分别提高菠菜产量32.78%~37.23%和18.70%~44.26%。而且,接种赤子爱胜蚓显著提高菠菜的可溶性糖和维生素C含量,同时降低了菠菜中Cu、Pb和Zn含量。另外,本研究还发现赤子爱胜蚓对菠菜产量和品质的影响与有机物料种类有关,综合效果最佳的处理是施用腐熟牛粪+食用菌渣+赤子爱胜蚓。     

2种蚯蚓对土壤锌离子的生理响应对比研究

以赤子爱胜蚓、太平二号蚓为试验对象,分别对其展开锌离子的急性毒性试验、耐受限度试验和富集效应试验,探究利用蚯蚓对污染土壤进行联合修复的可能性。结果表明,赤子爱胜蚓的LC50值在800～1200mg/kg,太平二号蚓的LC50值在400～800mg/kg;赤子爱胜蚓体重受Zn2+限制的程度低于太平二号蚓;赤子爱胜蚓及太平二号蚓对土壤中均具有一定的富集能力,赤子爱胜蚓对土壤中锌离子的富集量更大。     

利用农杆菌介导的共转化法获得含双Bt基因转基因水稻

利用农杆菌介导的共转化法,将分别携带不同Bt抗虫基因Cry1C*和Cry2A*的两个植物表达载体同时导入吉林省主栽超级粳稻品种吉粳88中,获得598株草丁膦(Basta)抗性独立转化植株。以Cry1C*和Cry2A*特异性引物对独立转化植株进行双引物PCR扩增,结果得到35株Cry1C*和Cry2A*基因双阳性植株,双基因共转化率为6.28%。以目的基因Cry1C*和Cry2A*为探针进行Southern blot分析,获得3个双基因单拷贝株系。利用RT-PCR检测Bt基因在这3个株系中的mRNA表达情况,结果表明两个Bt基因在转录水平上同时得到了有效表达。     

2种新的转双价基因棉种子越冬能力研究

以非转基因常规棉赣棉11号为对照,用当年的种子进行年前与年后室内发芽试验,研究转双价抗虫基因(Cry1Ac+Cry2Ab)棉和转双价抗虫抗除草剂基因(Cry1Ac+EPSPS)棉种子的越冬能力,结果表明:室内常温越冬后,转双价抗虫基因(Cry1Ac+Cry2Ab)棉的发芽势及发芽率并未衰减,反而有所增强,分别提高0.75和1.00个百分点,转双价抗虫抗除草剂基因(Cry1Ac+EPSPS)棉的发芽势及发芽率都提高1.00个百分点;而常规棉(CK)的发芽势及发芽率分别衰减2.75和1.00个百分点,这说明转双价抗虫基因棉及转双价抗虫抗除草剂基因棉种子的越冬竞争能力较强。此外,3个供试棉花品种的种子野外越冬均不能发芽和生苗,野外播种无越冬能力。     

接种蚯蚓对土壤过氧化氢酶和转化酶活性的影响

通过选择两种生活习性不同的威廉环毛蚓(Pheretima guillelmi)和赤子爱胜蚓(Eisenia foetida)在试验土壤中进行接种,跟踪测量蚯蚓－土壤交互作用下土壤过氧化氢酶和土壤转化酶活性的变化.结果表明,土壤经蚯蚓接种后,土壤过氧化氢酶和转化酶的活性发生了较大的变化,威廉环毛蚓对土壤过氧化氢酶活性的影响比赤子爱胜蚓要显著;赤子爱胜蚓对土壤转化酶活性的影响比威廉环毛蚓要显著.经组合接种后,土壤过氧化氢酶和土壤转化酶活性增加比较显著.产生这种差异性一个原因可能是与蚯蚓生活习性和接种土蚯蚓后引起土壤微生物类群的变化有关.     

