rnai用于基功能的鉴定和确证.pptVIP

Renilla Luciferase data Firefly Luciferase data miRNA的编辑及其靶标的变化 seed elment： A I miR376a-5p 经过编辑后，其靶标的变化 FulenGen OmicsLink? Human miRNA Target Sequence (3’ UTR) Expression Clones SV40 hLuc RLuc CMV polyA polyA pUC ori Kan/Neo 3’ UTR SV40 GLuc sAP CMV polyA polyA pUC ori Kan/Neo 3’ UTR pEZX-MT01 pEZX-MT02 FulenGen Gaussia luciferase expression in CHO cells FulenGen 构建克隆 转染细胞 分泌表达Luciferase 取上清 加入底物 读取荧光值 FulenGen miRNA Deliver into cell, tissue or animal Restore the silence gene function mRNA cleavage or translation inhibition Silence gene function 基因沉默后的功能拯救 RNAi临床药物开发 Thank You! Phone： 020 FAX：020 Email：Support@, Sales@ * * RNAi(shRNA/miRNA)用于基因功能的鉴定和确证 广州复能基因有限公司 李华平 博士 研发部主任 FulenGen Gain of function ORF expression clones Tansfection/Transduction Loss of function shRNA/miRNA Chemical genetics Inhibitors and activators of biological process Automated read-out mechanism Therapeutic strategy cell FulenGen Gene Function Screening Mechanism of RNAi (shRNA/miRNA) Comparison of shRNA and miRNA properties Architecture short hairpin Single-strand Original Exogenous Endogenous No. of intended target 1 Many Degree of gene silence High Low Complementary to mRNA target Perfect Imperfect Effect on mRNA Endonucleolytic cleavage Repressed translation shRNA miRNA FulenGen FulenGen shRNA的设计与构建 shRNA的验证 miRNA的克隆与检测 miRNA靶标的确证 基因沉默后的功能拯救 FulenGen FulenGen shRNA 表达克隆的构建 载体选择 筛选标记和报告基因 目的细胞 shRNA序列设计 高准确性的预测方法 茎和环序列 有效性检测 3-5个shRNA /基因 目的基因是否表达 克隆和测序 合成引物的长度 茎环结构难以有效测序 27mer shRNA Oligo Design 5’ GATCGAGAGTGGAGGACGCTTTGCCTCCAAGCTCAA