12字咒语

<正>【上集回顾】古古卡老师用宝石钥匙打开了神秘门,进入了一个奇妙的世界。在那里,他认识了鸭子管家克拉拉、小驴立耳奇以及杜立德医生。杜立德医生说,他给箭毒蛙做的再造毒素手术失败了,而古古卡就是他们的救星。这究竟是怎么一回事呢?     

白细胞介素12研究进展

白细胞介素１２（ＩＬ－１２）是近年新发现的一种细胞因子,主要来源于Ｂ淋巴细胞系。它是目前发现的细胞因子中唯一由异二聚体组成的因子,两亚基的分子量分别为４０ｋＤ和３５ｋＤ。两亚基的基因定位于不同的染色体上。ＩＬ－２具有多种生物学效应,只存在单一亲和力的膜受体。对ＩＬ－１２的研究将有助于确定其在抗肿瘤和抗感染免疫中的重要作用。     

白细胞介素12研究进展

白细胞介素１２是一种异源二聚体细胞因子。ＩＬ－１２能对Ｔ细胞和ＮＫ细胞的功能施加多种影响,促进细胞免疫,诱导干扰素－γ产生,具有抗肿瘤作用,且毒副作用很小。本介绍了ＩＬ－１２的分子结构及生物学功能。     

第12号木棒

正思维跳跳糖里"堆雪人"大赛正在火热进行中,动物选手们玩出了新花样。中场休息时,糖兄打算出一道题考考大家。他用大伙儿"堆雪人"的道具——13根木棒摆了个"迷阵",要求大家经过仔细观察,按顺序一根一根地将需要移动的木棒拿开,最终"解放"第12号木棒。试试看,你能做到吗?(注意:每根木棒被拿掉时,上面不能压着其他木棒。)     

A newborn child with karyotype 47,XX,+der(12) (12pter→12q12::8q24→8qter),t(8;12) (q24;q12) pat

Summary A case of Meckel or Gruber syndrome is reported, together with a survey of the relevant literature of recent years (1971–1977), in reference to a probably autosomal recessive inheritance of this malformation.     

白细胞介素12的研究进展

白细胞介素１２（ＩＬ－１２）是由巨噬细胞和Ｂ淋巴细胞等产生的糖蛋白类细胞因子,由４０ｋＤ和３５ｋＤ两个亚基组成。ＩＬ－１２能促进Ｔｈｌ细胞的分化与增殖,控制细胞介导免疫;促进Ｔ淋巴细胞、ＮＫ细胞的发育与增殖,并刺激这些细胞产生ＩＦＮ－γ;增强ＮＫ细胞和细胞毒淋巴细胞的细胞毒活性以及ＩＬ－２诱导ＬＡＫ细胞的活力。ＩＬ－１２在培育新疫苗、防治利什曼病和ＡＩＤＳ等免疫系统疾病及肿瘤的免疫治疗中,有着广阔     

重组人卵泡刺激素在昆虫细胞中的表达

为了研究在昆虫细胞中表达重组人卵泡刺激素,我们以人胎盘组织提取的染色体DNA为模板,利用重叠PCR方法扩增出hFSHβ亚基的cDNA的编码区。将此cDNA克隆入核型多角体病毒(AcNPV)非融合蛋白基因表达载体pVLl393,我们得到了表达载体pVLl393-hFSHβ,然后与BaculoGold^TM线性杆状病毒DNA共转染昆虫细胞SF9,经多次扩增后获得高滴度的重组病毒AcNPV-hFSHβ。将此重组病毒感染昆虫细胞,我们得到了在胞浆中表达的hFSHβ亚基,Western blot显示分子量大约为21kDa。以重组病毒AcNPV-hFSHβ与AcNPV-hCGoL一同感染昆虫细胞得到了具有分泌性的重组hFSH异二聚体,在非还原的条件下Western blot显示分子量大约为33kDa。     

人绒毛膜促性腺激素β亚基和绵羊垂体促性腺激素共有α亚基组成的单链多...

