质粒

Preventing and controlling crown gall disease applied the early biological controls strain K84, itself did not have the pathogenicity, its plasmid may synthesize one kind of bacteriocin Agrocin84, may suppress many pathogenic strains, but did not have pathogenicity to non-pathogenic strain .

防治根癌病应用较早的生防菌株是K84，本身无致病性，它的质粒可以合成一种细菌素Agrocin84，可以抑制很多有致病性的菌株，而对非致病性的菌株则无影响。

Recombinant ptotein made up 39.2%(pET-20b-bglA) and 33.5%(pET-28a-bglA) of the total proteins in the intracellular fraction, the solubility proportion of the enzyme up to 32.8%(pET-20b-bglA) and 40.1%(pET-28a-bglA), the activities of the enzyme were 66.8 (pET-20b-bglA) and 71.2 U/mg (pET-28a-bglA). These results showed that E. coli BL21-CodonPlus (DE3)-RIL with argU tRNA, ileY tRNA and leuW tRNA genes helped to improve expression of the enzyme through enhanced identification of the rare codons AGA/AGG, AUA and CUA. Further optimized the conditions for inducing, the solubility proportion of the enzyme was 70.6% at 16 °C and 1.2 times higher than 37 °C. The solubility proportion of the enzyme increased from 12.3% to 32.8% when IPTG concentrations dropped from 1000 μM to 25 μM. The recombinant enzyme was purified by heat treatment, DEAE Sepharose anion-exchange chromatography and TOYOPEARL HW-55F.

maritima MSB8 的-葡萄糖苷酶基因 bglA克隆至表达载体 pET-20b 和 pET-28a，构建重组质粒 pET-20b-bglA 和 pET-28a-bglA，然后转化不同大肠杆菌 E.coli 宿主，Tm-SIGlA 在 E.coli BL21-CodonPlus(DE3)-RIL 中获得高效表达，重组蛋白的表达量为 33.5%(pET-28a-bglA)和 39.2%(pET-20b-bglA)，可溶性比例为 32.8%(pET-20b-bglA)和 40.1%(pET-28a-bglA)，比酶活达 66.8 (pET-20b-bglA)和 71.2 U/mg (pET-28a-bglA)，这些结果表明，E.coli BL21-CodonPlus(DE3)-RIL 宿主带有的 argU tRNA、ileY tRNA 和 leuW tRNA 基因，分别增强对稀有密码子 AGA/AGG、AUA 和 CUA 的识别，有助于提高该酶在 E.coli 中的表达；进一步优化诱导条件，重组 E.coli BL21-CodonPlus(DE3)-RIL/pET-20b-bglA 在 37 ℃下诱导培养，IPTG 浓度由 1000 μM 降至 25 μM，目的蛋白可溶性表达由 12.3%增至 32.8%，提高 1.7 倍，16 ℃下低温诱导实现目的蛋白 70.6%的可溶性表达，较 37 ℃下诱导培养提高 1.2 倍。

The coat protein gene of Cucumber mosaic virusfrom melon isolate (CH99/90) was amplified by RT-PCR from the total RNA of infected zucchini leaves and cloned into pUCm-T vector.

把该分离物接种西葫芦，从发病的叶片中提取总 RNA，并以此为模板经 RTPCR 扩增获得 CMV 的外壳蛋白基因，将其克隆到 pUCmT 质粒上。

In this research we obtained the banana ACO gene with about 1.2kb length by cuffing from the cloning vector pUCm-ACO with restriction endonucleases Xba Ⅰ and Sac Ⅰ, and then directionally linked the gene to the plasmid vector pBⅠ-121 in the same endonucleases digestion sites, finally we got the construct of plant expression vector pBI-aACO.

用限制性内切酶Xba Ⅰ和sac Ⅰ从克隆载体pUCm-ACO上切下约1.2kb的香蕉ACO基因，将其定向连接在经相同酶切的质粒载体pBⅠ-121上，构建成植物表达载体pBⅠ-aACO。

The expression plasmid pGEX-5X-1/DFF45 was constructed and transformed into E.coli BL21, and the expression of GST-DFF45 protein was induced by IPTG. After purification, the fusion protein GST-DFF45 was used to immunize Oryctolagus cuniculus to obtain the antibody serum.

构建pGEX-5X-1/DFF45原核表达质粒，转化大肠杆菌BL21，用IPTG诱导融合蛋白表达，经纯化后免疫日本大耳白兔得到多克隆抗体并用CNBr-activated Sepharose？

By using Roche kit as standard, the amplification efficiency of different reagents were compared by Roche Light Cycler instrument for establishing real-time PCR reaction system. The primers were designed based on signature sequence of chromosomal DNA fragment and caf 1 gene in plasmid pMT1 of Y.pestis. And sensitivity and specificity assay were done with serial diluted Y. pestis EV76 strain and other bacterias. Then 275 Y.

采用SYBRGreenI随机掺入法，以Roche试剂盒为标准，比较两公司的DNA聚合酶，用鼠疫菌310 0 4建立荧光PCR反应体系；针对鼠疫耶尔森氏菌特异的染色体标识序列和质粒上F1抗原基因设计引物，检测其灵敏度和特异性，以盲测试验进行验证；在此基础上鉴定2 75株鼠疫耶尔森氏菌，并测试该法检测脏器污染模拟标本的灵敏度。

Immunofluorescence cytochemistry and Western blot showed that angiostatin protein was expressed and secreted by GBC-SD cells in the experimental group.

1。实验得到的含血管抑素基因的真核表达质粒pcDNA3.1-angiostatin经酶切鉴定和测序证实：碱基序列和阅读框架正确。2。

Src l was an acidic cytolysin, which was reported forthe first time. The cDNA encoding the mature Src l was cloned into non-fusion expression vectorpBV220 and expressed successfully in E.coli DH5α in inclusion body. After washing anddenaturing-renaturing, the recombinant Src l protein was purified by Q Sepharose FastFlow ion exchange chromatograph and Phenyl Sepharose hydrophobic interactionchromatograph.

首先构建了Src I基因的非融合蛋白表达质粒pBV220-Src I，并成功地在原核细胞E.coli进行了重组表达，筛选出稳定表达的工程菌株DH5α，摸索了稳定表达的条件，摸索了包涵体的洗涤、变性和复性条件，摸索出了复性蛋白Src I的纯化工艺，纯化过程简单，经QSepharose Fast Flow阴离子交换层析和Phenyl Sepharose疏水层析两步纯化，就可获得了高纯度的重组蛋白样品(纯度达99％以上)，该纯化方法适合重组蛋白的大量纯化，为Src I的性质、结构和功能研究奠定了良好的基础。

RESULTS: We cloned cytosine deaminase gene and the results from gene sequencing were the same as Genbank.

将胞嘧啶脱氨酶基因亚克隆到pIRES2-AcGFP1质粒上，构建了pIRES2-AcGFP1-CD真核表达载体。

PartⅡTransform the yeast, screen the excellent expression yeast strain, induce the yeast to express the fusion protein and depurate the target protein. The recombination plasmid was transformed into the yeast GS115 successfully by means of electrotransformation. The excellent yeast strain was got by G418 screening. We achived the yeast supernatant including target protein induced by methanol.

第二部分：酵母的转化、蛋白高表达菌株的筛选、重组蛋白的诱导表达及纯化实验利用电转化的方法，成功地将重组质粒pPIC9k-PTD-hA20导入毕赤酵母GS115，并通过G418浓度梯度筛选得到了蛋白高表达菌株，并对蛋白高表达菌株进行甲醇诱导表达，得到了含有目的蛋白的酵母上清液。

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！