怎样用蚯蚓作饲料

蚯蚓是一种多功能的生物资源,目前国内已普遍开展了蚯蚓的人工养殖和生产利用,并且已经取得了很大成绩,尤其是用作蛋白质补充饲料,已在全国普遍推广应用。一、蚯蚓的营养价值蚯蚓干重约为鲜重的12～21％,水分占鲜重的79～88％。在蚯蚓干体的化学成分中,主要有蛋白质、脂肪、碳水化合物和灰分。其中蛋白质的含量约占风干重的53.5～65.1％,蚓体中还含有丰富的畜禽所必需的氨基酸。经净化处理的赤子爱胜蚓、背暗异唇蚓、威廉环毛蚓的氨基酸含量都比较丰富,尤其是赤子爱胜蚓粉,优于豆饼和玉米,接近于鱼粉,和饲料酵母相比,各有优缺点。蚯蚓的不足之处是,除了赤子爱胜蚓以外,其它蚯蚓含硫氨基酸和赖氨基酸较     

呋喃丹对赤子爱胜蚓体内蛋白含量、SOD和TChE活性的影响

选用杀虫剂呋喃丹对赤子爱胜蚓(Eisenia foetida)进行染毒试验,研究农药污染对赤子爱胜蚓的致死率和体内蛋白含量、SOD和TChE活力的影响。结果表明:当蚯蚓受到低浓度呋喃丹胁迫时,其体内蛋白含量和TChE的活性均表现为升高趋势,SOD活性表现为降低;当受到高浓度呋喃丹胁迫时,其体内蛋白含量和TChE活性表现为不同程度的降低,SOD却表现为升高;其剂量-效应关系曲线均为抛物线型。可利用蛋白含量、SOD和TChE活性的变化来指示呋喃丹对赤子爱胜蚓的毒性作用。     

蚯蚓生态类型和密度对水浇地土壤综合质量的影响

【目的】通过室内培养试验探究不同生态类型蚯蚓及其接种密度对土壤物理指标、pH、养分指标和阳离子交换性能的影响，为改善华南地区水浇地土壤质量提供参考依据。【方法】分别设无蚯蚓对照土壤（CK）、低密度赤子爱胜蚓处理（E1: 20条，约10 g）、高密度赤子爱胜蚓处理（E2: 40条，约20 g）、低密度参状远盲蚓处理（A1: 2条，约10 g）和高密度参状远盲蚓处理（A2: 4条，约20 g），测定并对比分析不同处理的土壤机械组成、pH、养分含量和阳离子交换性能。【结果】与CK相比，各蚯蚓处理均显著降低土壤砂粒含量（P<0.05，下同），并显著提升土壤的粉粒和黏粒含量，然而相同密度下不同生态类型及相同生态类型下不同密度处理之间的机械组成均无显著差异（P>0.05）。低密度接种条件下，与CK相比，接种赤子爱胜蚓显著降低土壤pH，提高土壤交换性镁含量；接种参状远盲蚓显著降低土壤电导率和有效磷含量；相比接种赤子爱胜蚓处理，接种参状远盲蚓处理的土壤pH更高，电导率、有效磷和交换性镁含量更低。高密度接种条件下，与CK相比，接种2种生态类型蚯蚓均显著降低土壤电导率，而接种参状远盲蚓也显著降低有效磷含量及显著提高土壤pH、有机碳含量和碳氮比；相比接种赤子爱胜蚓处理，接种参状远盲蚓处理的土壤pH、有机碳、碳氮比更高，有效磷含量更低。主成分分析结果显示，接种蚯蚓处理的土壤养分含量、阳离子交换性能及黏粒、粉粒含量均有提高；接种高密度蚯蚓有更高的碳氮比、有机碳、粉粒和黏粒含量，接种低密度蚯蚓有更高的砂粒和有效磷含量；赤子爱胜蚓处理的阳离子交换性能、电导率、有效磷、全氮和碱解氮含量较高，参状远盲蚓处理的pH、碳氮比和有机碳含量较高。基于主成分分析结果进行土壤综合质量指数计算，结果显示接种低密度赤子爱胜蚓处理可显著提高土壤综合质量。【结论】在华南地区水浇地的生态管理上，选择接种低密度的表栖型赤子爱胜蚓有利于改变土壤质地并提升土壤综合质量。     

