苹果退绿叶斑病毒组织印迹的免疫法检测与定位

用组织培养技术将富士苹果茎尖外植体建立在ＭＳ培养基中,形成试管苗。取４—６周龄的茎与愈伤组织,徒手切取约１ｍｍ厚的横切面,印迹在硝化纤维膜上。印迹组织经封闭后,与ＣＬＳＶ碱性磷酸酶标抗体反应。对反应结果进行显色。感染ＣＬＳＶ的印迹组织呈紫色反应,正常组织无显色反应。结果表明,该方法十分容易检测印迹组织中的ＣＬＳＶ。带毒愈伤组织较茎的显色反应敏感,显色反应时间仅为茎的一半（１５ｍｉｎ）。ＣＬＳＶ在愈伤组织中有两种分布情况：一是所有组织均有显色反应;二是呈不均匀分布。ＣＬＳＶ在茎组织中有三种分布情况：第一,除髓部外,表皮、皮层及维管系统中均有分布;第二,不均匀分布,ＣＬＳＶ集中分布于表度组织中,在皮层及韧皮部仅有少量分布;第三,“嵌合”分布,即同一种组织中部分组织带毒,部分为正常组织。     

Importance of Cell Wall Degrading Enzymes Produced by Fusarium graminearum during Infection of Wheat Heads

Cytological studies were carried out to elucidate the importance of cell wall degrading enzymes (CWDE) during infection of wheat spikes by Fusarium graminearum. Scanning electron micrographs revealed that at 6–24 hours after inoculation (hai) of single spikelets with macroconidia of F. graminearum, the fungus germinated by forming several germ tubes and developed a dense hyphal network in the cavity of the spikelet. At 24–36hai, the fungus formed infection hyphae which invaded the ovary and inner surface of the lemma and palea. Transmission electron microscopical studies revealed that the fungus extended inter- and intracellularly in the ovary, lemma and rachis and caused considerable damage and alterations to the host cell walls. In different tissues of healthy and F. graminearum-infected wheat spikes the cell wall components cellulose, xylan and pectin were localized by means of enzyme-gold and immuno-gold labelling techniques. Localization of cellulose, xylan and pectin showed that host cell walls which were in direct contact with the pathogen surface had reduced gold labelling compared to considerable higher labelling densities of walls distant from the pathogen–host interface or in non-colonized tissues. The reduced gold labelling densities in the infected host cell walls indicate that these polysaccharide degrading enzymes might be important pathogenicity factors of F. graminearum during infection of wheat spikes. The results revealed that, infection and colonization of wheat spikes by F. graminearum and reactions of infected host tissue were similar to those reported for F. culmorum.     

‘新红星’苹果果实蔗糖合酶的活性及亚细胞定位

 在‘新红星’苹果果实发育过程中, 伴随果糖、葡萄糖和蔗糖的积累, 蔗糖合酶的分解活性逐渐下降, 而合成活性逐渐升高。苹果果实的蔗糖合酶由分子量为90 kD 的亚基组成, 其免疫信号强度随发育进程而增加。蔗糖合酶主要定位于果肉细胞的细胞质中, 发育中后期该酶的分布密度有一定增加。综合结果认为, 蔗糖合酶调控发育早期蔗糖的降解和发育后期蔗糖的积累。     

Expression and subcellular localization of urocortin in syncytiotrophoblast of human term placenta

AIM: To obverse the expression and localization of urocortin on ultrathin cryosections of syncytiotrophoblast of human term placenta with immunocytochemistry technique under transmission electron microscope. METHODS: The human term placenta tissue from Cesarean delivery and normal labor were fixed in 4% paraformaldehyde, and then divided into two parts. One part was for regular immunocytochemistry under microscope, and the other part was used to prepare ultrathin cryosections for immunocytochemistry under transmission electron microscope. RESULTS: 1.Uroncortin mainly distributed in cytoplasm of syncytiotrophoblast of human term placenta under microscope. Urocortin also appeared in cytoplasm in some stromal cells. 2. Under transmission electron microscope, the anti-urocortin gold particles were observed in cytoplasm of syncytioptrophoblast ultrathin cryosections and sited on rough-surfaced endoplasmic reticulum. The anti-urocortin gold particles also appeared on nucleus and nuclear membrane of syncytiotrophoblast. CONCLUSION: Syncytiotrophoblast of human term placenta synthesized and secreted urocortin. The internalization of urocortin within syncytiotrophoblast nuclear indicates that urocortin may act as intracrine.     

