马铃薯花青素转录激活基因(stmyc)的克隆及表达分析

花青素（anthocyanin）为糖苷类物质,属于类黄酮色素。在花青素生物合成途径中,调控基因主要编码R2R3-MYB、bHLH和WD40转录因子,共同调控花青素合成的结构基因的表达,进而控制花青素在植物中的时空表达和沉积的模式（Holton and comish,1995）。本研究以紫色马铃薯为材料,克隆花色素合成的转录激活基因stmyc并研究其表达模式,为阐明马铃薯花色素生物合成和沉积机制提供依据。     

草酸胁迫下拟南芥三个差异表达microRNA的分析

草酸是多种植物病原真菌(如核盘菌,灰葡萄孢等)的重要致病因子,这些草酸产生菌在侵染植物时,会分泌草酸来酸化寄主组织,使得病原真菌的细胞壁降解酶能更迅速地协同发挥作用,加速细胞的降解,而且草酸可夺取寄主细胞壁的钙离子形成草酸钙结晶从而破坏寄主细胞壁。植物microRNA广泛参与植物生长发育各阶段的基因表达调控,在抗生物或非生物胁迫的应答中发挥着重要的作用。本研究基于植物microRNA芯片研究3周龄拟南芥(Arabidopsis thaliana)在30 mmol/L草酸胁迫下microRNA的表达。获得3个差异表达的microRNA:miRNA-2988(Ptc-miR3911),miRNA-3090(Ath-miR858),miRNA-3131(Ppt-miR1211)。根据psRNATarget,对下调表达的小立碗藓(Physcomitrella patens)Ppt-miR1211的同源物,找到一个最匹配的靶mRNA是At5g35753;另一下调表达的毛果杨(Populus trichocarpa)Ptc-miR3991(与Ath-miR399b同源)的同源物,对应的靶mRNA是一个泛素连接酶(At2g33770);上调表达的AthmiR858有4个靶mRNA,分别为At2g47460(AtMYB12)、At5g49330(AtMYB111)、At1g06180(AtMYB13)和At1g66230(AtMYB20)。草酸胁迫下,qRT-PCR对其中下调表达的两个Ppt-miR1211及Ptc-miR3991同源物的靶基因的表达情况分析表明,At5g35753及At2g33770在草酸胁迫2 h后被诱导表达,于12 h达到峰值,24 h后表达量下降;qRT-PCR还表明:在草酸胁迫1 h后,Ath-miR858确实被诱导表达,于12 h达到峰值,24 h后表达量下降,其靶基因MYB在草酸胁迫早期(2 h内)多数有下调的趋势,表明microRNA对其靶mRNA确有负调控。此外,还对Ath-miR858与Ath-miR399b的启动子进行了生物信息学分析。本研究结果为microRNA在拟南芥抗草酸胁迫中的作用提供了依据。     

降低UV-B辐射强度对小麦类黄酮含量及分布的影响

在降低目前南京地区近地面UV—B辐射强度的条件下,研究了小麦叶片类黄酮含量及分布的变化。试验结果表明：UV—B紫外辐射强度降低,类黄酮含量显著下降,拔节—孕穗期最为敏感;小麦上部叶片类黄酮含量变化显著大于中、下部叶片,在单张叶片中,叶片基部类黄酮含量显著较叶片尖、中部低。UV—B紫外辐射强度降低,小麦叶片叶绿素含量增加。     

六种生物降解地膜对马铃薯生育期及产量的影响初探

为了在马铃薯生产中推广使用安全环保的地膜,选择6种生物可降解地膜进行试验,研究其对马铃薯生育期及产量的影响,结果表明:1)K9-8和V1-8两个类型的降解膜降解时间短,适合中、低海拔地区早市早熟马铃薯品种地膜覆盖栽培需要。2)K9-10和V1-10两个类型的降解膜降解时间长一些,能够满足中晚熟马铃薯品种的地膜覆盖栽培需要。3)K9-12和V1-12两个类型的降解膜降解时间较长,不太适合本地区的马铃薯地膜覆盖栽培需要。     

广西免耕马铃薯种出“世界新模式”