Initial function determination for the open reading frame (ORF) region of Pib gene via rice transformation

Three plant binary expression vectors—pNAR501, pNAR502 and pNAR503—were constructed, carrying fragments of exon2-exon3, 5′partial deletion exon1 and 5′partial deletion exon1-exon2-exon3 of Pib gene driven by 35S. These three vectors were transformed into the japonica rice variety Nipponbare through agrobacterium-mediated transformation. More than 30 transgenic rice plants were obtained and confirmed by polymerase chain reaction (PCR), Southern hybridization and the hygromycin resistance test in seed germination of their progeny. A rice blast resistance test for in vitro leaves of T_o transgenic plants in the tillering stage showed higher resistance to the races of E1, F1 and G1 of rice blast than that of the control Nipponbare. However, results of rice blast resistance test for seedlings of T₁ transgenic plants in the 3-to 4-leaf stage were different. All T₁ transgenic seedlings had a lower level of resistance to E1, F1 and G1 races than that of the control Nipponbare. Translated from Journal of Nanjing Agricultural University, 2006, 29(3): 1–5 [译自: 南京农业大学学报]     

二穗短柄草DREB1G基因表达模式分析和在拟南芥中的异源过量表达

CBF/DREB基因是一类植物特异性转录因子,通过调控下游抗逆相关基因的表达,在植物逆境胁迫应答过程中发挥重要作用。为了研究新型的禾本科模式植物二穗短柄草DREB基因的功能,克隆BdDREB1G的cDNA序列,分析该基因在各种非生物胁迫条件下的表达,构建过表达载体,获得转基因拟南芥。结果表明:高温、低温和水杨酸胁迫处理显著诱导BdDREB1G的表达,暗示该基因响应高温和低温胁迫。初步观察结果说明,该与野生型拟南芥相比,转基因植株生长发育迟缓。这些为研究该基因响应非生物胁迫的功能奠定基础。     

Cloning and Analysis of the Promoters of Two OsRhoGDIs Genes from Rice

OsRacD, belonging to rice （Oryza sativa L. ssp. japonica） Rho family of the small GTPases, is a pivotal gene involved in rice photoperiod fertility conversion of photoperiod sensitive genic male sterile rice which influences the rice fertility via controlling the pollen tube growth. Using OsRacD as bait, two GDP dissociation inhibitor genes designated as OsRhoGDI1 and OsRhoGDI2 were screened by yeast two-hybrid system. To further study the regulation mechanism of OsRhoGDIs, and also the relationships between OsRhoGDIs and OsRacD, the upstream regulation sequences of OsRhoGDI1 and OsRhoGDI2 were cloned by PCR, and the cis-elements in the promoters were predicted using internet tools in this study. The results showed that there were several kinds of response elements, such as phytohormone response elements, lightregulated elements and adversity induced elements in the promoters of OsRhoGDIs, some of which were similar. Comparing with the promoter of OsRacD, several pollen tube germination related elements were found. On one hand, the expression and regulation mechanism of OsRhoGDIs were complex, they may be regulated by several signaling pathways commonly, and the regulation mechanisms were similar to some extent. On the other hand, it was suggested that the fertility of photoperiod sensitive genic male sterile rice may be controlled by the cooperative expression of OsRhoGDIs and OsRacD.     

