质子对植物蔗糖转运蛋白和己糖转运蛋白活性的调节

蔗糖是植物体内进行长距离运输的主要营养物质和信号分子。蔗糖转运蛋白和己糖转运蛋白是蔗糖从源到库的运输过程中的重要参与者。质子不仅通过形成质子动子势为蔗糖和己糖逆浓度梯度跨膜转运提供动力,还调节它们的活性,从而影响蔗糖的运输和分配。该文总结了质子对蔗糖转运蛋白的调节和质子对己糖转运蛋白活性及功能的调节,并加以展望。     

拟南芥Mg2+转运蛋白AtMGT10的原核表达及多克隆抗体制备

【目的】在大肠杆菌系统中原核表达拟南芥叶绿体Mg~(2+)转运蛋白AtMGT10,通过免疫健康成年家兔,获得针对AtMGT10蛋白的多克隆抗体,为研究AtMGT10蛋白在植物生长发育中的功能奠定基础。【方法】利用PCR技术克隆AtMGT10基因编码区187~786bp的cDNA片段,连接到pET-28a中构建原核表达载体pET-28aAtMGT10~(63-262),转化大肠杆菌BL21(DE3)菌株,经IPTG诱导表达带有His标签的His-AtMGT10~(63-262)截短重组蛋白。用镍柱亲和纯化后的重组蛋白作为抗原皮下注射免疫家兔,制备针对AtMGT10蛋白的多抗血清。提取拟南芥野生型叶片总蛋白,用AtMGT10多抗血清进行蛋白免疫印迹检测。【结果】经PCR扩增获得了600bp的AtMGT10基因cDNA片段,成功构建原核表达载体pET-28a-AtMGT10~(63-262),在大肠杆菌系统中表达出His-AtMGT10~(63-262)蛋白,经亲和纯化后免疫家兔获得了Anti-AtMGT10多抗血清。在拟南芥野生型总蛋白中用Anti-AtMGT10经免疫印迹检测到大约50ku的蛋白条带,利用抗原竞争试验进一步证明检测到的信号就是拟南芥内源AtMGT10蛋白。【结论】原核表达了His-AtMGT1063-262蛋白,成功制备了特异性较强的AtMGT10蛋白多克隆抗体。     

Strategies for increasing the selenium content of wheat

Selenium (Se) is essential for humans and animals but has no known function in plants. Excess accumulation is toxic to both plants and animals. Dietary intake of Se is low in a large number of people worldwide. This is due to low bioavailability of Se in some soils and consequently low concentrations of Se in plant tissues.Both selenate and selenite are taken up by plants and subsequently translocated around the plant. Selenate, an analogue of sulphate, is transported by the sulphate transporter family. Some plants are able to accumulate high internal concentrations of Se (hyperaccumulators); however, genetic variation in accumulation ability amongst non-accumulators such as cereals, is relatively small.Within plant tissues, Se enters the pathways for sulphate assimilation and metabolism and will replace cysteine and methionine in proteins, often with detrimental effect. Alternatively, Se may be accumulated as methylated derivatives or lost from the plant following volatilisation.Agronomic biofortification of crops with Se-containing fertilisers, which is practised in some countries, provides the best short-term solution for improving Se content of wheat. Longer-term genetic improvement, particularly by targeting substrate discrimination of transporters between selenate and sulphate, for example, may provide a means to enhance uptake and promote accumulation.     

