人超氧化物歧化酶（SOD）基因的原核表达与蛋白纯化

从人直肠癌细胞株Colo320中提取总RNA,经RT-PCR扩增出SOD基因片段,经限制性内切酶（BamHI,Sinai）酶切后按正确的读码框顺序插入到pGEX-4T-2表达载体上,重组质粒转化大肠杆菌,经菌落PCR和质粒双酶切鉴定、序列测定确认,证实成功地构建了人SOD基因融合表达载体．转化菌经IPTG诱导表达,SDS-PAGE显示有与预期大小约44kD相吻合的融合蛋白带,获得了可溶性表达的目的蛋白,采用Glutathione Sepha—rose4B亲和层析对重组蛋白进行纯化,获得了纯度很高的目的蛋白．     

小鼠nulp1基因多克隆抗血清的制备及检测

为了在小鼠模型中更深入地研究nulp1蛋白在心脏发育中的功能,采用PCR技术扩增出小鼠nulp1基因的部分编码区并连接入pET-28a表达载体中,之后将重组质粒转入大肠杆菌(E.coli)通过IPTG诱导表达Hisnulp1融合蛋白,对该融合蛋白采用Ni-IDA凝胶柱层析纯化后,免疫新西兰兔制备了多克隆抗体,并用western blotting对抗体进行分析.结果表明,获得了高效价的特异性兔抗nulp1多克隆抗体.这为nulp1功能的进一步研究奠定了基础.     

对虾白斑综合症病毒囊膜蛋白Vp28基因的克隆、表达及蛋白纯化

以对虾白斑病毒DNA为模板扩增出Vp28基因,经限制性内切酶（SmaI,Not I）酶切后按正确的读码框顺序插入到pGEX-4T-2表达载体上,重组质粒转化大肠杆菌,经菌落PCR和质粒双酶切鉴定、序列测定确认,证实成功地构建了Vp28基因原核表达载体．转化菌经IPTG诱导表达,SDS-PAGE显示有与预期大小约52kD相吻合的融合蛋白带,18℃诱导24h后获得了可溶性表达的目的蛋白,采用Glutathione Sepharose 4B亲和层析对重组蛋白进行纯化,获得了纯度很高的目的蛋白．     

对虾白斑病毒核糖核苷酸还原酶基因原核表达载体的构建及蛋白表达、纯化

采用PCR方法扩增对虾白斑病毒核糖核苷酸还原酶基因,插入到pGEX-4T-2表达载体上构建出带有目的基因的重组质粒pGEX-4T-2RR,然后将重组质粒转化大肠杆菌.转化菌经IPTG诱导后大量表达重组蛋白,通过降低诱导温度获得可溶性表达的重组蛋白,采用Glutathione Sepharose 4B亲和层析对重组蛋白进行纯化,获得了高纯度的目的蛋白.     

水生植物凤眼莲Ca2＋-ATPase的原核表达与蛋白纯化

从水生植物凤眼莲叶片中提取总 RNA,经 RT-PCR扩增出Ca2＋-ATPase基因片段,经限制性内切酶(Sma I,Not I)酶切后按正确的读码框顺序插入到pGEX-4T-2表达载体上,重组质粒转化大肠杆菌,经菌落PCR和质粒双酶切鉴定、序列测定确认,证实成功地构建了Ca2＋-ATPase基因融合表达载体,．转化菌经IPTG诱导表达,获得了大小约48 kD的可溶性目的蛋白,与预期相吻合．利用谷胱甘肽琼脂糖凝胶4B(Gluta-thione Sepharose 4B)亲和介质对重组蛋白进行纯化,获得了高纯度的目的蛋白．     

绿僵菌Argonaute基因片段原核表达载体的构建及其表达

绿僵菌是一类广泛应用于生物防治的昆虫病原真菌.研究表明小RNA能够调控基因的表达,其中Argonaute基因在整个小RNA通路中发挥重要的作用.本研究通过RT-PCR的方法从绿僵菌中获得了Argonaute基因的部分功能片段,构建了其重组原核表达载体,将重组载体转化至大肠杆菌进行诱导表达;采用镍柱亲和纯化重组的目的蛋白并通过Western blot技术鉴定.结果发现：通过RT-PCR的方法获得长度约为950bp的基因片段;重组原核表达载体经诱导表达后,SDS-PAGE检测发现分子量约为34kDa的目的蛋白条带;诱导5h后蛋白的表达量最高,采用镍柱亲和层析纯化重组蛋白,经Western blot技术检测,重组蛋白可与His-tag抗体发生特异性反应.该纯化重组蛋白的获得为将来绿僵菌Argonaute蛋白抗体的制备,并进一步通过该抗体获得绿僵菌体内的小RNA及其靶基因提供了基础.     

胭脂鱼弹状病毒糖蛋白基因在原核细胞中的表达

克隆、构建弹状胭脂鱼弹状病毒糖蛋白基因重组质粒,并进行表达和鉴定.用RT-PCR方法扩增出大小为1.5kb的CSRV-G基因片断,构建重组克隆质粒pRSET-C/CSRV-G,并将其转化进入大肠杆菌DH5α原核表达系统,酶切鉴定证明重组质粒的正确性.经IPTG诱导后表达重组蛋白,利用SDS-PAGE对表达产物进行分析鉴定,诱导后能够表达预期的分子量为55 kD的融合蛋白.     

