海口地区产脂肪酶菌株的分离与16S rRNA序列分析

以橄榄油为碳源,采用富集和复筛的方法,于夏季从海口市食堂和汽修场等富油土壤中筛选出9株高产脂肪酶活性菌株,对其形态、生长和16SrRNA序列进行分析,结果表明：各菌株菌落皆呈暗绿色,镜检为革兰氏阴性、杆状单细胞,最适生长pH值为7,最适生长温度高于42℃,9株菌株的16SrDNA序列完全一致,皆为绿脓假单胞菌。     

The effects of seasonal,sex and size on the digestive enzyme activities in freshwater crayfish (Astacus leptodactylus Eschscholtz, 1823)

The effects of seasonal, sex and size on digestive enzyme activities of Astacus leptodactylus in natural habitat were investigated in the present study. The freshwater crayfish were sampled 584 individual as seasonally from Egirdir Lake. Ten male and 10 female individuals were sampled randomly from caught crayfish in each two different size ranges (4.5–7.0, 7.1–9.9 cm) and each season. The gastrointestinal tract, gastric, intestine and midgut gland were individually dissected on ice for enzyme analysis and stomach contents. At the end of the study, amylase activities were affected by interaction of season × sex × size in intestinal, season × sex in midgut gland. However, any interaction was not determined in amylase activities of the gastric. Lipase activities were affected by interaction of season × sex × size in intestinal and gastric, season × sex in midgut gland. The protease activities in each organ were affected by interaction of season, sex and size (p ? .05). In all organs, amylase and lipase activities were similar. However, the highest protease activity was in stomach while the lowest protease activity was the intestine. The protease activities were higher than amylase and lipase activities. There is a positive correlation between protease enzyme activities in gastric with gonad maturation and active feeding period. It can be said based on data of digestive system and high protease activity that A. leptodactylus is an omnivore species in need high protein. In addition, the reason for the low levels of lipase activity may be A. leptodactylus not prefer high fat foods or not fed high fat foods.     

油用向日葵GDSL脂酶/脂肪酶基因的序列分析

为挖掘油用向日葵中的GDSL脂肪酶基因,本研究以油用向日葵转录组测序得到的Unigenes为对象,筛选GDSL脂酶/脂肪酶基因并对其理化性质和序列结构等特点进行生物信息学分析。结果表明:油用向日葵中筛选到7个GDSL脂酶/脂肪酶基因,编码的蛋白质为稳定存在的亲水性蛋白,由255~396个氨基酸组成,分子量介于28.75~42.61ku,等电点分布在4.91~8.89;除unigene 60916外,编码的蛋白质都含有较多的跨膜区域和较高的磷酸化程度;α-螺旋和无规则卷曲是主要的二级结构;进化树分析表明,这7个基因聚为三类,保守性较高,其中unigene 58624与AT4G01130.1在同一分支上高度保守,unigene 60916与AT1G74460.1相似度最高,含有多个α螺旋和β折叠,推测与种子的油脂代谢有关。     

铜绿假单胞菌脂肪酶基因(LipA)在枯草芽孢杆菌pab02中的整合表达

对铜绿假单胞菌脂肪酶基因(Lip A)进行克隆,构建整合表达质粒pHSG398–16S–lip A,并在枯草芽孢杆菌pab02中整合表达。根据铜绿假单胞菌Lip A基因设计引物,扩增Lip A基因,并进行测序,构建整合表达质粒pHSG398–16S–lip A,转化枯草芽孢杆菌pab02,并对转化后的阳性菌株进行诱导表达。结果显示,成功从铜绿假单胞菌A05菌株中扩增到长1 092 bp的Lip A基因,该基因与铜绿假单胞菌Lip A基因(登录号为AB290342)的同源性高达98%;该基因共编码311个氨基酸(aa),包含26个aa组成的信号肽和285个aa组成的成熟肽;成功构建了整合表达质粒pHSG398–16S–lip A,并将Lip A整合到枯草芽孢杆菌pab02染色体上,获得了重组枯草芽孢杆菌pab02–lip A;SDS–PAGE分析结果表明,脂肪酶蛋白得到了有效表达,该蛋白的相对分子质量约为37 000。     

