Navitoclax_HNMR_26062_MedChemExpress
大动脉炎患者外周血单个核细胞RT-qPCR内参基因的选择
January 2021Vol.41 No.12021 年 1 月 第 41 卷 第 1 期基础医学与临床Basic & Clinical Medicine文章编号：1001-6325 ( 2021 ) 01-0087-06研究论文大动脉炎患者外周血单个核细胞RT-qPCR 内参基因的选择田苡箫，李菁*收稿日期：2019-11-18 修回日期：2020-04-30*通信作者(corresponding author ) ：lijing6515@ （中国医学科学院北京协和医学院北京协和医院风湿免疫科风湿免疫病学教育部重点实验室国家皮肤与免疫疾病临床医学研究中心，北京100032）扌摘要：目的筛选适于在大动脉炎（TAK ）患者和健康人群（HC ）之间比较外周血单个核细胞（PBMC ）中mRNA 表达水平的内参基因。
方法提取PBMC 中的总RNA,应用RT-qPCR ，分别采用geNorm 、NormFinder 、BestKeeper 3种软 件程序，分析 3-glucuronidase ,GAPDH ,ACTB ,SDHA ,HPRT1,RPL13A ,B2M , YWHAZ 和 PKG1 9 个基因的 mRNA 表达稳定性。
以T-bet 、GATA3和RORC 作为目的基因，比较不同稳定性的内参基因对mRNA 相对丰度的影响。
结果geNorm 筛选得到的基因组合为B 2M-SDHA , Nor^nFinder 和BestKeeper 筛选出最稳定的内参基因均为HPRT1 ； 3种方法均显示GAPDH 的稳定性较差。
结论自身免疫病患者在接受免疫抑制药物治疗时,原本稳定表达的基因可能会 上调或下调;在样本量较小时，稳定性更好的内参基因可能更有助于检测组间差异。
关键词：大动脉炎;实时定量聚合酶链式反应;RNA 稳定性；内参基因选择中图分类号:R593.2 文献标志码:AValidation of reference genes for the normalization of the RT-qPCRin peripheral blood mononuclear cells of patients with Takayasu arteritisTIAN Yi-xiao ， LI Jing *(Department of Rheumatology and Immunology , Key Laboratory of Rheumatology and Clinical Immunology , Ministry of Education ,National Clinical Research Center for Dermatologic and Immunologic Diseases ( NCRC-DID ),Peking Union Medical College Hospital , CAMS & PUMC , Beijing 100032, China)Abstract ： Objective To validate proper reference genes for quantitative real-time polymerase chain reaction ( RT-qPCR) used for comparing mRNA expression levels in Takayasu arteritis" (TAK) and healthy controls' ( HC ) pe ripheral blood mononuclear cells ( PBMC ). Methods Total RNA in PBMCs was extracted and used RT-qPCR to determine the profiles of 9 candidate genes , including 0-glucuronidase, GAPDH , ACTB , SDHA , HPRT1, RPL13A , B2M , YWHAZ and PKG1. Then compared their transcription stability by geNorm , NormFinder , and Best Keeper. Afterwards , with T-bet , GATA3 and RORC as the targeted genes , explored the influence of reference genes with different stability on mRNA relative abundance. Results The gene combination of B2M-SDHA was selected bygeNorm , and HPRT1 was the most stable one in analysis results of NormFinder and BestKeeper , while GAPDH was less stable. Conclusions Genes that have been expressed stably may be upregulated or downregulated whenpatients with autoimmune diseases received immunosuppressive drugs. When the sample size is small ， the more sta ble internal reference may facilitate the identification of inter-groups difference.Key words : Takayasu arteritis ； real-time polymerase chain reaction ； RNA stability ； selection of reference gene88基础医学与临床Basic&Clinical Medicine2021.41(1)反转录实时荧光定量聚合酶链式反应（reverse quantitative real-time polymerase chain reaction,RT-qPCR）是目前分析基因表达水平的黄金标准，却经常表现出重复性欠佳的问题，选取合适的内参基因有助于改善这一情况［1］。
TMA-DPH-SDS-MedChemExpress
