二代测序实验与测序原理讲义.ppt

二代测序的建库与测序原理;内容;TRUSEQ RNA AND DNA SAMPLE PREPARATION;CLUSTER GENERATION OVERVIEW;OH;diol;TEMPLATE PREPARATION-BRIDGE RCR;;CYCLIC REVERSIBLE TERMINATION;BASE CALLING;FLOWCELL LAYOUT ON GAII;PRIMARY DATA ANALYSIS BY FIRECREST AND BUSTARD IN RTA/OLB ;;SEQUENCE MULTIPLE SAMPLES IN THE SAME LANES;PAIR-END 测序优势;MATE-PAIR 建库和测序;TEMPLATE PREPARATION- EMULSION PCR;PYROSEQUENCING;BASE CALLING;*;454、SOLEXA测序模式;Detect H+ released as a voltage change—fast Common microchip design standards—low-cost manufacturing Sequencing volume is increasing ;FASTA序列格式 ;ILLUMINA SEQUENCE IDENTIFIERS;ILLUMINA SEQUENCE IDENTIFIERS;序列质量;SSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSSS......................................... ..........................XXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXX.......... ...............................IIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIII.......... .................................JJJJJJJJJJJJJJJJJJJJJJJJJJJJJJJJJJJJJJJ.......... LLLLLLLLLLLLLLLLLLLLLLLLLLLLLLLLLLLLLLLLLL........................................ !#$%()*+,-./0123456789:;=?@ABCDEFGHIJKLMNOPQRSTUVWXYZ[\]^_`abcdefghijklmnopqr | | | | | 33 59 64 73 104 0........................26...31.......40 -5....0........9.............................40 0........9.............................40 3.....9.............................40 0........................26...31........41 S - Sanger Phred+33, raw reads typically (0, 40) X - Solexa Solexa+64, raw reads typically (-5, 40) I - Illumina 1.3+ Phred+64, raw reads typically (0, 40) J - Illumina 1.5+ Phred+64, raw reads typically (3, 40) with 0=unused, 1=unused, 2=Read Segment Quality Control Indicator (bold) (Note: See discussion above). L - Illumina 1.8+ Phred+33, raw reads typically (0, 41) ;;;;454原始数据图片、SFF格式、FASTA格式（Q