CAJ | 학술논문

研制人线粒体 DNA（mitochondrial DNA，mtDNA）表达谱芯片，鉴定芯片的特异性和稳定性，完善芯片的制备和使用程序，优化实验参数，为线粒体功能研究提供物质基础及技术保障。以人 mtDNA 剑桥序列为标准，设计 mtDNA 编码的13种 mRNA、2种 rRNA 和9种 tRNA 基因及蛋白产物定位于线粒体的核 DNA （nDNA）编码的5种凋亡相关基因寡核苷酸探针。芯片点样仪点制芯片，进行荧光显示和洗脱，用其检测人宫颈癌上皮 Hela 细胞和人肾小管上皮 Hc1细胞 cDNA 文库 mtDNA 编码基因及 nDNA 编码凋亡相关基因的差异表达情况。所点制的芯片扫描结果显示，样点分布均匀、清晰，规整度好，无漏点、连点。cDNA 文库杂交鉴定结果显示，整张芯片荧光信号均匀一致，背景清晰，各基因重复样点杂交结果一致，各质控点能正常显示。成功制备了人 mtDNA 基因表达谱芯片，完善了该芯片的制备和使用程序，通过初步应用，证明所研制的人 mtDNA 基因表达谱芯片具有特异性高、稳定性好等优点，可用于不同组织或细胞样本 cDNA 文库 mtDNA基因的差异表达分析。
연제인선립체 DNA（mitochondrial DNA，mtDNA）표체보심편，감정심편적특이성화은정성，완선심편적제비화사용정서，우화실험삼수，위선립체공능연구제공물질기출급기술보장。이인 mtDNA 검교서렬위표준，설계 mtDNA 편마적13충 mRNA、2충 rRNA 화9충 tRNA 기인급단백산물정위우선립체적핵 DNA （nDNA）편마적5충조망상관기인과핵감산탐침。심편점양의점제심편，진행형광현시화세탈，용기검측인궁경암상피 Hela 세포화인신소관상피 Hc1세포 cDNA 문고 mtDNA 편마기인급 nDNA 편마조망상관기인적차이표체정황。소점제적심편소묘결과현시，양점분포균균、청석，규정도호，무루점、련점。cDNA 문고잡교감정결과현시，정장심편형광신호균균일치，배경청석，각기인중복양점잡교결과일치，각질공점능정상현시。성공제비료인 mtDNA 기인표체보심편，완선료해심편적제비화사용정서，통과초보응용，증명소연제적인 mtDNA 기인표체보심편구유특이성고、은정성호등우점，가용우불동조직혹세포양본 cDNA 문고 mtDNA기인적차이표체분석。
To manufacture and identificate the expressing microarray of human mtDNA genes.In the process of conformation,the specificity and stability of the new microarray were evaluated,the procedure of preparing and applying of the new microarray were con-summated,and the experimental parameter were optimised to provide the substance support and technology guarantee for the function research of mitochondrial genome.Design of the oligonucleotide probes of 13 mRNA、2 rRNA、9 tRNA encoded by mtDNA was based on the mtDNA Cambridge Sequence,and the oligonucleotide probes of 5 apoptosis related genes encoded by nuclear genome was based on the database of Genebank.The microarray was printed with microarray robots.Fluorescence background manifest and deletion were ac-cording to the standard process.The primary microarray was firstly used to detect the differential expressions of mtDNA genes and apop-tosis related genes between cDNA library of human cervix epithelia carcinoma cell line Hela and human renal tubule epithelia cell line Hc1.The printed spots on the primary microarray show distinct and regular on the scanned imagine,and the background of the slides is nearly undetected.The hybridized imagine of cDNA library from cell lines show a good quality in size and color contrast.Twenty mini -blocks are printed in the first round.