烟草玉米黄质环氧化酶基因的转录表达模式及其功能研究

通过Real-time PCR检测了普通烟草(Nicotiana tabacum)红花大金元的玉米黄质环氧化酶(Zeaxanthin epoxidase,ZE)ZE基因在盛花期不同器官中的表达量.利用烟草脆裂病毒诱导的基因沉默(VIGS)技术抑制本氏烟草(Nicotiana benthamiana)ZE基因的表达,在该基因沉默后,Real-time PCR检测其上游基因的表达变化;同时检测烟草中质体色素(β-胡萝卜素、紫黄质、新黄质、叶黄质、叶绿素a和叶绿素b)含量的变化.结果显示:ZE基因在盛花期的第10位叶片、第15位叶片和花萼中表达量较高.与对照组相比,在ZE基因沉默后,其上游的基因八氢番茄红素合成酶(PSY)、八氢番茄红素脱氢酶(PDS)、ζ-胡萝卜素脱氢酶(ZDS)、类胡萝卜素异构酶(CRTISO)、番茄红素β-环化酶(β-LCY)、胡萝卜素β-环羟化酶(β-OHase)和紫黄质脱环氧化酶(VDE)的表达量降低;烟草中质体色素的含量降低.以上结果说明:ZE基因作为类胡萝卜素合成通路下游的基因在该通路中发挥着重要的调控作用,该基因表达量的变化可以影响烟草中质体色素的含量,与烟草的光合生理过程也存在着密切关系.     

Negatively and positively charged bacterial aerosol concentration and diversity in natural environments

Bioaerosol charge information is of vital importance for their electrostatic collection. Here, electrostatic means and molecular tools were applied to studying bioaerosol charge dynamics. Positively or negatively charged bioaerosols were collected using an electrostatic sampler operated with a field strength of 1.1 kV cm 1 at a flow rate of 3 L min 1 for 40 min. Those with fewer or no charges bypassing the sampler were also collected using a filter at the downstream of the electrostatic sampler in one environment. The experiments were independently conducted three times in three different environments. The collected bacterial aerosols were cultured directly on agar plates at 26°C, and the colony forming units (CFU) were manually counted. In addition, the CFUs were washed off from the agar plates, and further subjected to polymerase chain reaction (PCR) and denaturing gradient gel electrophoresis (DGGE) for culturable diversity analysis. The results revealed remarkable differences in positively and negatively charged culturable bacterial aerosol concentration and diversity among the studied environments. In the office environment, negatively charged culturable bacterial aerosols appeared to dominate (P = 0.0489), while in outdoor and hotel environments both polarities had similar concentration levels (P = 0.078, P = 0.88, respectively). DGGE patterns for positively charged culturable bacterial aerosols were shown strikingly different from those of negatively charged regardless of the sampling environments. In addition, for each of the environments positively charged culturable bacterial aerosols collected were found to have more band pattern similarity with those positively charged for respective regions of agar plates than those negatively charged, and vice versa. The information developed here is useful for developing efficient electrostatic sampling protocols for bioaerosols.     

视网膜静脉周围炎与HLA的初步研究

为了解HLA—DBB、-DQB1及-B各等位基因在正常人群和视网膜静脉周围炎患者中的频率分布，寻找HLA-DRB、-DQB1及-B各等位基因位与视网膜静脉周围炎的关联基因，并进一步分析该基因与视网膜静脉周围炎的关系，特应用病例一对照分析方法，采用PCB-序列特异性引物基因分型技术进行HLA-DBB、-DQB1及-B等位基因分型，本研究共检测到HLA-DBB位点上15种等位基因、HLA-DQB1位点上7种等位基因以及HLA-B位点上22种等位基因；发现在患者组中DBB1＊04基因频率高于对照组，DBB1＊04与Eales病发病呈正相关；在健康对照组HLA-B＊15基因位点出现频率明显高于患者组，HLA-B＊15与Eales病发病呈负相关。结果表明：获得HLA-DBB、-DQB1及-B等位基因在中国北方汉族正常人群和Eales病患者中的频率分布的初步特征；DBB1＊04可能是Eales病的易感基因；HLA-B＊15可能是Eales病的保护性基因。     