武运粳7号超表达OsNRT1.2 后对硝酸盐的响应

 应用公开的植物基因组数据库和生物信息学分析确定了一个水稻低亲和硝酸盐运输蛋白候选基因OsNRT1.2。利用农杆菌介导Ubiquitin启动子过量表达OsNRT1.2,研究了该基因在武运粳7号中的获得性功能特征。OsNRT1.2的cDNA序列从KOME网站中获得,该序列全长为2178 bp,编码阅读框为1732 bp,编码533个氨基酸。采用半定量RT PCR技术分析,OsNRT1.2在武运粳7号中表达存在器官特异性,主要在根系表达,地上部分几乎不表达。通过将pUbi OsNRT1.2 在武运粳7号中转化,得到OsNRT1.2基因超量表达材料。0.2 mmol/L NO3-和5.0 mmol/L NO3-处理野生型（WT）和T2转基因植株OE1、OE2 和OE8  30 d,每10 d跟踪记录株高和根长。转基因植株（除OE8在5.0 mmol/L NO3-供氮水平外）与WT株高无明显差异,根长增加量显著降低。在0.2 mmol/L NO3-供氮水平下,转基因植株和WT地上部分和根系的全氮含量均无显著差异;在5.0 mmol/L NO3-供氮水平下,除OE1外,OE2和OE8地上部分全氮含量显著增加,3个转基因株系比WT根系全氮含量显著降低。在两个氮素水平处理条件下,转基因植株根冠比都显著降低;生物量比WT都增加,在0.2 mmol/L NO3-供氮水平下增加了9.4% ~ 31.1%,在5.0 mmol/L NO3-供氮水平下增加了12.5% ~ 43.8%。研究表明,OsNRT1.2基因在武运粳7号中可能参与了氮素从根系向地上部的转运,从而导致地上部生物量增加。     

盐胁迫下宁夏枸杞Na+吸收及Na+/H+转运蛋白与H+-ATPase基因表达的研究

为从分子水平揭示宁夏枸杞钠的吸收积累机理,本试验采用原子吸收分光光度法和实时荧光定量PCR(RT-qPCR)方法,对盐胁迫下宁夏枸杞根中Na⁺、K⁺含量以及质膜和液泡膜Na⁺/H⁺转运蛋白与H⁺-ATPase基因表达水平进行测定分析。结果表明,相同胁迫时间下,随着NaCl处理浓度的增加,枸杞根系中Na⁺浓度总体呈缓慢增加趋势,K⁺含量呈先增加后减少趋势,Na⁺/K⁺比值呈先减少后增加趋势;编码质膜和液泡膜的Na⁺/H⁺转运蛋白基因LbSOS1、LbNHX1以及液泡膜H⁺-ATPase基因LbVHA-C1表达量均呈升高趋势,质膜H⁺-ATPase基因LbHA1表达量呈先升高后降低趋势。相同NaCl处理浓度下,随着胁迫时间的延长,Na⁺含量总体呈增加趋势,K⁺含量呈先增加后减少趋势,Na⁺/K⁺比值呈增加趋势。LbSOS1、LbNHX1表达量总体呈先升高后降低趋势,LbVHA-C1、LbHA1表达量总体呈降低的趋势。相关性分析显示,不同胁迫时间下,枸杞根中LbSOS1、LbNHX1、LbVHA-C1和LbHA1表达量与Na⁺含量存在一定的正相关或负相关。上述结果表明,在低浓度NaCl胁迫时,维持枸杞体内较高的K⁺/Na⁺比值是宁夏枸杞耐盐的主要方式之一,同时也说明在胁迫初期,质膜和液泡膜的Na⁺/H⁺转运蛋白与H⁺-ATPase参与了枸杞细胞中Na⁺及时排出胞外和区隔于液泡,从而保持了根细胞内Na⁺的稳定性。此外,随着胁迫时间的延长和NaCl处理浓度的增加,LbSOS1、LbNHX1、LbVHA-C1和LbHA1的表达水平均降低,而 Na⁺积累量大幅增加,致使枸杞抗盐性降低。本研究揭示了宁夏枸杞的耐盐机理,为利用宁夏枸杞改良宁夏大面积盐碱地提供了理论依据。     

糖在苹果果实中卸载机制的研究

在建立了活体研究苹果果实糖卸载体系－“卸载穴技术”的基础上,对金冠苹果果实糖迅速积累期糖卸载的生理机制进行了研究。１ｍｍｏｌ／Ｌ ＰＣＭＢＳ,１ｍｍｏｌ／Ｌ ＤＮＰ,１ｍｍｏｌ／Ｌ ＥＢ和８００ｍｍｏｌ／Ｌ Ｂｅｔａｉｎｅ明显抑制苹果果实糖的卸载,１０＾－５ｍｏｌ／Ｌ ＡＢＡ,５ｍｍｏｌ／Ｌ ＥＧＴＡ和５０／Ｌ ＮＯ＾－３明显促进苹果果实糖的卸载。     

Taurine alone or in combination with fish protein hydrolysate affects growth performance,taurine transport and metabolism in juvenile turbot (Scophthalmus maximus L.)

