大豆GmNRT1.2a和GmNRT1.2b基因的克隆及功能探究

拟南芥中硝酸盐的吸收、转运和分配是通过硝酸盐转运蛋白(nitrate transporter, NRT)实现的。尽管之前的生物信息学分析推测大豆GmNRT1.2s可能参与共生固氮过程,但尚未开展相应的功能研究。本研究通过对其表达模式分析収现,GmNRT1.2a和GmNRT1.2b分别在根和叶中高表达,且受硝酸盐诱导,在接种根瘤菌与结瘤因子(nodfactors,NFs)后表达量明显升高。功能研究结果显示,过表达GmNRT1.2a或GmNRT1.2b后大豆根瘤数目显著增加。本研究为深入探究GmNRT1.2a和GmNRT1.2b调控大豆共生固氮过程的分子机制提供了一定的数据支持。     

棉花蔗糖合成酶基因家族在纤维发育期时空表达模式分析

蔗糖合成酶(Sucrose Synthase)是调控植物糖类代谢的关键酶之一,在植物生长发育及渗透调节过程中起着重要的作用。该酶为多基因家族,各成员在植物不同组织及不同发育阶段发挥不同的作用。陆地棉(Gossypium hirsutum)中有15个Sus成员,本研究利用转录组高通量测序及实时定量PCR技术,检测了该基因家族的表达模式。结果表明,Gh Sus1D、Gh Sus3A、Gh Sus3D和Gh Sus6D在起始期(0~3 DPA)大量表达;GhSus7D在10 DPA的纤维中表达量快速增加,达到同期突变体胚珠的4.7倍;Gh Sus1A、Gh Sus5D在纤维伸长期(5~15 DPA)表达量显著高于胚珠,分别在10 DPA、15 DPA时达到峰值;Gh Sus3A、Gh Sus3D、和Gh Sus6D在纤维发育(15 DPA)时表达量重新上升,且在20 DPA时仍处于较高表达水平。Sus基因家族组织表达模式分析表明,它们在陆地棉品种徐州142野生型和突变体中的组织表达模式基本相同,根、茎、叶、花中没有显著差异。但不同Sus基因具有不同的组织表达特异性。     

大鼠成熟的肺表面活性蛋白B的原核表达及纯化

为表达和纯化SP-B蛋白,先对SP-B基因进行稀有密码子优化,PCR反应获得SP-B片段,构建重组质粒pGEX4T-1/SP-B,转化到E.coli BL21(DE3)中诱导表达;用SDS-PAGE与Western blot进行检测;用GSTPrep FF 16/10对融合蛋白进行纯化。经双酶切鉴定证实质粒中插入基因长250bp,测序结果与大鼠SP-B cDNA序列相符;质粒转化后用IPTG进行诱导,在分子质量约34ku处出现1个新条带,与pGEX4T-1/SP-B蛋白预期大小一致,且该融合蛋白能可溶性表达并被纯化。结果表明成功构建了pGEX4T-1/SP-B重组质粒,表达、纯化得到GST/SP-B蛋白,为研究肺表面活性物质替代药物奠定了基础。     

文心兰HSP70基因的克隆及表达分析

为研究文心兰热激蛋白70基因（命名为OnHSP70）的分子特性及表达特点,以文心兰‘柠檬绿’（Oncidium hybridum ‘Honey Angel’）为材料,采用RT-PCR技术克隆OnHSP70,利用生物信息学方法分析其分子特性,通过qRT-PCR技术分析其在不同组织及不同非生物胁迫处理下的表达特点。生物信息学分析表明：该基因开放阅读框为1944 bp,编码647个氨基酸,翻译的蛋白为稳定蛋白,属于HSP70超家族,其蛋白二级结构由41.11%的α-螺旋、17.77%的延伸链、7.73%的β-转角和33.38%的无规卷曲组成,具有2个高度保守的功能域。聚类分析表明：该蛋白与铁皮石斛和深圳拟兰的HSP70亲缘关系较近。qRT-PCR分析表明：文心兰HSP70在4种不同组织器官中具有表达差异性,在花中的表达量最高;高温处理后基因的表达量明显上升;40 ℃高温胁迫4 h时表达量达峰值;茉莉酸甲酯及水杨酸处理时表达量呈现下调响应。本研究为后期该基因功能研究及提高文心兰对高温的适应性提供理论基础。     

Study on Developmental Expression of ApoCⅡ Gene mRNA in Liver Tissues of Pigs

The aim of this study was to investigate the developmental patterns of ApoCⅡ gene mRNA in liver in Mashen and Large White pigs, and study the relationship between the expression level of ApoCⅡ and the lipid metabolism in pigs.The mRNA relative expressions of ApoCⅡ gene in liver at seven stages of 1,30,60,90,120,150,and 180-day old in Mashen and Large White pigs were determined by quantitative Real-time PCR.The results showed that the developmental trend of ApoCⅡ mRNA expression in liver between Mashen and Large White pigs was different.The ApoCⅡ mRNA abundance was decreased from birth to 60-day old,then increased at 90-day old,and decreased again after that in Mashen pig.However,the relative expression amount in Large White pig was gradually decreased from birth to 150-day old and increased again at 180-day old.Except for the ApoCⅡ mRNA expression amount at 1-day old,the differences of the expression amount at other stages in Mashen and Large White pigs were significant or extremely significant (P<0.05 or P<0.01).The ApoCⅡ mRNA expression in liver was affected by age and breed,and could play an important role in lipid metabolism in pigs.     