乳腺癌患者外周血中MnSOD基因表达检测及其临床意义的研究

目的探讨锰超氧化物歧化酶(manganese superoxide dismutase,MnSOD)在乳腺癌患者和正常人外周血中的表达差异及其临床意义.方法采用实时荧光定量PCR方法检测218例乳腺癌患者和77例正常女性外周血中MnSOD基因表达水平,并进行分析.结果乳腺癌患者外周血中MnSOD基因表达水平为4.38±0.74,正常女性外周血中MnSOD基因表达水平为4.16±0.64,P0.001,有统计学意义.淋巴转移患者组和临床早期患者组与正常女性组比较,P0.001.结论淋巴转移和临床早期乳腺癌患者外周血中MnSOD基因表达水平显著低于正常女性,提示MnSOD的低表达与肿瘤的发生和发展有关.应用荧光定量PCR方法检测外周血MnSOD基因水平对监测乳腺癌有一定意义.     

抗牙周病厌氧菌（Bg）单链抗体基因的构建与表达

从牙周病厌氧菌（Ｂａｃｔｅｒｏｉｄｅｓｇｉｎｇｉｖａｌｉｓ）单克隆抗体的杂交瘤细胞株出发，通过基因工程方法──ＰＣＲ技术，调出其抗体的重链和轻链基因可变区序列，体外重组后得到单链抗体ＳｃＦｖ（ｓｉｎｇｌｅｃｈａｉｎｆｒａｇｍｅｎｔｖａｒｉａｂｌｅ）片段，克隆到了ｐＣＡＮＴＡＢ５载体上，在ＥｓｃｈｅｒｉｃｈｉａｃｏｌｉＴＧｌ中得到了成功表达。     

小麦叶绿体DNA的提取与psbA基因的扩增

利用高离子强度，低ｐＨ值缓冲液匀浆细胞的方法，提取小麦叶绿体ＤＮＡ，并以此为模板，通过聚合酶链反应（ＰＣＲ）对小麦叶绿体ｐｓｂＡ基因进行了特异性扩增。     

应用gE-MGB-TaqmanPCR方法检测冰冻动物组织样品

伪狂犬病毒是严重影响养猪业发展的重要病毒,病毒粒子具有较强的抵抗力。gE基因是伪狂犬病毒的毒力基因和gE-基因缺失疫苗的缺失基因。本实验应用gE-MGB-TaqmanPCR检测18份冰冻组织样品,阳性率为15/16(除2份弃去),与血清中和结合细胞MTT比色实验结果比较,符合率为100%。此方法以闭管的模式操作,减少了以后步骤污染的可能性,整个PCR检测过程少于2个小时。     

SLE患者血清抗Sm抗体免疫PCR的方法学评价

目的为SLE患者血清抗Sm抗体提供免疫PCR测定方法.方法评价免疫PCR方法检测抗Sm抗体的特异性、敏感性、重复性和相关性.结果免疫PCR方法测定SLE患者血清抗Sm抗体特异性强,敏感性是ELISA方法的107倍,而且与ELISA方法呈高度正相关.结论免疫PCR方法用于血清抗Sm抗体测定具有临床实用价值.     

不同品种猪解耦联蛋白基因3′调控区的遗传变异

对猪UCP3基因3′调控区进行了克隆测序，通过序列比对分析发现长白猪、内江猪、民猪和二花脸猪存在15个碱基位点的多态，利用PCR-RFLP分析长白猪、大白猪、内江猪、民猪、二花脸猪及梅山猪存在连续的9个多态位点，经X^2检验发现二花脸猪与长白猪、大白猪、内江猪和民猪差异极显著，与梅山猪差异显著，梅山猪与长白猪、民猪差异显著，提示太湖猪具有独特的种质特性。     

转基因花卉的DNA提取与多重PCR分析

分别以CTAB法、DNA提取试剂盒提取菊花、矮牵牛、非洲菊3种花卉的基因组DNA，凝胶电泳结果表明试剂盒提取的效果较好。设计合成了3对引物，分别扩增花椰菜花叶病毒（Cauliflower mosaic virus,CaMV)35S启动子、根癌农杆菌胭脂碱合成酶基因（nos）终止子、大肠杆菌K12菌株新霉素磷酸转移酶Ⅱ(nptⅡ）基因。普通PCR检测结果表明，3种花卉6个样品中仅矮牵牛的1个样品含有这3种转基因成分，酶切鉴定结果验证了PCR产物为非假阳性。尝试以多重PCR同时检测转基因矮牵牛与对照阳性质粒中的2个或3个转基因成分，得到预期结果。