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The PCR products were examined by agarose gel electrophoresis. The target gene fragments were purified by gel extraction kit and ligated to cloning vector pMD18-T. The recombinant vectors were transformed into host strain E. coli K802 by lithium chloride method, screened and identified with PCR and restrictive enzymatic digestion. Their sequences were confirmed by DNA sequencing.(2) sTWEAK1 gene was subcloned into expression vector pProEx HTb and transformed into E. coli BL21. sTWEAK2 gene was subcloned into expression vector pMAL-C2x and transformed into E. coli TB1. The recombinant vectors were screened and identified with PCR and restrictive enzymatic digestion. The recombinant fusion proteins were induced to express with IPTG, detected by coomassie brilliant blue-stained SDS-polyacrylamide gel electrophoresis , and confirmed by Western blot analysis.(3) The sTWEAK1 fusion protein was purified with Ni-NTA Spin Kit.(4) The biological activity was assayed on transformed and tumor cells by microplate photometer after crystal violet or sulfur rodamine B staining.(5) The contents of IL-8 in the supernatant of 1990 cell cultures were determined by ELISA.(6) The morphological changes of the sensitive cells were observed by light and transmission electron microscopies.(7) The cell cycle and apoptotic rate were assayed by flow cytometry in 1990 and M85 cells.(8) The effect of fusion proteins on induction of NF-κB in 1990 and LOVO cells was detected with Dual-Luciferase Reporter Assay system.(9) The TWEAK gene was subcloned into Adeno-X Viral DNA with pShuttle vector and transfected into HEK293 cells by lipofectamine method.

(1)本研究用RT-PCR方法，从人组织细胞总RNA中扩增可溶性TWEAK胞外区（sTWEAK1和sTWEAK2）的cDNA序列及全长编码序列，用琼脂糖凝胶电泳分析PCR产物，胶回收目的基因片段，连接到pMD18-T克隆载体中，转化大肠杆菌K802，PCR和酶切筛选阳性克隆，全自动DNA测序验证序列；(2)sTWEAK1和sTWEAK2分别亚克隆到pProEx HTb和pMAL-C2x表达载体中，分别转化大肠杆菌BL21和TB1，PCR筛选和酶切鉴定，阳性克隆用IPTG诱导表达，表达产物用SDS-PAGE分析和Western blot验证融合蛋白；(3)用NTA-Ni Spin试剂盒初步分离纯化sTWEAK1融合蛋白；(4)用体外培养的肿瘤细胞和正常对表达产物进行活性检测，贴壁细胞用结晶紫染色法，悬浮细胞用磺酰罗丹明B染色法，酶标仪检测OD值；(5)敏感细胞用ELISA法检测细胞培养上清中IL-8的含量；(6)用光镜和电镜观察敏感细胞死亡和细胞凋亡情况；(7)用流式细胞仪分析表达产物对敏感细胞凋亡率和细胞周期的影响；(8)用双荧光素酶报告基因检测法，测定表达产物对敏感细胞NF-κB的影响；(9)用pShuttle穿梭质粒将TWEAK重组到腺病毒载体上，用脂质体转染法转染HEK293细胞，PCR鉴定重组质粒。

Here's a list of Lex Luthor's known lairs straight from the Superman Encyclopedia: a complex of buildings held aloft by a giant dirigible high above the stratosphere, a glass-enclosed city of ancient, weird design, an abandoned factory, a gigantic man-made meteor floating in outer space, an abandoned barn, a secret underground lab, a giant spaceship, a secret mountaintop laboratory, an electronics firm, a hidden laboratory on the outskirts of Metropolis, a laboratory hideout that has been lined with lead to conceal it from Superman's X-ray vision, a massive fortress-like citadel on a lonely mountaintop north of Metropolis, a lonely farmhouse in the mountains north of Metropolis, a secret lead-lined subterranean hideout built into the side of a grassy hill, and the elaborately equipped Luthor's Lair, Luthor's Lair II, and Luthor's Lair No.

关于莱克斯·卢瑟已知的据点，超人的百科全书上是这样记载的："他的藏身处包括由巨大飞船托浮在半空的复杂建筑群；古代样式的玻璃城，设计诡异；废弃的工厂；悬浮在外太空的巨大人造陨石；废弃的谷仓；秘密的地下实验室；巨大的太空船；建于山顶的秘密实验室；电子公司；藏匿于城郊的实验室，这个实验室通体灌铅，能够躲过超人的X光眼；巨大的堡垒--它就像孤零零伫立在城市北面山顶上的一个据点；坐落在城市北面山上的孤零零的农舍；还有建在绿岭山脊上的秘密铅制地下室；以及装备精良的卢瑟堡一号，二号和5号。"

Because 95% of China's e-bikes use lead batteries, they emit more lead into the atmosphere than other forms of transportation, according to some studies.

