青稞HbSnRK2.4的克隆及其序列特征与表达特性分析

干旱胁迫是制约农作物生产的重要限制因素之一,研究并增强作物的抗旱性具有重要意义。Sn RK2(Sucrose nonfermenting 1-related protein kinase 2)基因编码一类蔗糖非酵解型蛋白激酶,该酶在ABA信号转录途径和抗渗透胁迫中起着重要作用。以青稞(Hordeum vulgare subsp.vulgare)抗旱品种喜马拉雅10号为材料,利用RT-PCR技术克隆获得了Sn RK2基因全长c DNA序列,命名为Hb Sn RK2.4(登录号:KJ699389)。生物信息学分析表明,该基因全长1 310 bp,编码362个氨基酸序列,蛋白分子量为41.94 k D,等电点(p I)为5.96。Prosite Scan分析结果表明,Hb Sn RK2.4含有多个干旱胁迫响应蛋白的作用位点,如酪蛋白激酶Ⅱ磷酸化位点、酪氨酸激酶磷酸化位点、蛋白激酶C磷酸化位点及N-豆蔻酰化位点等。利用实时定量PCR方法研究了Hb Sn RK2.4在干旱胁迫条件下及复水后不同时间点的表达情况,发现Hb Sn RK2.4在土壤绝对含水量为33.4%时表达量最高,随着土壤绝对含水量的下降而下调表达;当达到作物正常生长所需的土壤绝对含水量时又开始上调表达;进行干旱胁迫后(15.5%)基因表达量下降;复水后8 h时恢复正常表达水平。     

麻疹减毒活疫苗检定用Vero细胞代次对病毒滴度稳定性试验的影响

目的了解麻疹减毒活疫苗检定用Vero细胞代次的稳定性范围。方法用DMEM培养液将Vero细胞137代连续传至173代,检测其细胞活力,并观察Vero细胞的形态,接种麻疹减毒活疫苗参考品进行病毒滴定及热稳定性试验。结果 170代以下Vero细胞形态良好,细胞活力在85%以上,麻疹减毒活疫苗病毒滴度热稳定性试验的结果:137~169代Vero细胞活力及原麻疹减毒活疫苗病毒滴度热稳定性试验无统计学差异(P>0.05),170代以上代次的Vero细胞活力及原麻疹减毒活疫苗热稳定性试验有明显统计学差异(P<0.01)。结论 169代以下Vero细胞为检定用麻疹减毒活疫苗最佳细胞代次。     

Cloning and Functional Characterization of the Maize (Zea mays L.) Carotenoid Epsilon Hydroxylase Gene

The assignment of functions to genes in the carotenoid biosynthesis pathway is necessary to understand how the pathway is regulated and to obtain the basic information required for metabolic engineering. Few carotenoid ε-hydroxylases have been functionally characterized in plants although this would provide insight into the hydroxylation steps in the pathway. We therefore isolated mRNA from the endosperm of maize (Zea mays L., inbred line B73) and cloned a full-length cDNA encoding CYP97C19, a putative heme-containing carotenoid ε hydroxylase and member of the cytochrome P450 family. The corresponding CYP97C19 genomic locus on chromosome 1 was found to comprise a single-copy gene with nine introns. We expressed CYP97C19 cDNA under the control of the constitutive CaMV 35S promoter in the Arabidopsis thaliana lut1 knockout mutant, which lacks a functional CYP97C1 (LUT1) gene. The analysis of carotenoid levels and composition showed that lutein accumulated to high levels in the rosette leaves of the transgenic lines but not in the untransformed lut1 mutants. These results allowed the unambiguous functional annotation of maize CYP97C19 as an enzyme with strong zeinoxanthin ε-ring hydroxylation activity.     

一株产琼胶酶细菌的分离、鉴定及其琼胶酶基本性质

【目的】分离海洋来源的琼胶酶产生菌,对其进行分类鉴定,并研究其所产琼胶酶的基本酶学性质,为琼胶酶的应用研究及开发利用奠定基础。【方法】通过以琼脂为唯一碳源的选择培养基分离产琼胶酶的菌株;利用16S rRNA基因序列分析、表型和生理生化特征对菌株进行鉴定;通过DNS-还原糖法测定琼胶酶活性;利用显色底物法测定琼胶酶的类型;对菌株所产琼胶酶粗酶的酶学性质进行初步研究。【结果】分离到一株产琼胶酶的菌株NTa,16S rRNA基因序列分析显示该菌株属于寡养单胞菌属(Stenotrophomonas sp.);该菌株主要产胞外琼胶酶,可分泌α-琼胶酶和β-琼胶酶;琼胶酶粗酶的最适反应温度和pH分别为40℃和7.0,并且琼胶酶在温度低于30℃,pH为7.0-9.0时稳定;Ca2+对琼胶酶粗酶具有促进作用,Ag+、Fe2+、Ba2+、Mn2+、Cu2+、Zn2+和Fe3+均可不同程度地抑制酶的活性;EDTA对琼胶酶粗酶活性具有抑制作用;琼胶酶粗酶对检测的抑制剂、去垢剂及变性剂有较好的抗性。【结论】海洋细菌Stenotrophomonas sp.NTa是一种新型的产琼胶酶菌株,可同时分泌α-琼胶酶和β-琼胶酶,具有潜在开发利用价值。     