人C6orf210基因原核表达载体的构建及蛋白的初步表达

为了构建人C6orf210基因原核表达载体,并在E．coliBL21中表达并纯化。采用RT—PCR扩增人C6orf210基因片段,并将其克隆到原核表达载体Pet21a中,构建重组质粒pET21a—C6ort210。经限制性内切酶EcoRI与XhoI双酶切鉴定及序列测定后,转化E．coliBL21,经IPTG诱导表达融合蛋白。结果显示获得全长为1188bp的人的CAiorf210基因片段。以构建的重组质粒pET21a—C6orf210转化E．coliBL21后,经IPTG诱导,表达出相对分子质量（Mr）约66000的融合蛋白。经SINS—PAGE、Western blot分析显示,诱导表达的蛋白为C6orf210。     

假单胞茵产弹性蛋白酶基因的克隆与表达

从假单胞菌（Pseudomonassp．）XZG36中克隆弹性蛋白酶基因,构建原核表达载体,实现其在大肠杆菌（Escherichiacoli）中的高效表达,并对表达产物进行酶学性质分析,为微生物发酵生产弹性蛋白酶奠定基础．以假单胞菌基因组DNA为模板,PCR扩增弹性蛋白酶基因,并将其开放阅读框（0RF）克隆至融合表达载体pET30a（＋）进一步IPTG诱导表达;表达产物经His·Bind亲和层析纯化后对弹性蛋白酶进行酶学性质分析．实验成功克隆了弹性蛋白酶基因,DNA基因片段为1672bp、编码497个氨基酸残基的多肽,与预计长度相符合;实现了其在E．coli中的高效表达,表达量约占菌体总蛋白的20％;经SDS-PAGE分析,相对分子质量为48000,与预期的一致;提纯后的表达蛋白SDS-PAGE分析可见单一条带,纯度可达92％以上．表达蛋白具有良好活性．     

昆虫神经毒素AaIT原核表达、纯化及抗体制备

人工合成Aa IT的成熟基因,连接到p ET32载体并转化大肠杆菌,IPTG诱导表达,纯化重组Aa IT蛋白并免疫小鼠,经蛋白质-G柱对抗血清进行纯化.研究结果表明,纯化得到了Aa IT蛋白经免疫小鼠和纯化抗血清得到了特异性强的Aa IT抗体蛋白.     

卡思卡特城堡

It is worthy of noting that, whilst Crookston Castle witnessed the earlier and happier portion of Mary's variegated life,     

不同的吃喝玩乐

一、吃和喝吃苹果 eat an apple, 吃药 take medicine,吃糖 have some sweets,吃饭 have one's meals,吃馆子 dine out,吃惊 be surprised/     

漏洞百出的教育

Many college freshmen arrive woefully unprepared to do college work, and as disadvantaged populations continue to grow, the share of the American work force that has made it through college is expected to plummet. Many experts blame that educational failure not just on high schools but also on colleges. School & College, a special report by The Chronicle, looks at efforts to fix the system. What reforms would better prepare students for college? What should schools and colleges be doing differently? How should state and federal officials help?     

只要努力,不要后悔

There are numbers of crossroads on our long and unpredictable life journey where we totally have no idea about which direction to choose. No matter what our decision is, we should not turn back, but face the music and go ahead instead. I am this kind of girl who always does try without regretting, one example is how I dealt with my love.     

浅谈移民问题

Migration occurs behind a variety of reasons and has a great effect on the whole world. People may migrate in order to improve their economic situation, or in order to escape civil strife, persecution, and environmental disasters. The impact of migration is complex, bringing both benefits anddisadvantages. This paper briefly talks about the causes of migration, the allocation of benefits, and the ways in which individual countries and the international community deal with this important subject.     

Sister Carrie, an Adherent of Desires

Sister Carrie is one of the most controversial characters in American literature.Thought as a "fallen woman" firstly,she was defined as a "new woman" by some critics later. However, by digging into the motivaton behind the whole process of Carrie's "success", the relationship between Carrie and her creator (the author), the social conditions of then American, it can be found that Carrie has never been free-standing on her thought and she has never found her real-sdf even after becoming a famous actress. In a society dominated by mass consumerism Carrie is only an adherent of her own desires. She also is a representative of all those country girls flooded into cities, a symbol and a sacrifice of the urbanization of America in a time countryside was overcome by cities.     

Improvement of one-cycle controller by use of proportional integral differential controller

1.IntroductionOne-cyclecontrolmethod,whichwasproposedaboutonedecadeago[1],hasbecomeanattractivemethodinspecialfieldssuchaspowerfactorcorrection[2-6],switchingamplifiers[7,8],etc.Themainideaofthiscontrollerisbasedonintegrationofdiodevoltageinone-cycleandforcesittobeexactlyequaltothereferencevalue.Themainadvantageofthiscontrollerisitsrealtimeabilitytorejectthevariationofinputvoltage[1].Despitethisgreatability,ithasnogoodperformancesinrejectingofloaddisturbanceandfollowingreferencecommands.Espec…     

库克斯顿城堡

城堡, 是欧洲典型的建筑之一, 每个城堡都有一个美丽的传说或是一段历史。从本期开始, 我们将推出“城堡之旅”, 带各位读者去浏览、观赏欧洲各地的城堡, 开始一段文化与历史的旅途!     

The Reader Response Approach to Teaching of English Literature

During interview with English majors, it was found that a majority of students expressed disappointment with their experience of English literature classes. Students' dislike of learning English literature appears to be the major problem of teaching English literature in China. Reader response approach is a good way to solve this problem. In this paper, I explain the rationale behind the reader response approach, such as its origin, definition, features, assumptions, and strategies. Then I illustrate how reader response approach works as a teaching strategy by presenting several teaching models. At last I evaluate its usefuiness for teachers and present the advantages of reader response approach in Chinese context.