时下，正是广西免耕马铃薯收获的季节。近日，广西壮族自治区组织有关专家组来到武鸣县双桥镇八桥村现场验收免耕马铃薯。经测产，免耕马铃薯平均单产达到2564kg／667m^2，超过全国马铃薯平均单产1．5倍以上。国内外著名马铃薯专家孙慧生称：“广西稻草覆盖免耕栽培马铃薯，创新了我国马铃薯栽培新模式，也是世界新模式，对促进粮食增产、农业增效和农民增收具有十分重要的意义。我们要把这一新的栽培模式补充到《中国马铃薯栽培学》书中。”     

单/间作条件下氮肥水平对水稻总酚和类黄酮的影响

采用田间试验研究了主栽杂交稻合系-41和优质糯稻黄壳糯间作条件下,高氮供应（N300）对易感病品种黄壳糯叶片中总酚和类黄酮含量、生物量、产量和稻瘟病发生的影响。结果表明,高氮供应（N300）显著降低了单作黄壳糯的生物量、产量及体内总酚和类黄酮含量,增加了稻瘟病发生的严重程度;而对间作黄壳糯产量和总酚含量无显著影响,对类黄酮含量的降低及稻瘟病发生的增加不及单作显著。在低氮供应 (N180)下,合系-41与黄壳糯间作中黄壳糯较其单作明显提高了的生物量和产量,提高了间作黄壳糯叶片中总酚和类黄酮含量,降低了稻瘟病发生。黄壳糯叶片中总酚和类黄酮含量与稻瘟病的发生呈负相关趋势。     

微生物菌剂对张北冷凉坝上地区马铃薯产量、品质及活化土壤磷钾的效果

采用田间小区试验,以溶磷解钾微生物菌剂(功能菌为巨大芽孢杆菌和胶质芽孢杆菌)和"冀张薯12""紫色马铃薯""大西洋"3个品种马铃薯为研究对象,探讨了在张北冷凉坝上地区施用微生物菌剂对不同品种马铃薯产量、品质、土壤速效磷钾以及土壤磷酸酶活性的影响。结果表明:施用微生物菌剂可以显著增加3个品种马铃薯的产量(P0.05),其中"冀张薯12"产量增幅最大为24.54%,其他2个品种产量平均增幅仅为6.96%;可以改善马铃薯的品质,尤其是紫色马铃薯,其Vc、可溶性蛋白质和可溶性糖含量均显著提高,增幅分别为320.21%,29.40%,7.12%(P0.05)。此外,菌剂施用调控着整个生育期土壤速效磷钾的含量和相关土壤酶活性,种植"大西洋"和"冀张薯12"的土壤施用菌剂后速效磷钾含量高于对照且在中后期达到显著水平,同时提高了土壤酸性和碱性磷酸酶的活性,平均增幅为37.52%和32.30%。总之,溶磷解钾微生物菌剂适用于冷凉张北坝上地区马铃薯种植,且对"冀张薯12"的促生效果最为明显。     

基因工程及花色素合成途径在花色改良中的研究进展

随着经济的发展,转基因技术在花卉中的应用也越来越关键,通过基因工程使花色发生变化已经在商业中运用。生物碱类、胡萝卜素和类黄酮化合物等一类活性物质共同作用使植物的花色更具观赏性,目前,在花色改良上应用较多的是类黄酮色素,其研究机理也较为清楚。本文主要介绍了F3＇5＇H（类黄酮-3＇,5＇-羟基化酶）和F3＇H（类黄酮-3＇-羟基化酶）的生物合成途径,阐述了基因工程技术对花色修改的作用,综述了目前花卉转基因工程的研究现状和花色素合成的主要途径及影响因素,同时简单介绍了目前改良花色的几种策略和就其存在的问题、发展前景做出了讨论。     

马铃薯叶片特异性启动子克隆分析及其驱动的人白细胞介素-12表达载体构建

利用植物生物反应器表达具有应用价值的药用蛋白是近年来生物技术研究热点。本研究利用PCR技术从马铃薯基因组中克隆其叶片组织特异性启动子Prbcs后,利用PLACE等数据库对序列进行启动子元件分析,接着通过重叠PCR、限制性内切酶酶切、体外连接、转化等技术,以双元载体pCambia-2301为骨架,构建Prbcs驱动的药用蛋白人白细胞介素12（hIL12）表达载体。结果显示,本研究成功从马铃薯基因组中扩增得到了623 bp大小的启动子片段,序列分析表明该片段含有典型的启动子元件如TATA-box,以及一些组织特异性的相关响应元件,符合组织特异性启动子的特征;成功获得了基于pCambia-2301的Prbcs驱动的hIL12植物表达载体。本研究获得的Ppatatin启动子及其驱动的hIL12表达载体,为提高hIL12在植物生物反应器中表达量以及优化马铃薯生物反应器打下了基础。     