Hsp90 inhibitor 17-AAG sensitizes Bcl-2 inhibitor (-)-gossypol by suppressing ERK-mediated protective autophagy and Mcl-1 accumulation in hepatocellular carcinoma cells

Natural BH3-memitic (-)-gossypol shows promising antitumor efficacy in several kinds of cancer. However, our previous studies have demonstrated that protective autophagy decreases the drug sensitivities of Bcl-2 inhibitors in hepatocellular carcinoma (HCC) cells. In the present study, we are the first to report that Hsp90 inhibitor 17-AAG enhanced (-)-gossypol-induced apoptosis via suppressing (-)-gossypol-triggered protective autophagy and Mcl-1 accumulation. The suppression effect of 17-AAG on autophagy was mediated by inhibiting ERK-mediated Bcl-2 phosphorylation while was not related to Beclin1 or LC3 protein instability. Meanwhile, 17-AAG downregulated (-)-gossypol-triggered Mcl-1 accumulation by suppressing Mcl-1^Thr163 phosphorylation and promoting protein degradation. Collectively, our study indicates that Hsp90 plays an important role in tumor maintenance and inhibition of Hsp90 may become a new strategy for sensitizing Bcl-2-targeted chemotherapies in HCC cells.     

Substrate consumption and excess sludge reduction of activated sludge in the presence of uncouplers: a modeling approach

A mathematical model with a consideration of energy spilling is developed to describe the activated sludge in the presence of different levels of metabolic uncouplers. The consumption of substrate and oxygen via energy spilling process is modeled with a Monod term, which is dependent on substrate and inhibitor. The sensitivity of the developed model is analyzed. Three parameters, maximum specific growth rate (μ _max), energy spilling coefficient (q _max), and sludge yield coefficient (Y _H) are estimated with experimental data of different studies. The values of μ _max, q _max, and Y _H are found to be 6.72 day^-1, 5.52 day^-1, and 0.60 mg COD mg^-1 COD for 2, 4-dinitrophenol and 7.20 day^-1, 1.58 day^-1, and 0.62 mg COD mg^-1 COD for 2, 4-dichlorophenol. Substrate degradation and sludge yield could be predicted with this model. The activated sludge process in the presence of uncouplers that is described more reasonably by the new model with a consideration of energy spilling. The effects of uncouplers on substrate consumption inhibition and excess sludge reduction in activated sludge are quantified with this model.     

再引入黑颈长尾雉夏季栖息地选择

2008年6～7月,在广西岑王老山国家级自然保护区采用样带法对再引入黑颈长尾雉(Syrmaticus humiae)的栖息地选择进行了研究。共测定了7条样带上的22个栖息地样方和17个对照样方的24个生态因子。结果表明,再引入黑颈长尾雉的夏季栖息地偏向于选择海拔低、坡位低、乔木较低和灌木较高、落叶盖度较大、灌木层盖度大、草本高且盖度大的阔叶林、针阔混交林或者针叶林;逐步判别分析的结果显示坡位、草本层盖度是重要的生态因子,由这两个变量构成的方程在对栖息地和对照区进行区分时,正判率可以达到77.3%,这种选择与黑颈长尾雉的隐蔽条件和食物密切相关。     

大额牛产犊行为观察

采用直接观察法对大额牛(Bos frontalis)的产前离群与产后带犊归群、分娩等产犊行为进行了观察。结果表明,(1)野外观察17头临产母牛中,76.47%(13头)的个体在产前(11.23±7.14)d离开社群独行觅食,寻找分娩场所;分娩地点多选择在靠近溪河、地势较高、食物丰富、光照好、地面相对平坦的草地、林边或林中。母牛野外产犊后带犊归群的时间为产后(5.57±2.64)d。(2)在设围栏的人工草地上放牧条件下对6头母牛分娩行为观察表明,产前24～12 h母牛的警觉巡视行为明显增多;产前12～5h表现为烦燥不安和频繁爬跨;产前3～1 h领地行为明显,对分娩地点4～6 m2内长势较高的牧草等进行清理,为新生犊牛行动清除障碍;分娩时间为6:00～18:00时,其中在6:00～7:00时分娩的占66.67%(4/6);母牛从卧地进入分娩至犊牛产出期间起卧(7.00±6.00)次,从卧地到犊牛完全产出需(25.00±5.00)min,胎衣排出的时间为产后(5.64±1.80)h;犊牛(n=6,♂1+♀5)初生体重为(16.46±2.56)kg,母牛无难产现象;母牛对犊牛能及时舔饰,定时授乳和用心守护。犊牛从产出至第一次站稳需(52.70±29.69)min,第一次站稳到吮吸初乳需(38.7±29.14)min,第一次吃初乳到向周边活动需(61.33±1.53)min。