大肠杆菌新型菌毛(F1987)的基因定位

对动物产肠毒素性大肠杆菌 (ETEC)的一种新型菌毛 (F1987)进行了相应基因的定位研究 ,通过对表达该新型菌毛 (F1987)相应 ETEC菌株的培养、质粒提取、对大肠杆菌受体菌株 DH5α的转化及阳性转化子筛选与相应菌毛表达的检定等 ,初步表明该新型菌毛 (F1987)的基因定位于质粒上     

苦瓜转录因子McEIL2的克隆及其表达分析

本研究从已构建的苦瓜果实均一化文库筛选得到一个EIN3-like同源EST序列,结合RACE技术,克隆得到EIN3基因c DNA全长序列,命名为Mc EIL2(Gen Bank登录号:KF595122)。该基因全长2 122 bp,开放阅读框1 890 bp,编码629个氨基酸,预测该蛋白的分子量为71.58 k D,与黄瓜Cus EIN3、甜瓜Cm EIL1、苜蓿Mt EIL1、绿豆Vr EIL1和番茄Le EIL1的蛋白同源性分别为89.61%、78.37%、71.23%、69.09%和65.66%。Mc EIL2基因的编码蛋白亚细胞定位于细胞核,荧光定量分析表明Mc EIL2基因表达量在幼果期先强后弱,绿熟期最低,破色期表达量大幅增加至最高随后下降,推测该基因参与苦瓜果实成熟软化调控过程。本研究初步明确了Mc EIL2基因在苦瓜成熟过程表达模式,可为今后进一步揭示苦瓜果实成熟软化的分子机制奠定基础。     

鸡Perilipin1基因的克隆及亚细胞定位

本研究旨在克隆鸡(Gallus gallus)脂滴包被蛋白基因(Perilipin1)的cDNA序列,并检测其在鸡前脂肪细胞分化过程中的亚细胞定位.本研究以7周龄肉鸡腹部脂肪组织为材料,采用RT-PCR和RACE的方法扩增并克隆了鸡Perilipin1基因的5'UTR、CDS和3'UTR片段,分析了该基因的结构;以鸡原代前脂肪细胞为材料,利用免疫荧光及激光共聚焦技术,在诱导分化的不同时间点(12～120 h),检测了鸡Perilipin1在前脂肪细胞中的表达位置.结果表明,本研究获得的鸡Perilipin11基因5'UTR、CDS和YUTR序列总长度为2 379 bp,该基因由9个外最子、8个内含子组成(ATG位于第二外显子上);在鸡原代前脂肪细胞诱导分化过程中,Perilipin1始终包被在脂滴周围.综上,本研究成功克隆了鸡Perilipinl基因的完整cDNA序列并确定了Perilipin1在鸡前脂肪细胞分化过程中与脂滴的空间位置关系,该结果为深入开展鸡Perilipinl基因的功能研究,揭示鸡脂肪代谢的分子遗传机理提供了重要的理论依据.     

