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Abortus, amplify BP26 gene by PCR and clone to pMD18-T simple vector. Identify the constructed recombinant plasmid pMDBP26 by sequencing, then subclone to vector pET-28a. Transform the constructed recombinant plasmid pETBP26 to E. coli BL21(DE3) for expression under induction of IPTG. Purify the expressed product by histidine-binding resin column chromatography and identify by Western blot.

提取布鲁氏菌基因组DNA，用PCR扩增出BP26基因，并克隆到pMD18-T载体上，测序正确后，再亚克隆至pET-28a载体上，转化到大肠杆菌BL21(DE3)中，用IPTG诱导表达，经组氨酸结合树脂柱纯化，并对纯化后的表达产物进行Western blot鉴定。

METHODS摘要: Molecular cloning techniques were used to construct the recombinant plasmid pLMP1p53mt containing mutant p53 gene and EB virus LMP1 gene. Linear recombinant plasmid pLMP1p53mt was transfected into mouse androgenesis of germ cells by microinjection and then survival germ cells treated were planted into fallopian tubes of artificial pregnant female mice.

方法摘要：通过分子克隆技术构建含突变型p53基因和EB病毒LMP1基因的真核表达载体pLMP1p53mt；采用显微注射法将线性化的表达载体注射至小鼠受精卵的雄性原核中，然后将注射存活的受精卵植入假孕母鼠的输卵管；取其产3 wk龄子代鼠进行PCR初选，再通过Southern杂交进一步确证；利用HE染色法观察4 mo龄转基因小鼠不同组织病理变化。

The constructed pcDNA3-IL-15 plasmid was identified by Bgl Ⅱ digestion,PCR amplification and sequenceanalysis respectively,to ensure that the hIL-15 cDNA was properly inserted into thevector and the sequence was correct.Then the pcDNA3-IL-15 plasmid was instantlyexpressed in COS-7 cells by means of lipofectamine transfection,and followed by astable expression in Chinese hamster oval cells.8 positive cell colonies wereobtained after the G418 selection,designated CHOA~H cells.

经过BglⅡ酶解分析，PCR扩增鉴定和DNA测序分析，证明IL-15 cDNA已正确插入克隆载体且序列正确后，采用脂质体法，首先将pcDNA3-IL-15重组质粒转染至非洲绿猴肾细胞（COS-7）做瞬时表达，再转染至中国仓鼠卵巢细胞进行稳定表达。

The fragment was connected to pcDNA3.0 carrier which was cut by the same two enzymes to form the plasmid which was transformed into Escherichia coli DH5a.Appropriate clone was cultured to extract plasmid.The extract was analysed to confirm that it was constructed correctively.

将该片段连接到同样被KpnⅠ和XhoⅠ双酶切的pcDNA3.0载体中，转化大肠杆菌DH5α感受态细菌，挑取阳性克隆进行质粒提取，进行酶切鉴定，并获得阳性克隆。

The construction process includes the following steps: constructing recombinant plasmid containing phhA and phhB; transforming the recombinant plasmid to hygrophilous aeromonad; and screening recombinant hygrophilous aeromonad with the recombinant plasmid.

该重组菌中含有β-酮硫解酶基因phbA和乙酰乙酰辅酶A还原酶基因phbB。其构建方法包括以下步骤：1构建含phbA和phbB基因的重组质粒；2转化该重组质粒到嗜水气单胞菌中；3筛选出含有重组质粒的重组嗜水气单胞菌。

CaN Aβ gene silencing can reduce myocardial hypertrophy in cultured cells, si1280 (21bp) of CaN Aβ gene is the most effective target site for siRNA. The method of intrapericardial injection of plasmid, microbubbles and erzymes can improve transfection efficiency of non-viral plasmid with satisfying targeted transfection. But the scope of transfected myocytes is still limited. CaN Aβ shRNA expressing plasmid transfection in vivo by pericardial injection results in decreased CaN Aβ protein expression of small part of myocytes, and CaN Aβ mRNA only shows decreased trend. The dosage of non-viral vector and the parameters of ultrasound energy should be optimized in further study.