由于中国95%的电动自行车使用铅电池，一些研究结果显示，电动自行车向大气中排放的铅要高于其他类型的交通工具。

Methods: Based on the value added and the absorbance of the sample in the beer into lead series solution, increasing the absorbance values, enhance the sensitivity of the method, the direct determination of lead content in beer was performed.

依据吸光度值的加和性，在啤酒样品中加入铅标准系列溶液，增大吸光度值，提高方法的灵敏度，直接测定啤酒中铅含量。

Through seeking for a scientific and effectual evaluation theory and method on operating achievement for non-stated listed companies, it will urge them to know clearly about their real management status, to lead their enterprises to put scientific management into practice and to level up their operation. Thus non-stated listed companies can be in an indefectible position in the drastic market competition, what's more they can lead all non state-owned enterprises to grow in strength endlessly.

通过探索建立科学有效的民营上市公司的经营业绩评价理论和方法，将促使民营上市公司认清企业的真实经营状况，引导企业进行科学管理，促进民营上市公司的管理水平提高到一个新的层次，这样才有可能在激烈的市场竞争中立于不败之地，进一步带动所有的民营企业不断发展壮大。

Lure,entice,inveigle,decoy,tempt,seduce These verbs mean to lead or attempt to lead into a wrong or foolish course, as of action.

这些动词意思是引导或试图诱导至错误或愚蠢的道路，如行动的道路。

In addition, many small businesses there is no perfect environmental protection facilities, high levels of lead smelting process steam, lead dust, sulfur dioxide and other harmful substances into the atmosphere, not only the job-site labor conditions also cause serious environmental pollution.

此外，许多小型企业没有完善环保设施，铅冶炼过程中的蒸汽高，到大气中铅尘，二氧化硫和其他有害物质，不仅是施工现场劳动条件，同样会造成严重的环境污染。

Mine of zinc of lead of yellow sanded level ground was 1967 the mine of plumbic zinc backbone of put into production, zinc of house home lead produces mine the 7th.

黄沙坪铅锌矿是1967年投产的铅锌骨干矿山，居国内铅锌生产矿山第七位。

Our so-called " lead ers" couldn't lead children into a sweetshop, it seems.

看来我们所谓的"领导"并不能把孩子们领到糖果店去。

C Supports both Firmware Hub and LPC Memory Read and Write Cycles Auto-detection of FWH and LPC Memory Cycles C Can Be Used as FWH for Intel 8xx, E7xxx, and E8xxx Series Chipsets C Can Be Used as LPC Flash for Non-Intel Chipsets Flexible, Optimized Sectoring for BIOS Applications C 16-Kbyte Top Boot Sector, Two 8-Kbyte Sectors, One 32-Kbyte Sector, Three 64-Kbyte Sectors C Or Memory Array Can Be Divided Into Four Uniform 64-Kbyte Sectors for Erasing Two Configurable Interfaces C FWH/LPC Interface for In-System Operation C Address/Address Multiplexed Interface for Programming during Manufacturing FWH/LPC Interface C Operates with the 33 MHz PCI Bus Clock C 5-signal Communication Interface Supporting Byte Reads and Writes C Two Hardware Write Protect Pins: TBL for Top Boot Sector and WP for All Other Sectors C Five General-purpose Input Pins for System Design Flexibility C Identification Pins for Multiple Device Selection C Sector Locking Registers for Individual Sector Read and Write Protection A/A Mux Interface C 11-pin Multiplexed Address and 8-pin Data Interface C Facilitates Fast In-System or Out-of-System Programming Single Voltage Operation C 3.0V to 3.6V Supply Voltage for Read and Write Operations Industry-Standard Package Options C 32-lead PLCC C 40-lead TSOP

0第0页，本页显示记录0-0，共0条记录分0页显示C支持两种固件中心和LPC内存读取和写入周期自动的FWH和LPC的记忆圈C检测可以用于英特尔8xx系列，E7xxx，E8xxx系列芯片组和C可以用作FWH与至于非英特尔芯片组的BIOS应用柔性优化扇区开放16字节热门引导扇区，两个8 - Kbyte的，一个32字节部门，3个64 - Kbyte的C或存储阵列，线性预测编码闪光可分为四个统一为两个可配置的接口擦除的FWH / LPC接口为64 - Kbyte的行业，系统运行C地址/地址多路复用在制造过程中用于编程接口的FWH /线性预测编码界面C与33 MHz的PCI总线时钟 5信号通信接口进行操作，支持字节读取和写入引导扇区的顶部和WP C两硬件写保护引脚：任务型为所有其他部门 5个通用输入的系统设计的灵活性识别的多种设备选型部门登记销锁定为个别部门读取和写保护的A /阿复用界面C 11引脚复用引脚地址和8引脚的数据界面C促进快速系统内或外的系统编程的单电压3.0V至3.6V的操作供应的读取和写入操作业界标准的封装选项电压 32 -引脚PLCC 40引脚的TSOP

No, it's unnecessary. I don't think I have a fever.

不，没有必要，我想我没有发烧。

We live in a culture of instant gratification.

我们生活在即时满足的文化中。