Inhibitors, Agonists, Screening LibrariesSafety Data Sheet Revision Date:Mar.-25-2019Print Date:Mar.-25-20191. PRODUCT AND COMPANY IDENTIFICATION1.1 Product identifierProduct name :TMA-DPHCatalog No. :HY-D0986CAS No. :115534-33-31.2 Relevant identified uses of the substance or mixture and uses advised againstIdentified uses :Laboratory chemicals, manufacture of substances.1.3 Details of the supplier of the safety data sheetCompany:MedChemExpress USATel:609-228-6898Fax:609-228-5909E-mail:sales@1.4 Emergency telephone numberEmergency Phone #:609-228-68982. HAZARDS IDENTIFICATION2.1 Classification of the substance or mixtureNot a hazardous substance or mixture.2.2 GHS Label elements, including precautionary statementsNot a hazardous substance or mixture.2.3 Other hazardsNone.3. COMPOSITION/INFORMATION ON INGREDIENTS3.1 SubstancesSynonyms:NoneFormula:C28H31NO3SMolecular Weight:461.62CAS No. :115534-33-34. 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安捷伦产品目录
15
Real-Time PCR
16
Mx3000P QPCR System
17
Brilliant III Ultra-Fast SYBR Green QPCR and QRT-PCR Reagents
18
Brilliant III Ultra-Fast QPCR and QRT-PCR Reagents
Agilent / STRATAGENE
Agilent website: /genomics
Welgene | Agilent Stratagene
威健股份有限公司 | Stratagene 總代理
Table of Content
Table of Contents
/ XL1-Red Competent Cells SoloPack Gold Supercompetent Cells
/ TK Competent Cells Specialty Cells
/ Classic Cells / Fine Chemicals For Competent Cells
適用於 UNG 去汙染或 bisulphite
sequencing
適用於 TA Cloning
最高敏感性
取代傳統 Taq 的好選擇
-
2
威健股份有限公司 | Stratagene 總代理
PCR Enzyme & Instrument
Agilent SureCycler 8800
市場上領先的 cycling 速度和 sample 體積 10 ～ 100 μL 簡易快速可以選擇 96 well 和 384 well 操作盤 優秀的溫控設備讓各個 well 都能保持溫度的穩定 七吋的高解析度觸控螢幕讓操作上更為簡便 可以透過網路遠端操控儀器及監控儀器 Agilent 專業的技術支援可以幫助您應對各種 PCR 的問題
抗D说明书
生产厂家：英国Millipore (UK)Limited中国大陆地区总代理上海博力医学生物科贸有限公司产品简介：抗D（IgM+IgG）血型定型试剂直接凝集法可识别D，弱D和不完全D间接抗人球蛋白实验可识别DVI。
效价达1：256，IgM克隆号Th-28，IgG克隆号MS-26，该产品已通过微米的过滤膜过滤，并在胰蛋白酶大豆琼脂中培养3至6天和在沙氏葡萄糖琼脂中培养5至7天后通过一个生物负荷测试<1个细菌群落每毫升（CFU/ml）HIV、HCV、HBsAg、EBV、MAP病毒检测阴性，有效期3年。
注册证号：国食药监械（进）字2010第3403117号使用此产品的临床意义：抗D （IgM）不与DVI细胞反应，不能检测DVI （弱D）；抗D （IgM+IgG）中的IgG能与DVI 细胞反应，能检测DVI （弱D）。
该产品应用于玻片法、室温盐水试管法、微孔板法和间接抗人球蛋白法检测人红细胞中的D抗原。
产品的预期应用项目和使用量：Rh 血型定型试剂；检验方法：玻片法、室温盐水试管法、间接抗人球蛋白实验（用于弱D样本的检测）、微孔板法；使用量：根据实验室实验方法的不同而定。
详细请看下面：？抗D（IgM+IgG）血型定型试剂（单克隆抗体）操作规程【产品名称】通用名称：抗D（IgM+IgG）血型定型试剂（单克隆抗体）英文名称：Anti-D Blood Grouping Reagent (Monoclonal Human IgM/IgG Blend)【包装规格】10mL/瓶，10瓶/盒。
【预期用途】该产品应用于玻片法、室温盐水试管法、微孔板法和间接抗人球蛋白法检测人红细胞中的D抗原。
红细胞RhD亚型系统最早是由Levine和Stetson在1939年提出来的。
RhD阴性的人在接受RhD阳性血液输血或者孕有RhD阳性的胎儿时，会产生抗-D抗体，从而导致严重的新生儿溶血症或输血性溶血反应。
“弱D型”通常用来表述RhD抗原数目偏少的红细胞；“部分D型”通常用来表述缺乏部分RhD抗原表位的红细胞；“D VI”属于“部分D型”的范畴。
Keap1
非小细胞肺癌(non-small cell lung cancer，NSCLC)发病率占据肺癌的75%~80%。
肿瘤细胞进展快且易扩散转移，临床常采用手术、放化疗等进行治疗，但5年生存率低于60%[1-2]。
氧化应激是由活性氧(ROS)生成量增加所致，ROS积累可诱导肺癌细胞凋亡，清除ROS 可阻止癌细胞凋亡，即肺癌细胞存活依赖于癌细胞自身抗氧化能力[3]。
Kelch样环氧氯丙烷相关蛋白-1 (kelch-like epichlorohydrin-associated protein-1，Keap1)/核因子E2相关因子2(nuclear factor E2related factor 2，Nrf2)信号通路在癌症中发挥重要调控作用，氧化应激可激活Keap1，促使Keap1-Nrf2复合物裂解，Nrf2转移至细胞核内，可激活下游靶基因表达，参与肺癌发生发展过程[4]。
Nrf2可维持氧化还原稳态，ROS侵袭细胞时，Nrf2可进入细胞核，结合抗氧化反应元件(ARE)转录编码各种抗氧化蛋白、代谢酶基因，抑制氧化应激反应[5-6]。
目前氧化应激、Keap1/Nrf2信号通路在NSCLC发生过程中的机制尚未明确。
基于此，本研究尝试分析Keap1/Nrf2信号通路与临床病理参数、氧化应激指标的相关性，探讨其在NSCLC氧化应激机制中的作用，为临床研制新药提供参考依据。
1资料与方法1.1一般资料选取2017年4月至2020年4月郑州市第三人民医院收治的100例NSCLC患者为研究对象。
纳入标准：符合NSCLC诊断标准[7]；术前未接受放化疗、免疫治疗者；预计生存期≥6个月；符合手术适应证、禁忌证；Karnofsky功能状态评分≥70分；签署知情同意书。
排除标准：合并凝血功能障碍、肝肾功能障碍、其他恶性肿瘤者；伴有急/慢性感染者；伴有精神疾病者；既往腹部相关外科手术史者。
所有患者均行肺癌根治性切除术，术中收集癌组织、癌旁组织(距离癌组织5cm范围内正常组织)，其中男性63例，女性37例；年龄46~67岁，平均(56.32±3.16)岁；体质量指数(BMI)17~30kg/m2，平均(23.16±2.03)kg/m2；病理类型：鳞癌58例、腺癌42例；病理分级[8]：Ⅰ~Ⅱ级51例、Ⅲ级49例；T分期[9]：T1~T253例、T3~T447例；N分期：N055例、N1~N245例。
15523666_迷迭香酸对哮喘小鼠氧化性肺损伤的保护作用
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