Diversity in the carotenoid profiles and the expression of genes related to carotenoid accumulation among citrus genotypes

Carotenoids are not only important to the plants themselves but also are beneficial to human health. Since citrus fruit is a good source of carotenoids for the human diet, it is important to study carotenoid profiles and the accumulation mechanism in citrus fruit. Thus, in the present paper, we describe the diversity in the carotenoid profiles of fruit among citrus genotypes. In regard to carotenoids, such as β-cryptoxanthin, violaxanthin, lycopene, and β-citraurin, the relationship between the carotenoid profile and the expression of carotenoid-biosynthetic genes is discussed. Finally, recent results of quantitative trait locus (QTL) analyses of carotenoid contents and expression levels of carotenoid-biosynthetic genes in citrus fruit are shown.     

A review of visual soil evaluation techniques for soil structure

Soil structure forms a key component of soil quality, and its assessment by semi‐quantitative visual soil evaluation (VSE) techniques can help scientists, advisors and farmers make decisions regarding sampling and soil management. VSE techniques require inexpensive equipment and generate immediate results that correlate well with quantitative measurements of physical and biochemical properties, highlighting their potential utility. We reviewed published VSE techniques and found that soils of certain textures present problems and a lack of research into the influence of soil moisture content on VSE criteria. Generally, profile methods evaluate process interactions at specific locations within a field, exploring both intrinsic aspects and anthropogenic impacts. Spade methods focus on anthropogenic characteristics, providing rapid synopses of soil structure over wider areas. Despite a focus on structural form, some methods include criteria related to stability and resiliency. Further work is needed to improve existing methods regarding texture influences, on‐farm sampling procedures and more holistic assessments of soil structure.     

利用钾吸收基因表达评价葡萄叶面喷施钾肥效果和喷施浓度

【目的】叶面喷施钾肥可以快速、 高效地为葡萄补充钾营养,促进葡萄的高产和优质,已经被广泛应用于葡萄生产中。本实验通过对葡萄叶面喷施不同种类及浓度钾肥,测定葡萄叶片和果实等生理指标变化,以及钾吸收相关基因的表达变化,从生理和基因水平上评价这些钾肥的喷施效果,为葡萄生产中钾肥的施用提供一定指导。【方法】本实验以‘夏黑’葡萄为试材,选择两个葡萄生长关键时期盛花期和果实膨大期分别对葡萄叶片喷施0.2%、 0.5%和0.8%三种浓度的K2SO4、 K2CO3、 K2SO4·2MgSO4和KCl。然后对葡萄叶片和新梢的生长率,坐果率,叶绿素含量,单粒重,可溶性固形物等生理指标进行统计分析,并利用荧光定量PCR技术分析4个钾吸收相关基因VvHAK13、 VvKEA2、 VvSIRK和VvSORK的表达情况。【结果】叶面喷施4种钾肥后,葡萄叶片和新梢的生长率,坐果率,叶绿素含量,单粒重,可溶性固形物等各项生理指标均有不同程度的提升。钾肥种类不同,最适喷施浓度不同,同种钾肥在葡萄盛花期和果实膨大期的最适喷施浓度也有所不同,四种钾肥在果实膨大期的最适喷施浓度普遍高于盛花期。四个钾吸收相关基因在喷施不同种类及浓度钾肥后也表现出不同的表达模式,总体来讲VvKEA2、 VvSIRK 、 VvSORK的表达上调,而VvHAK13的表达下调。果实膨大期,需喷施较高浓度的钾肥,钾吸收相关基因才表现出较强烈的响应,而在盛花期则只需喷施较低浓度的钾肥。综合生理指标和基因表达两方面结果,得出4种钾肥效果依次为K2SO4·2MgSO4＞K2SO4＞K2CO3＞KCl。盛花期K2SO4和K2CO3的最适喷施浓度为0.5%,K2SO4·2MgSO4和KCl的最适喷施浓度为0.2%; 果实膨大期K2SO4、 K2SO4·2MgSO4和KCl的最适喷施浓度为0.5%,K2CO3的最适喷施浓度为0.8%。【结论】葡萄叶面喷施钾肥可以有效促进葡萄叶片和果实的生长发育,四种钾肥的效果依此为: K2SO4·2MgSO4＞K2SO4＞K2CO3＞KCl。盛花期K2SO4和K2CO3的最适喷施浓度为0.5%,K2SO4·2MgSO4和KCl的最适喷施浓度为0.2%; 果实膨大期K2SO4、 K2SO4·2MgSO4和KCl的最适喷施浓度为0.5%,K2CO3的最适喷施浓度为0.8%。适宜的喷施浓度可以有效提高钾吸收相关基因的表达,是其提高钾吸收利用的机理之一。