CHARACTERIZATION OF THE CELLULAR LOCALIZATION OF BASE

基地细胞定位的表征

• 批准号: 8360634
• 负责人: KRISTI A EGLAND
• 金额: $ 0.25万
• 依托单位: UNIVERSITY OF SOUTH DAKOTA
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-05-01 至 2012-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8360634
• 关键词: Amino Acid Sequence; Amino Acids; Biomedical Research; Breast Cancer Cell; Cancer cell line; Cell Extracts; Cells; Culture Media; Evaluation; Family; Funding; Genes; Grant; Hour; Human; Immunoblot Analysis; In Vitro; Libraries; Membrane Proteins; Monoclonal Antibodies; N-terminal; National Center for Research Resources; Normal tissue morphology; Oncogenes; Open Reading Frames; Peptide Signal Sequences; Plasmids; Principal Investigator; Proteins; Research; Research Infrastructure; Resources; Salivary Glands; Source; South Dakota; System; Tumor Markers; United States National Institutes of Health; Western Blotting; base; cost; disorder risk; latherin; malignant breast neoplasm; outcome forecast; response; vector

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Tumor markers provide information for assessing disease risk, prognosis, metastatic potential and evaluation of response to therapy. Previously, a new breast cancer gene designated BASE (breast cancer and salivary gland expression) has been identified by generating a library that was enriched for genes encoding secreted and membrane proteins. Of 62 normal tissues analyzed, BASE was only expressed in salivary gland. BASE was also expressed in human breast cancers. The deduced amino acid sequence of BASE predicts that it is secreted. BASE is predicted to have a 20 amino acid N-terminal signal sequence, and has homology to other secreted proteins, such as Latherin and the PLUNC family. To further characterize BASE, we need to verify its cellular localization. As a first step, an in vitro expression system using 293T cells and the ZR-75-1 breast cancer cell line will be developed. A plasmid will be constructed containing the BASE ORF cloned 5' of a Myc tag using the vector pcDNA3.1-Myc/His. This expression construct will be transfected into 293T and ZR-76-1 cells. After 48 hours, the culture medium and total cell extracts will be examined for the presence of BASE-Myc by Western immunoblot analysis using a monoclonal antibody against the Myc tag.

该副本是利用资源的众多研究子项目之一 由NIH/NCRR资助的中心赠款提供。对该子弹的主要支持 而且，副投影的主要研究员可能是其他来源提供的 包括其他NIH来源。 列出的总费用可能 代表subproject使用的中心基础架构的估计量， NCRR赠款不直接向子弹或副本人员提供的直接资金。 肿瘤标记提供了评估疾病风险，预后，转移性潜力和对治疗反应的评估的信息。 以前，通过生成富含编码分泌和膜蛋白的基因的文库来鉴定出一种新的乳腺癌基因指定碱（乳腺癌和唾液腺表达）。 在分析的62个正常组织中，仅在唾液腺中表达碱。 碱还在人类乳腺癌中表达。 推导的碱酸序列预测其分泌。 预计基碱具有20个氨基酸N末端信号序列，并且与其他分泌的蛋白质（例如燃烧林和PLUNC家族）具有同源性。 为了进一步表征碱，我们需要验证其细胞定位。 作为第一步，将开发使用293T细胞和ZR-75-1乳腺癌细胞系的体外表达系统。 将使用vector pcDNA3.1-myc/His构建质粒，其中构建包含MYC标签的基本ORF 5'。 该表达构建体将转染到293T和ZR-76-1细胞中。 48小时后，将检查培养基和总细胞提取物通过西方免疫印迹分析使用抗MYC标签的单克隆抗体来检查碱基Myc。