PCV2衣壳蛋白羧基端基因在T4噬菌体表面的展示及ELISA方法的建立

用PCR扩增猪圆环病毒Ⅱ型广东分离株的衣壳蛋白羧基端基因,将PCR产物连接到pR质粒,转化DH5α细胞,筛选阳性克隆进行PCR鉴定并测序后,转化E2菌,将重组E2菌与缺陷型噬菌体T4-Z1同源重组后,得到重组噬菌体,经SDS-PAGE和Western blotting分析,表明衣壳蛋白片段在噬菌体表面正确展示,表达的融...     

封闭型植物工厂发展现状与展望

经过多年探索和发展,封闭型植物工厂无论在材料、装置和技术都取得了快速进步,生产产量成数倍增长。在介绍发展封闭型植物工厂必要性的基础上,着重分析了封闭型植物工厂的发展现状、设施分类、栽培形式选择、栽培技术选择,最后指出了存在的问题,并对未来的封闭型植物工厂外部设施及其栽培技术进行了展望。     

不同光质LED光源对鱼腥草幼苗膜脂过氧化及抗氧化酶的影响

以鱼腥草为材料,以日光灯处理为对照,分析了不同光质LED光源(白、红、黄、绿、蓝LED灯)对鱼腥草叶片活性氧、类黄酮含量及抗氧化酶的影响。结果表明,蓝光下叶片中TBARS(硫代巴比妥酸)含量最高,较对照高30.18%,与希夫试剂染色结果相一致;过氧化氢产生最多,较对照高190.21%,与DAB染色结果一致;超氧阴离子产生速率在蓝光下最高,为对照组的1.54倍,白光下最低;蓝、绿光下脯氨酸含量是对照的2倍;蓝光和黄光处理下,黄酮含量较对照提高了21.4%。蓝、绿光提高了SOD2同工酶的表达,且在蓝光下最高;蓝、绿、红、黄光处理分别诱导了更多的POD基因位点的表达;不同LED光质均提高了CAT抗氧化酶活性,其中蓝、绿光下CAT同工酶的表达量明显高于对照。综上所述,蓝色光处理可以有效提高幼苗的抗氧化能力,可为鱼腥草品质的控制提供参考。     

猪链球菌蛋白表面展示系统的构建及展示蛋白的初步观察

为构建猪链球菌(Streptococcus suis)蛋白表面展示系统,本研究通过序列分析,确定猪链球菌的LPxTG蛋白及其信号肽(SP)和胞壁锚定基序(CWA),通过PCR扩增Peno-SP、GFP、CWA的DNA片段并融合,构建强启动子Peno控制表达编码SP-GFP-CWA融合蛋白的DNA片段,将该重组DNA片段连接pSET2载体,获得蛋白表面展示质粒,转化猪链球菌,构建得到以GFP为报告蛋白的猪链球菌蛋白表面展示系统。结果显示,利用猪链球菌的10个LPxTG蛋白及其SP和CWA序列,构建了10个含有Peno-SP-GFP-CWA融合片段的重组pSET2表面蛋白展示质粒pSsPSD1至pSsPSD10,分别转化猪链球菌05ZYH33,PCR鉴定显示其中7个转化猪链球菌。采用western blot初步检测其展示蛋白,结果显示,7个转化阳性菌株均能有效表达GFP蛋白,以成熟GFP条带为指标,均表现出了一定的外源GFP表面展示水平,分别命名为SsPSD1、SsPSD2、SsPSD4、SsPSD7-SsPSD10,其中SsPSD1、SsPSD4、SsPSD8和SsPSD9表面展示水平相对较好,在猪链球菌表面展示外源蛋白方面具有很好的潜力。本研究首次尝试建立猪链球菌蛋白表面展示系统,为猪链球菌表面递呈外源蛋白或抗原提供了新的策略。     

Construction and Identification of Yeast Surface Display Libraries of Ovis aries DRA Gene Exon 2