The study was conducted to investigate the effects of taurine (Tau) alone or in combination with fish protein hydrolysate (FPH) on growth performance, the expression of Tau transporter (TauT) and metabolic profile in juvenile turbot. FM, FPH0, FPH0+T, FPH10 and FPH10+T diets, respectively, contained 300, 150, 150, 80, and 80 g/kg fishmeal. FPH10 and FPH10+T diets contained 62 g/kg FPH. FPH0+T and FPH10+T diets were, respectively, prepared by supplementing the FPH0 and FPH10 diet formulations with 8 g/kg Tau. Specific growth rate was the highest in FM group and the lowest in FPH10 group. TauT mRNA levels in fish fed Tau supplemented diets were significantly lower than that in Tau unsupplemented diets. NMR‐based metabolomics analysis showed that Tau contents in liver of FPH0+T and FPH10+T were significantly higher than that of FM, FPH0 and FPH10. In muscle, Tau contents were significantly decreased in the FPH10+T versus FPH0 and the FPH10+T versus FPH10 comparisons. In conclusion, 62 g/kg FPH to replace fishmeal may not affect Tau synthesis, transport and metabolism. However, Tau supplemented alone or in combination with a certain level of FPH could reduce the requirement for Tau synthesis and transport and increased Tau levels in muscle and liver.     

早期断奶对仔猪空肠和回肠兴奋性氨基酸载体1表达的影响

本试验旨在研究仔猪出生后10~20 d,早期断奶仔猪小肠谷氨酸转运载体基因表达情况与哺乳仔猪的差异。试验分别从40头不同母猪的仔猪中各选出体重相近,10日龄的"杜×长×大"三元杂交仔猪1头,共40头仔猪,随机不配对分为2组,每组20头仔猪,对照组(哺乳组)为哺乳仔猪,随母猪喂养;试验组(断奶组)为断奶仔猪,隔离断奶饲养;试验期10 d。饲养结束,每组随机取12只仔猪,宰杀取空肠和回肠,测定谷氨酸转运载体兴奋性氨基酸转运载体1(EAAC1)蛋白质表达情况和游离氨基酸含量。结果显示,断奶显著降低了仔猪空肠和回肠EAAC1(57和73 ku)及其相关蛋白谷氨酸转运联合蛋白(GTRAP3-18)(50 ku)的蛋白质和mRNA表达量(P0.05)。断奶提高了仔猪空肠游离谷氨酸和总氨基酸含量,却降低了仔猪回肠游离谷氨酸和总氨基酸含量,差异显著(P0.05)。结果提示,早期断奶降低EAAC1和GTRAP3-18的蛋白质含量,这可能与早期断奶仔猪遭受营养谷氨酸缺乏导致的肠道氨基酸吸收转运障碍有关。     

丽蚜小蜂糖转运蛋白基因克隆及其诱导表达

丽蚜小蜂Encarsia formosa Gahan是粉虱类害虫的重要寄生性天敌昆虫，补充糖分等营养可以延长丽蚜小蜂寿命及增加其产卵量，但对其糖转运相关蛋白基因及其糖诱导表达尚不清楚。本研究克隆获得丽蚜小蜂的糖转运蛋白EfST1基因，与其他寄生蜂的糖转运蛋白基因序列同源性达65%以上；进一步采用实时荧光定量qPCR技术检测发现，饲喂不同浓度的葡萄糖溶液均可显著提高EfST1基因的表达量，且饲喂不同时间段（2～48 h）后，丽蚜小蜂EfST1基因表达量均显著高于对照组，2 h和48 h处理组间差异不显著。综上，EfST1基因可能与糖分补充及其转运密切相关，该基因诱导表达受糖分浓度影响小。研究结果为进一步研究寄生蜂生物防治过程中补充营养及其转运机制奠定基础。