hCG beta-oLH alpha chimeric cDNA was constructed by using overlapping PCR to contact the codons of C-terminal end of hCG beta with the codons of N-terminal end of oLH alpha, then it was subcloned into nuclear polyhedrosis virus (AcNPV) expression vector pVL1393 to construct expression vector pVL1393-hCG beta-oLH alpha. The insect cells (Sf9) were cotransfected by the expression vector pVL1393-hCG beta-oLH alpha and BaculoGold AcNPV linearized genomic DNA, and recombinant viruses AcNPV-hCG beta-oLH alpha were screened out by plaque assay. Further the insect cells were infected by the recombinant viruses, the recombinant hCG beta-oLH alpha was purified by immunoaffinity chromatography column coupling anti-hCG beta monoclonal antibody from the conditioned media of infected cells. The results of SDS-PAGE silver staining and western blotting showed that hCG beta-oLH alpha single peptide chain had apparent molecular weights of 40.5 kD and 38.0 kD under non-reducing and reducing conditions respectively, indicating the occurrence of disulfide bonds and significant tertiary structure in the single peptide chain. From the results of competitive inhibition of 125I-hCG beta binding we can conclude that the anti-hCG beta antibody-binding activity of hCG beta-oLH alpha chimera is lower than that of native hCG, but higher than that of native hCG beta. Therefore, we assume that the hCG beta-oLH alpha chimera should have potential application as a target antigen of anti-hCG fertility regulatory vaccine.     

人乳头瘤病毒16型E6基因的T—A克隆及在真核细胞中的表达

由于ＨＰＶ１６Ｅ６蛋白能诱导机体保护性免疫反应,可作为基因治疗的靶抗原。用杆状病毒昆虫细胞表达系统制备了ＨＰＶ１６Ｅ６基因工程蛋白,拟用于宫颈癌细胞系小鼠模型抗癌的免疫治疗。用ＰＣＲ技术从ＨＰＶ１６基因组中扩增获得转化基因Ｅ６的完整ＯＲＦ,按ＴＡ策略将其克隆到自行制备的杆状病毒转移载体ｐＶＬ１３９３Ｔ尾载体中,置于杆状病毒ＡｃＭＮＰＶＰｏｌｈ晚期启动子控制之下,用此重组转移质粒ｐＶＬ１３９３Ｅ６与杆状病毒ＤＮＡ共转染昆虫细胞Ｓｆ９,经噬斑筛选获得带有编码Ｅ６蛋白基因的重组杆状病毒株,并在昆虫细胞Ｓｆ９中表达为非融合性Ｅ６蛋白。ＳＤＳＰＡＧＥ电泳分析其分子量约为１８ｋＤ,免疫印迹实验表明,此重组蛋白能被兔抗ＨＰＶ１６Ｅ６抗体所识别。     

人绒毛膜促性腺激素β亚基和补体C3d片段组成的单链多肽的生物合成

In view of the strong immunity-enhancing function of HEL-C3d3 designed by Dr. Paul W. Dempsey, we made our efforts to produce a similar recombinant protein of hCG beta. With polymerase chain reaction, we introduced a Bam HI restriction site into the 3' terminal of hCG beta cDNA. The new cDNA and its terminal's correctness has been confirmed by sequencing. Then we have it covalently attached to the C3d3 cDNA at the pre-designed Bam HI/Bgl II site. Having the chimeric DNA correctly cloned into the protein nuclear polyhedrosis virus (AcNPV) expression vector pVL1393, we constructed the expression vector pVL1393-(hCG beta-C3d3). The insect cells were co-transfected with the expression vector and linearized nuclear polyhedrosis virus DNA, and recombinant viruses AcNPV-(hCG beta-C3d3) were screened out. Through anti-hCG beta immunoaffnity chromatography, the recombinant hCG beta-C3d3 chimera polypeptide was purified from culture supernatant of insect cells infected by the recombinant viruses. In RIA test, the expressed product competitively inhibits the binding of 125I-hCG beta to hCG beta-antibody. On SDS-PAGE and Western blot, the recombinant peptide hCG beta-C3d3 obviously appears to be with a molecular weight of 116KD. Therefore, we arrive at a conclusion that it has a normal immunogenic ability.     

水稻矮缩病毒外壳蛋白基因S8在昆虫细胞中的表达

将水稻矮缩病毒 (RiceDwarfVirus,RDV)外壳蛋白基因S8克隆到杆状菌毒转移载体 pVL1 393中 ,重组转移载体 pVL1 393 S8与线性杆状病毒RP2 3.LacZ共转染草地夜蛾细胞sf9,经过细胞体内同源重组、PCR筛选得到重组病毒RP2 3 S8。重组病毒RP2 3 S8感染sf9细胞后 ,收集细胞 ,并进行SDS PAGE及Western blot检测。结果表明 ,S8基因在昆虫细胞中成功表达 ,且在感染后 96h表达量最高     

人重组白细胞介素12在CHO细胞中的表达

将人白细胞介素12（interleukin 12, IL-12）两条链p35及p40全长cDNA分别亚克隆至真核表达载体pcDNA3中,构建了pcDNA3/p35a,pcDNA3/p40a,pcDNA3/p35b,pcDNA3/p40b四种真核细胞重组表达质粒,利用磷酸钙共沉淀法转染中国苍鼠卵巢(CHO)细胞. 通过对阳性克隆的筛选鉴定,获得了稳定表达人IL-12的CHO细胞株,活性最高的一株表达量为105 U/ml.此细胞株经半年的传代培养,能够稳定地分泌IL-12.结果显示：IL-12在CHO细胞中的稳定表达受到重组质粒结构、DNA整合、mRNA转录、蛋白质翻译等多因素的影响,IL-12两个亚基在CHO细胞中的共同表达是产生有生物活性IL-12的基础.     