The specific primers were designed according to Ovis aries DRA gene sequence deposited in GenBank and the multiple cloning site of the plasmid pYD1,which was a vector used for protein surface display on Saccharomyces cerevisiae.The gene encoding DRA was amplified by PCR using the genomic RNA of Ovis aries.The 762 bp fragment was cloned and released in GenBank and registration number was KR422362.The PCR product was inserted into the yeast surface display plasmid vector pYD1 by double enzyme digestion.It was indicated that DRA gene was successfully integrated into the genome.Dot mutation was made at both ends of exon 2 in DRA gene for making restriction enzyme cutting site and design the exon 2 specific primers according to mutated Ovis aries DRA gene sequence.Sequenced exon 2 amplification products based on DNA pooling of sheep large sample template was analyzed the polymorphic loci.The polymorphic exon 2 246 bp fragment was obtained by double enzyme digestion and connected to surface display restructuring mutation carriers pYD1-DRA by the same double enzyme digestion,and then we successfully constructed yeast surface display libraries.We transformed it into Saccharomyces cerevisiae EBY100 cell.Yeast monoclone was identified by PCR amplification and sequencing,and we confirmed that DRA gene had been integrated into Saccharomyces cerevisiae genome.After galactose induced,it was detected that DRA gene library had been successfully demonstrated on the yeast cell surface under the fluorescence microscope by immunofluorescence method.     

Utilization of digital differential display to identify differentially expressed genes related to rumen development

This study aimed to identify the genes associated with the development of the rumen epithelium by screening for candidate genes by digital differential display (DDD) in silico. Using DDD in NCBI's UniGene database, expressed sequence tag (EST)‐based gene expression profiles were analyzed in rumen, reticulum, omasum, abomasum and other tissues in cattle. One hundred and ten candidate genes with high expression in the rumen were derived from a library of all tissues. The expression levels of 11 genes in all candidate genes were analyzed in the rumen, reticulum, omasum and abomasum of nine Japanese Black male calves (5‐week‐old pre‐weaning: n = 3; 15‐week‐old weaned calves: n = 6). Among the 11 genes, only 3‐hydroxy‐3‐methylglutaryl‐CoA synthase 2 (HMGCS2), aldo‐keto reductase family 1, member C1‐like (AKR1C1), and fatty acid binding protein 3 (FABP3) showed significant changes in the levels of gene expression in the rumen between the pre‐ and post‐weaning of calves. These results indicate that DDD analysis in silico can be useful for screening candidate genes related to rumen development, and that the changes in expression levels of three genes in the rumen may have been caused by weaning, aging or both. © 2015 Japanese Society of Animal Science     

基于空间漫反射光的母兔早期妊娠诊断装置研制

妊娠诊断是肉兔繁殖管理中的重要环节,尤其是早期妊娠诊断,可提前未妊娠母兔的再授精时间,提高种兔利用率,缩短繁殖周期。针对人工摸胎诊断法存在对母兔腹内胚胎产生机械损伤、母兔应激反应大及对工人经验要求高等缺陷,该研究基于妊娠母兔与未妊娠母兔腹内有无孕囊组织所引起的光学特性差异,提出一种基于空间漫反射光的母兔妊娠诊断方法,研制了快速、无侵入式的便携诊断装置。该装置由具有2个红外发光二极管（Light Emitting Diode,LED,LED发光波长分别为850和930 nm）、3个硅基光电二极管和外围电路的传感探头和信号处理主机组成。利用该便携诊断装置采集130只人工授精14 d后的母兔（包括63只妊娠母兔和67只未妊娠母兔）腹部漫反射光强度数据,将采集的数据按照7∶3的比例划分为训练集和测试集,训练集数据分别用于建立偏最小二乘判别分析（Partial Least Squares Discrimination Analysis, PLS-DA）模型和支持向量机（Support Vector Machine, SVM）分类模型,测试集数据用于模型性能测试,并对比两种模型的分类性能。同时,利用PLS-DA对采样数据进行有监督的主成分分析和变量重要性分析,结果表明妊娠母兔与未妊娠母兔的采样数据之间存在差异,可以被较好的分类。对比两种分类模型的分类性能发现SVM对妊娠母兔和未妊娠母兔的分类性能均比PLS-DA好,对测试集数据的灵敏度、特异性和准确率分别为80.95%、83.33%和82.05%。研究结果表明,该研究提出的光学妊娠诊断方法可行,研制的诊断装置可对授精14 d后的母兔进行妊娠诊断,对提高兔产业体系智能化装备水平有积极的促进作用。     

辐照技术在果蔬贮藏保鲜中的应用研究进展

辐照保鲜技术是一种新型、绿色、便捷、高效、低能耗、去残留的保鲜技术,已广泛应用于果蔬贮藏保鲜领域,该技术可显著延长果蔬的贮藏期,延缓果蔬采后衰老,防止腐烂变质。笔者从辐照保鲜技术的基本原理、应用和存在问题三个方面进行阐述,并对其未来在果蔬贮藏领域中的应用前景进行展望。