富镉复垦基质中添加改良剂对作物镉含量及产量的影响

为了选择出可以在基质中安全种植的作物,该文研究了在粉煤灰加酒糟（基质Ⅰ）和粉煤灰加污泥（基质Ⅱ）混合成的基质中添加柠檬酸和熟石灰作为改良剂,对芸豆、豇豆和马铃薯镉含量及产量的影响。结果表明：熟石灰可以提高作物出苗率,柠檬酸对作物出苗率影响不大。熟石灰可以显著提高基质Ⅰ中芸豆和豇豆以及2种基质中马铃薯的产量,柠檬酸降低基质Ⅰ中芸豆和豇豆的产量,达到显著水平,而对马铃薯在2种基质中的产量影响都不显著。熟石灰可以显著降低芸豆和马铃薯果实中镉的含量,柠檬酸却可以显著增加芸豆果实中镉的含量。粉煤灰加酒糟中可以安全种植的作物为芸豆、豇豆和马铃薯;粉煤灰加污泥中可以安全种植的作物为马铃薯。     

Antioxidant capacity manipulation in transgenic potato tuber by changes in phenolic compounds content

The main goal of this study was to generate potato tubers with increased levels of flavonoids and thus modified antioxidant capacities. To accomplish this, the vector carrying multigene construct was prepared and several transgenic plants were generated, all overexpressing key biosynthesis pathway enzymes. The single-gene overexpression or simultaneous expression of genes encoding chalcone synthase (CHS), chalcone isomerase (CHI), and dihydroflavonol reductase (DFR) resulted in a significant increase of measured phenolic acids and anthocyanins. The increase in phenolic compounds synthesis is accompanied by decreases in starch and glucose levels in transgenic plants. The flavonoids-enriched plants showed improved antioxidant capacity; however, there is a complex relationship between antioxidant capacity and flavonoids content, suggesting the great participation of other compounds in the antioxidant potential of the plants. These other compounds are not yet recognized.     

patatin启动子调控烟草液泡转化酶抑制子在马铃薯抗低温糖化中的作用

为了降低马铃薯(Solanum tuberosum )块茎低温贮藏下还原糖的积累，实验构建了马铃薯块茎特异性启动子（CIPP）调控的烟草液泡转化酶抑制子Nt-VIF基因表达载体pBICNI，并转化马铃薯植株。PCR、Northern杂交和Southern杂交分析结果显示，CIPP调控的Nt-VIF基因全长cDNA成功地导入鄂马铃薯3号（E-3）植株。14个转基因株系块茎分别贮藏在4℃和20℃条件下，贮藏1个月后进行还原糖含量和液泡酸性转化酶（VI）活性测定。结果表明，在20℃条件下转基因株系块茎还原糖（RS）含量与对照相比差异不明显，在4℃条件下RS含量则显著下降，与对照相比下降幅度从34%（株系B-13）至76.8%（株系B-1），说明Nt-VIF cDNA在马铃薯中的表达，成功地抑制了液泡酸性转化酶的活性，导致还原糖含量降低。进一步分析表明，转基因块茎低温贮藏其液泡转化酶活性与还原糖含量呈显著的正直线相关（VI = 0.3084RS + 0.0673）。实验获得的B-1、B-2、B-6、B-9、B-14等5个转基因株系，块茎低温贮藏后能直接满足炸片加工对还原糖含量的要求。     

Increase in lipid content in potato tubers modified by 14-3-3 gene overexpression

Recently, transgenic potato plants were created with overexpression of the 14-3-3 protein derived from Cucurbita pepo. Detailed analysis of those plants suggested that the function of the isolated 14-3-3 isoform is in the control of carbohydrate and lipid metabolism in the plants. 14-3-3 protein overexpression gave rise to an increase in soluble sugar and catecholamine contents in both leaves and tubers. It is proposed that 14-3-3 protein affects carbohydrate metabolism in potato plants via regulation of catecholamine synthesis. Furthermore, genetically modified potato tubers with 14-3-3 protein overexpression showed changes in lipid content and composition. The transgenic potato tubers contained 69% more total fat compared to the wild-type plant. Separation of tuber lipids into polar and nonpolar fractions revealed that the transgenic potato tubers contained almost 3 times more nonpolar lipids than the control. Analysis of fatty acid composition, conducted by the means of gas chromatography, showed that linoleic acid was the main fatty acid present in the tubers of both modified and control potato plants. In the nonpolar fraction of the fat of the transgenic tubers the unsaturated fatty acids exhibited a higher participation in the sum of all fatty acids.     