结论RNAi技术抑制CaN Aβ基因表达可有效减轻Ald诱导的心肌细胞肥大程度；CaN Aβ基因中si1280(21bp)片段为实现RNAi的更有效片段；微泡+酶类心包腔内注射超声导入的方法可有效改善非病毒载体在体心肌的转染效率，同时具有一定的靶向性，但总的转染范围仍然有限；采用这一方法进一步进行&一肾一夹&心肌肥大模型大鼠在体心肌细胞的CaNAβ的RNAi干预，发现心肌肥大大鼠心外膜下局部心肌细胞CaN Aβ蛋白水平降低，CaN AβmRNA水平虽有下降趋势，但无统计学差异，提示质粒的用量及超声导入的参数有待进一步研究使其优化。

Formation of mating pairs, nicking of the F oriT sequence, and transfer of the 5' end of a single strand of F DNA proceed as in transfer of the F plasmid. Therefore, Part of the plasmid is transferred first, Chromosomal genes are transferred next, and the rest of the plasmid is transferred last.

雌雄菌的交配、F质粒oriT处缺口的形成、以及由单链F DNA5'末端开始转移的过程都与F质粒的转移相同，因此，首先转移的是部分质粒片段，其次是染色体基因，剩余的质粒片段最后被转移。

METHODS The HCV core gene coding region was inserted into the eukaryotic expressive plasmid pcDNA3, then the recombinant plasmid pcDNAHCV-C was constructed and expressed transiently with LipofectAMINE in the SP2/0 cells. After purification, these plasmid directly or encapsuled wit h Lipofect-AMINE were injected into BALB/c mice. HCV core antibody from immunized mice could be detected by ELISA. RESULTS The enzyme -cutting identification showed that HCV core gene fragment had been cloned into pcDNA3 eukaryote vectors.

人工构建包含HCV C基因片段的真核表达载体pcDNAHCV-C，在证实其可以在真核细胞中表达之后，将其用脂质转染剂包裹，形成LipofectAMINE-pcDNAHCV-C脂质混合物，将该混合物或单纯pcDNAHCV-C直接注射BALB/c小鼠股四头肌，以空载体pcDNA3做对照，ELISA法检测血清中抗体产生水平。

The pHybLex/Zeo-Idl plasmid and the cDNA library plasmid were sequentially transformed into the yeast swains and screened to obtain Leu2^＋ and Leu2^＋LacZ^＋ clones, with the false positive clones excluded using positive and negative controls, and the plasmid of the true positive clone was sequenced and blasted for homological analysis.

方法PCR方法扩增Id1的编码序列并定向克隆入诱饵质粒pHybLex/Zeo，构建重组诱饵质粒pHybLex/Zeo-Id1，酶切鉴定后转化入酵母株EGY48/pSH18-34，检测重组诱饵质粒有无非特异激活报告基因Leu2、LacZ作用；扩增并提取成人肺组织文库质粒，将文库质粒及重组诱饵质粒转化入酵母细胞，依次筛选Leu2^＋，Leu2^＋LacZ^＋克隆；设置阴性及阳性对照，重复筛选Leu2＋LacZ＋克隆并排除假阳性克隆，获取真阳性文库质粒，进行测序和同源性比对。

A process for preparing the recombinant chicken gamma-interferon (rCHIFN-gamma) with high activirus activity includes such steps as cloning ChIFN-gamma gene from eukaryotic plasmid to transfer carrier by dual enzyme servenings to obtain recombinant transfer plasmid pFASTBACI-ChIFN-gamma, taking DH10Bac competent cell, adding IngpFASTBAC1-ChIFN-gamma, transposition, diluting to culture object by SOD culture medium, coating on Luria plate, culturing, choosing white clony, purifying, naming positive plasmid as Bacmid-ChIFN-gamma, extracting its DNA, and transfecting Sf9 cells.

高抗病毒活性重组鸡γ-干扰素的制取方法及用途，涉及一种广谱高效抗病毒基因工程产品的制取方法。将ChIFN-γ基因从真核质粒中经双酶切后克隆到转移载体中，获得重组转移质粒pFASTBAC1-ChIFN-γ；取DH10Bac感受态细胞，加入1ng pFASTBAC1-ChIFN-γ，转座后，用SOC培养基稀释培养物，涂布Luria平板，培养后，挑取白色菌落，Luria平板进行纯化，阳性质粒命名为Bacmid-ChIFN-γ，提取阳性质粒DNA；取上述重组DNA质粒转染Sf9细胞，制得高抗病毒活性重组鸡γ-干扰素。

She gently rebuff ed him, but agreed that they could be friends

她婉言拒绝了，但同意作为朋友相处。

If in the penal farm, you were sure to be criticized.

要是在劳改农场，你等着挨绳子吧！