盐胁迫下OsDSR2RNAi转基因水稻的主要农艺性状分析

为探究盐胁迫下 OsDSR2 抑制表达后对水稻主要农艺性状和产量的影响,进一步阐明 OsDSR2 参与调控水稻耐盐机制,以野生型植株中花11(ZH11)和 OsDSR2 RNAi转基因水稻不同株系DR14和DR20为试验材料,植株幼穗分化期进行0.15mol/L盐胁迫处理,待植株完熟后测定穗长、每穗总粒数、每穗实粒数、结实率、单株穗数、单株粒质量、千粒质量等主要农艺性状,并进行相关性和主成分分析。结果表明,盐胁迫后 OsDSR2 RNAi转基因水稻的单株穗数、结实率均显著高于野生型ZH11,正常条件下, OsDSR2 RNAi转基因水稻单株粒质量均显著低于野生型ZH11,盐胁迫后二者均降低但无显著性差异。与正常条件下相比,盐胁迫处理后 OsDSR2 RNAi转基因植株的千粒质量与每穗实粒数呈显著负相关性,单株粒质量与穗长、结实率、千粒质量呈负相关性,单株穗数与穗长、结实率呈负相关性,单株粒质量与每穗总粒数呈正相关性;结实率与穗长、每穗总粒数呈负相关性。盐胁迫处理后 OsDSR2 RNAi转基因水稻第一、二主成分中,分别是单株粒质量和结实率特征向量最大。综上,盐胁迫下 OsDSR2 的抑制表达主要通过抑制水稻植株的单株穗数、结实率和单株粒质量的降低,协调单株粒质量与单株穗数、结实率、千粒质量的关系以及结实率与千粒质量、单株穗数的关系,从而调控盐胁迫下水稻的产量。     

转抗坏血酸过氧化物酶基因（APX）菊苣抗旱相关生理特性

在获得转抗坏血酸过氧化物酶基因(APX)菊苣株系的基础上,对3个转基因株系的抗旱相关生理特性进行分析。结果表明：正常供水条件下,转基因株系与非转基因植株各项生理指标差异不显著;干旱条件下, 3个转基因株系菊苣叶片APX质量摩尔浓度均显著高于非转基因对照。3个转基因株系中有2个株系丙二醛（MDA）质量摩尔浓度低于非转基因对照,表明APX基因可显著提高菊苣APX活性,降低氧自由基浓度,减轻对细胞的伤害。3个转基因株系叶片相对含水量、脯氨酸质量分数、叶绿素质量分数、单株幼苗的根系生长量,均高于非转基因对照。可见,转APX基因菊苣具有较强的抗旱能力,从而验证APX基因的抗旱功能,为转基因材料应用于菊苣的抗旱育种提供依据。     

花绒寄甲Caspase-1基因在不同虫态的表达

为明确花绒寄甲Caspase-1基因及其蛋白酶活性在不同虫态中的表达特性,采用RT-PCR和Caspase酶活测定方法进行研究。结果表明:Caspase-1酶活性变化趋势与其基因表达趋势基本一致,Caspase-1基因及其蛋白酶在5龄幼虫期和蛹期有较高的活性,且其活性高于所有成虫期;成虫期随存活时间的延长,Caspase-1酶活性总体呈现增长趋势。Caspase-1及其酶活在不同虫态中表达规律的揭示,为研究Caspase-1基因与花绒寄甲生理代谢和寿命关系提供依据。     

OsRAC3调控水稻籽粒性状的功能分析

目的 RAC/ROPs是一类植物特有的小G蛋白，作为分子开关参与众多的植物信号转导途径。OsRAC3通过与细胞分裂素信号组分互作，调控水稻Oryza sativa L.冠根发育，但是否影响水稻地上部分的性状尚不清楚。本研究主要分析OsRAC3对水稻穗发育以及产量性状的影响。方法 对水稻OsRAC3promoter::GUS转基因植株的花序组织进行GUS染色，分析OsRAC3的表达模式。分析转基因材料持续激活型突变体(CA-osrac3)和显性失活型突变体(DN-osrac3)2种试验材料的每穗粒数、粒长、粒宽、千粒质量等主要农艺性状。结果 OsRAC3在花序分生组织、雄蕊和雌蕊中强烈表达；CA-osrac3株系的穗粒数减少，籽粒更加饱满，粒长、粒宽、千粒质量均显著高于野生型；DN-osrac3株系的穗分枝数少且育性差，籽粒粒长、粒宽、千粒质量则显著低于野生型。结论 OsRAC3影响水稻穗的发育过程，持续激活型突变体CA-osrac3促进水稻籽粒的发育；OsRAC3在水稻高产育种和遗传改良中具有重要的应用价值。     