水稻赖氨酸和总黄酮含量的QTL定位及环境互作分析

为挖掘多环境下稳定存在的水稻赖氨酸和总黄酮含量相关QTL,以粳稻东农425和长白10号及其衍生的180个株系的F_(6:7)重组自交系(RIL)作为供试群体,采用完备区间作图法(ICIM)和基于混合线性模型的复合区间作图法(MCIM),对2014年和2015年水稻的赖氨酸含量和总黄酮含量进行加性QTL定位及环境互作分析。结果检测到10个影响赖氨酸含量的加性效应QTL和12个影响黄酮含量的加性效应QTL,分布在除第9、第10和第12染色体以外的9条染色体上,其中在第5染色体的RM538~RM1271标记区间内连续2年检测到总黄酮含量QTL。检测到6个存在环境互作效应的赖氨酸含量QTL、4个存在环境互作效应的总黄酮含量QTL,互作贡献率为0.15%~6.73%;一对影响总黄酮含量的上位互作效应的QTL,贡献率为0.99%。本研究结果为水稻赖氨酸和总黄酮含量QTL分子标记辅助育种提供了一定的理论依据。     

种植与组培9年的转AhDREB1基因毛白杨耐盐性比较

为了对栽培9年后转AhDREB1基因毛白杨的耐盐性进行评价,并探究田间种植与组培继代2种不同生长条件对转基因毛白杨耐盐性的影响,以从大田种植9年的转AhDREB1基因杂种毛白杨[(Populus tomentosa×Populus bolleana)×P.tomentosa]中获得的株系(T46-F)和组培继代9年的2个转基因杂种毛白杨株系(T46、T12),以及非转基因杂种毛白杨株系(CK)为试材,通过对外源基因进行PCR检测,发现9年后AhDREB1基因仍然稳定整合在转基因植株中。通过不同浓度的Na Cl胁迫试验,对各株系的相对电导率进行分析,进一步选择测定0.6%浓度Na Cl处理水平下各株系的株高、地径生长量、超氧化物歧化酶(SOD)活性、过氧化物酶(POD)活性、丙二醛(MDA)含量、脯氨酸含量和叶绿素相对含量,对不同转AhDREB1基因毛白杨株系的耐盐性进行分析和比较。结果表明,所有株系的相对电导率、MDA含量均随Na Cl浓度增加而增大,但高盐浓度下转基因株系T46-F、T46、T12的相对电导率、MDA含量均显著低于CK。无论是在非盐条件,还是0.6%Na Cl胁迫下,AhDREB1基因的导入均能显著提高毛白杨植株的SOD、POD活性,脯氨酸含量。其中0.6%浓度Na Cl胁迫下,T46-F、T46、T12的SOD活性分别是CK的2.51倍、3.20倍、2.55倍;POD活性分别是CK的1.23倍、1.63倍、1.10倍;脯氨酸含量分别是CK的1.51倍、1.69倍、1.62倍。此外,转AhDREB1基因毛白杨叶绿素含量下降显著低于CK,同时株高、地径生长量显著高于CK。组培继代9年的转基因株系的脯氨酸含量、株高生长量显著高于在大田种植9年的转基因毛白杨,而相对电导率显著低于大田种植转基因株系,说明组培继代培养更有利于保持转基因毛白杨的耐盐性。综上可知,转AhDREB1基因显著提高了毛白杨的抗氧化酶活性,调控渗透调节物质的积累,从而减轻细胞膜的氧化损伤,降低了相对电导率,提高了毛白杨的耐盐性,综合考虑各个指标,各株系耐盐性的大小顺序为T46T12T46-FCK。本研究为通过转AhDREB1基因提高植物耐盐性提供了理论参考。     

Cultivation of transgenic cyanophycin-producing potatoes does not negatively affect growth,reproduction and activity of the earthworm Lumbricus terrestris (L.)