休眠期中华鳖消化器官中主要消化酶活性分布

为研究休眠期中华鳖中主要消化器官消化酶活性,将平均体重为672.25g的4龄鳖6只,处死后取其胃、肝、肠,采用生化方法测定其胃蛋白酶、脂肪酶和淀粉酶活性。结果表明,在中华鳖的胃、肝、前肠、中肠、后肠中均有胃蛋白酶、脂肪酶、淀粉酶。肠中蛋白酶活性由中肠、前肠、后肠递减,前肠与中肠的酶活性没有显著差异(P>0.05);肠中脂肪酶活性较胃、肝中的高;淀粉酶活性,肠中最高,胃中次之,肝脏中最低,但在肠的不同部位淀粉酶的活性没有显著差异(P>0.05)。     

Expression of plasminogen activator-related genes in the adipose tissue of lactating dairy sheep in the early post-weaning period

There is growing evidence that plasminogen activator inhibitor type 1 (PAI-1) is expressed in adipose tissue and its expression is implicated in inflammation that accompanies obesity-associated diseases. The physiological role of other genes implicated in the plasminogen-activating cascade such as urokinase-type plasminogen activator (u-PA), u-PA receptor (u-PAR) and plasminogen activator inhibitor type 2 (PAI-2) in ovine adipose tissue remains unknown. The objective of this study was to examine the changes in the expression of four plasminogen activator (PA)-related genes during the early post-weaning period in dairy ewes. A total of 21 subcutaneous adipose tissue samples were obtained from seven lactating dairy ewes of the Chios breed at weeks 1, 2 and 4 after weaning. Results indicated that expression of all PA-related genes was detected in most of the samples examined. Greatest expression of u-PAR corresponded to highest (week 1), while greatest expression of PAI-2 corresponded to lowest (week 4) rate of lipolysis, as indicated by the expression of hormone-sensitive lipase, in the ovine adipose tissue. There were no significant differences in the expression of the other two PA-related genes (u-PA, PAI-1) throughout the experimental period. Plasminogen activator-related genes are not expressed in a coordinated manner in the adipose tissue of lactating dairy sheep in the early post-weaning period. In conclusion, adipose tissue mobilization is correlated with highest expression of u-PAR and lowest expression of PAI-2.     

脂肪酶及其在饲料中的应用

本文对脂肪酶的来源、性质、生产、检测及其在饲料中的应用研究作一综述。     

p E G F P - N 3 - L P L重组真核表达载体的构建及在V e r o细胞中的表达

目的:构建贵州白香猪LPL基因与增强型绿色荧光蛋白基因的融合表达载体pEGFP-N3-LPL,转染Vero细胞,观察重组质粒的表达.方法:采用HindⅢ和BamHⅠ两种限制性内切酶获得LPL基因编码区片段和pEGFP-N3线型载体,将目的片段与该载体连接、转化、挑取阳性克隆,进行菌落PCR、双酶切及测序鉴定.使用Lipofectamine2000将重组质粒转染Vero细胞,观察细胞中绿色荧光蛋白的表达并对其进行mRNA检测.结果:成功构建重组真核表达载体pEGFP-N3-LPL,经mRNA检测,实验组在1 500bp处出现特异性条带,说明重组载体已在Vero细胞中获得表达.结论:本试验构建的真核细胞表达载体pEGFP-N3-LPL,为进一步阐明LPL基因与肌内脂肪沉积间的生物学作用机制奠定了基础.     

脂肪酶及其产生菌的筛选与改良研究进展

脂肪酶(lipase,E.C.3.1.1.3,甘油酯水解酶)是一类特殊的酯键水解酶,能在异相系统(油—水)界面上水解特殊酯(脂肪酸甘油酯)类,具有重要而广泛的工业用途。综述了脂肪酶及其产生菌的筛选与改良研究进展。