Bruton酪氨酸激酶在昆虫细胞中表达的初步研究

用杆状病毒表达载体系统表达小鼠Ｂｒｕｔｏｎ酪氨酸激酶（Ｂｒｕｔｏｎｔｙｒｏｓｉｎｅｋｉｎａｓｅ,Ｂｔｋ）．构建重组转染载体时,于Ｂｔｋ起始码的上游插入了一段Ｈ９０２序列．用重组转染载体、苜蓿夜蛾核型多角体病毒线性ＤＮＡ和质脂体共转染Ｓｆ９昆虫细胞,经过对重组杆状病毒三轮扩增后,用Ｈ９０２抗体检测表明,昆虫细胞中Ｂｔｋ的表达已达最高水平．对表达后Ｂｔｋ自身磷酸化检测表明,该激酶具有自身磷酸化活性．从而证实Ｂｔｋ在昆虫细胞中的表达获得了成功     

Cloning and expression of murine IL-12.

Human IL-12 (NK cell stimulatory factor, cytotoxic lymphocyte maturation factor) is a heterodimeric cytokine that can act as a growth factor for activated human T and NK cells, enhance the lytic activity of human NK/lymphokine-activated killer cells, and stimulate the production of IFN-gamma by resting human PBMC. Because in our hands, human IL-12 did not elicit similar responses in murine lymphocytes, we have cloned and expressed the murine IL-12 subunit cDNA in order to obtain recombinant protein for murine studies. Comparison of the predicted amino acid sequences of the murine subunits with their human counterparts revealed that the p40 subunits are more highly conserved than the p35 subunits (70% vs 60% identity, respectively). The sizes of the p35 and p40 subunit mRNA were estimated to be 1.5 kb and 2.6 kb, respectively. RNA blot analysis showed that p35 mRNA was expressed in lymphoid tissues (spleen, thymus) and nonlymphoid tissues (lung, brain), whereas p40 mRNA expression was only detected in lymphoid cells. Incubation of splenocytes with pokeweed mitogen did not significantly affect p35 mRNA levels, however, it resulted in a decrease of p40 mRNA. Coexpression of the murine p35 and p40 cDNA clones in COS cells resulted in the secretion of IL-12, which was active in human and mouse T cell proliferation, murine NK cell activation, and murine IFN-gamma induction assays. Transfection of each subunit cDNA alone did not result in measurable secreted IL-12 activity. A hybrid heterodimer consisting of murine p35 and human p40 subunits retained bioactivity on murine cells; however, the combination of human p35 and murine p40 was completely inactive on murine cells. These results indicate that the observed inability of human IL-12 to act on murine cells is largely determined by the p35 subunit.     

序列优化的小鼠IL-33基因在哺乳动物细胞中的有效分泌表达

目的: 白细胞介素(IL)-33具有重要的免疫调控作用,在疾病中扮演着重要的角色。本文旨在通过基因优化实现IL-33在哺乳动物细胞中的高效表达,为疾病机理研究以及疫苗免疫佐剂应用等提供基础。方法: 根据小鼠白细胞介素-33成熟肽(mIL-33)的氨基酸序列,以哺乳动物细胞基因表达密码子偏好性进行基因优化设计;化学合成优化的mIL-33基因片段,通过搭桥PCR将编码人CD8α信号肽的核酸序列分别与优化或未优化的mIL-33基因连接,并与绿色荧光蛋白(EGFP)基因分别构建到双表达单元质粒 pBudCE4.1的不同启动子下;重组质粒经 lipofectamine 3000 和PEI转染293FT 细胞;以 Western blot和ELISA检测重组蛋白的表达;收集表达的mIL-33刺激巨噬细胞Raw264.7,ELISA检测培养上清的TNFα水平,以证明IL-33的生物学活性。结果: 重组质粒经酶切鉴定及测序分析证实构建成功; lipofectamine 3000转染效率较PEI转染更高;Western blot和ELISA 结果显示密码子优化的mIL-33表达水平较未优化序列更高,在EF-1α启动子和CMV启动子指导下mIL-33在293FT 细胞表达水平相当,CD8α信号肽成功引导mIL-33的分泌,产物具生物学活性。结论: 密码子优化操作显著改善了 mIL-33在哺乳动物细胞中的表达水平,为进一步研究奠定了基础。