A microcosm experiment was performed to investigate the effects of post-harvest potato tubers from transgenic cyanophycin-producing potatoes on Lumbricus terrestris (L.) activity and biomass, number of cocoons and their hatchability as well as the remaining cyanophycin content in soil and cast samples during a period of 80 days. Potato tubers from four transgenic potato events with different cyanophycin content in a range from 0.8 to 7.5% were compared to the near isogenic, non-transgenic control (Solanum tuberosum L. cv. Albatros) and a comparative potato cultivar (S. tuberosum L. cv. Désirée). One treatment with transgenic tuber residue but without earthworms was prepared as an additional control. Potato tuber loss from the surface of the microcosms was significantly higher in the treatments with transgenic potato tubers compared with non-transgenic treatments. It can be estimated that the earthworm contribution to potato tuber loss from the soil surface was approximately 61%. Mean number of cocoons in addition to the number of hatched cocoons varied from 2.6 to 6.2 and from 7 to 15 accounting for 45.2–83.35% hatchability, respectively, but no significant differences between the treatments were found. The same was true for the development of earthworm biomass in the various treatments. The cyanophycin content in soil samples was significantly higher when earthworms were present indicating that the cyanophycin content in the upper soil layer might have been enhanced through earthworm burrowing activity. Overall, it is concluded that tubers from transgenic cyanophycin potatoes are easily degradable and neither inhibit nor stimulate earthworm growth, reproduction, and activity.     

Repression of the 14-3-3 gene affects the amino acid and mineral composition of potato tubers

Recently, transgenic potato plants were created showing underexpression of the 20R isoform of the 14-3-3 protein. The transgenic plants grown in tissue culture showed a significant increase in nitrate reductase activity and a decrease in nitrate level. The transgenic line with the lowest 14-3-3 quantity was field-trialed (1997-2000) and analyzed. The reduction in the 14-3-3 protein level consistently resulted in a starch content increase and in an increase in the ratio of soluble sugars to starch in the tubers, although the latter was only barely visible. The determination of amino acid composition in the tubers showed a significant increase in methionine, proline, and arginine content and a slight but consistent increase in hydrophobic amino acid and lysine content in the cells of the transgenic potato plants. We also observed an increase in the crude protein content, from 19 to 22.1% of the control value in consecutive years. It is proposed that all of these changes might have resulted from the downregulation of nitrate reductase and sucrose phosphate synthase activities by 14-3-3, although other potential mechanisms cannot be excluded (e.g., an increase in enzyme protein level). 14-3-3-repressed transgenic plants showed a significant increase in calcium content in their tubers. It is thus proposed that a function of the isolated 14-3-3 isoform is in the control of amino acid synthesis and calcium metabolism. However, the mechanism of this control is as yet unknown.     

Effect of potato plants expressing snowdrop lectin (GNA) on the performance and colonization behaviour of the peach-potato aphid Myzus persicae

Abstract

Transgenic potato plants expressing snowdrop lectin (GNA potatoes) are developed to increase resistance against sap-feeding insects. When expressing GNA at relatively high levels such potatoes may have a negative effect on the fecundity and development of the first generation of the important pest, the peach-potato aphid Myzus persicae (Sulzer) (Hemiptera: Aphididae). However, the effects on M. persicae over several generations, and how such plants affect the alate aphids’ colonization behaviour have not been reported. In this laboratory study, the performance of single M. persicae on potatoes with low GNA expression, measured as developmental time, fecundity, size and survival, was compared with the control, following two successive generations of single apterous aphids. Aphid population growth on the two plant lines was also studied. In addition, colonization behaviour was investigated in a choice experiment where the alate aphids could choose between the GNA and an isogenic control potato plant in a cage. The present study showed that the apterous aphid performance was not significantly different on the two potato lines, although the aphids tended to perform slightly poorer on the transgenic potato. However, the transgenic potato was less likely to be colonized by alate aphids. It is concluded that such transgenic potato plants expressing the lectin at a relatively low level, maximum 0.2% of the soluble protein, have no significant impact on the performance of apterous M. persicae once on the plant, but may have a potential in controlling the aphids by altering the colonization behaviour of alates.