Insecticidal Constructure and Bioactivities of Compounds from Ficus sarmentosa var. henryi

Insecticidal activities of the petroleum ether-, chloroform-, ethyl acetate-, and water-soluble fractions of the methanolic extract of Ficus sarmentosa var. henryi were assayed against Musca domestica adults. The chloroform- and ethyl acetate-soluble fractions were the most active with 92.6 and 88.9% mortalities (24 h after treatment) respectively. Therefore, the two fractions were combined and four compounds, isolated from the fractions by activity-guided fractionation, were elucidated as 7-hydroxycoumarin, apigenin, eriodictyol, and quercetin by spectroscopic method and displayed excellent insecticidal activities against adults of M. domestica and 4th instar larva of Aedes albopictus. Among those, 7-hydroxycoumarin showed the strongest insecticidal activities with lethal concentrations (LC₅₀) values of 72.13 μg g⁻¹ sugar and 4.87 μg mL⁻¹ (48 h after treatment) against the test insects respectively. The cytoxicities of these compounds on BTI-Tn-5Bl-4 cell were also investigated for the insecticidal mechanism and found that quercetin represented superior inhibitory activity with MTT assay and reactive oxygen species (ROS) against BTI-Tn-5Bl-4 cell, but slightly weaker than that of the positive control (azadirachtin) and significantly greater than the negative control (DMSO only). Meanwhile, eriodictyol demonstrated the strongest effect on the mitochondrial membrane potentials (MMP). In conclusion, based on their comparative toxicities to commercial insecticides and their cytotoxic effects, some of the compounds from the F. sarmentosa have potential as botanical insecticides.     

间作对伴矿景天与红背桂花生长及镉锌吸收的影响

为探讨不同根系分隔方式间作对植物生长和Cd、Zn吸收的影响,以伴矿景天与红背桂花为供试植物,开展盆栽及田间试验。盆栽试验设置5个处理：伴矿景天与红背桂花塑料板分隔间作（地上部作用,COP）;伴矿景天与红背桂花300目尼龙网分隔间作（土壤溶液作用,COM）;伴矿景天与红背桂花无分隔间作（完全作用,CO）;红背桂花单作（EcM）;伴矿景天单作（SpM）。田间试验仅作无分隔间作处理。结果表明,COP条件下伴矿景天生物量略高于SpM处理,较COM及CO条件下高达100%、149%,差异显著。COP条件下每株伴矿景天的Cd、Zn吸收量分别较SpM显著高28.3%和14.0%。随着根系相互抑制作用的逐渐增强,COM和CO条件下伴矿景天每株Cd、Zn吸收量均显著低于COP及SpM处理。红背桂花的生长及Cd、Zn吸收性趋势与伴矿景天相似。田间条件下,因植物间距较大等因素,间作对二者生长及Cd、Zn吸收性均无显著影响。盆栽条件下,近距离间作时伴矿景天与红背桂花的根系相互作用将显著影响其生长和Cd、Zn吸收,田间条件下保持适宜的植物间距则不会出现抑制作用,二者在田间间作实现Cd/Zn污染土壤的边生产边修复是可行的。     

Functional Characterization of a NEM1-Like Gene in Magnaporthe oryzae

Magnaporthe oryzae, a filamentous ascomycete fungus, is well known as the causal agent of rice blast. With the technology of suppression subtractive hybridization (SSH), it was previously found that MGG_06001 (or named MoNEMl), a gene of M. oryzae homologous to the NEMl (nuclear envelope morphology protein 1) gene of baker's yeast (Saccharomyces cerevisiae), is differentially expressed between the mature appressium and the conidium and mycelium. This study aimed to characterize the function of MoNEMl gene by knocking it out using the method of target gene replacement. The AMoneml mutants exhibited reduced mycelial growth and conidiation. However, disruption of MoNEMl gene does not affect the pathogenicity of M. oryzae on barley and rice.