F-box domain is involved in regulating the proliferation of MCF-7 cells by FBXO39 protein

AIM: To investigate the effect of F-box domain on the regulation of MCF-7 cell proliferation by FBXO39 protein. METHODS: The effect of F-box domain on the localization of FBXO39 protein in the MCF-7 cells was investigated. MCF-7 cell cDNA library was used as the template resource. The full-length cDNA sequence of FBXO39 was amplified by PCR method and subcloned into eukaryotic expression vector pEGFP-C2. The pEGFP-FBXO39ΔF (F-box domain deletion mutation) plasmid was successfully constructed with the template resource of pEGFP-FBXO39 plasmid. The recombinant plasmids were transfected into the MCF-7 cells, and then the expression of FBXO39 and FBXO39ΔF were determined by Western blot. The cellular localization of FBXO39 and FBXO39ΔF were observed by confocal microscopy. The localization of endogenous FBXO39 in the MCF-7 cells was detected by immunofluorescence staining. In addition, MTT and EdU assays were used to measure the cell proliferation, flow cytometry was used to measure the cell cycle distribution, and immunohistochemical staining was used to observe the expression of FBXO39 in the breast cancer and para-carcinoma tissues. RESULTS: The eukaryotic expression vector pEGFP-FBXO39 and pEGFP-FBXO39ΔF were constructed successfully. F-box domain had no effect on the cell localization of FBXO39. FBXO39 promoted MCF-7 cell proliferation but FBXO39ΔF did not. FBXO39 was highly expressed in the breast cancer tissues. CONCLUSION: F-box domain had no effect on the cellular localization of FBXO39 protein. However, it plays an important role in the biological function of FBXO39. FBXO39 may be related to breast cancer tumorigenesis.     

Physiological processes associated with salinity tolerance in an alfalfa half‐sib family

We previously reported an alfalfa half‐sib family, HS‐B, with improved salt tolerance, compared to parental plants, P‐B. In this study, we conducted additional experiments to address potential physiological mechanisms that may contribute to salt tolerance in HS‐B. Vegetatively propagated HS‐B and P‐B plants were treated with a nutrient solution (control) or a nutrient solution containing NaCl (EC = 12 dS/m). Shoots and roots were harvested at various time points after treatment for quantification of proline, soluble sugar, and H₂O₂ using spectrophotometers. Subcellular localization and quantification of Na in roots were conducted using a Na⁺‐specific dye under a confocal microscope. HS‐B produced 86 and 89% greater shoot and root dry biomass, respectively, compared to parental plants, P‐B, under salinity in the greenhouse. Under saline conditions the HS‐B shoots accumulated 115% and roots 55% more soluble sugars than P‐B counterparts. The non‐saline HS‐B shoots, however, showed 72% less soluble sugars than the non‐saline P‐B plants. Under saline conditions HS‐B accumulated 39% less proline in shoots, while roots accumulated 56% more proline, compared to their P‐B parents. HS‐B plants also showed a greater reduction of stomatal conductance under mild saline stress. HS‐B shoots and roots contained 3–4 times less reactive oxygen species (H₂O₂) after salt treatment compared to P‐B plants. Sodium localization and distribution analysis using Na⁺‐specific dye revealed HS‐B plants accumulated 88% and 48% less Na⁺ in stele and xylem vessels compared to P‐B. The study provides insights into the potential mechanisms that may contribute to salt tolerance in HS‐B: maintaining RWC by accumulating soluble sugars while reducing transpiration, maintaining healthy status by reducing oxidative stresses, and preventing salt toxicity by reducing accumulation of Na⁺ inside root cells and xylem.     

绵羊毛囊及其角蛋白研究进展

毛囊是毛纤维的生长基础,决定毛纤维的性状,角蛋白是毛囊组成的重要成分之一。近年来,对毛囊生长发育和角蛋白基因的挖掘等取得了一定的进展。文章回顾了绵羊角蛋白基因的发现,已经与人类角蛋白基因相当,但在数量和分类上存在差异;笔者介绍了绵羊毛囊的形态学结构,简述了毛囊角蛋白基因的表达定位和影响毛纤维性状的角蛋白,并分析了毛囊中的蛋白质组学技术的应用对毛囊生长发育和毛纤维性状的重要作用,以期为绵羊毛囊生长发育和羊毛性状的分子选育提供新的研究思路。