基因表达系列分析(Serial Analysis of Gene Expression,SAGE)技术（Serial Analysis of Gene Expression, SAGE）.doc

基因表达系列分析(Serial Analysis of Gene Expression,SAGE)技术（Serial Analysis of Gene Expression, SAGE） Gene expression series analysis In 1995, Velculescu and others proposed the Serial Analysis of Gene Expression, SAGE, which can simultaneously study thousands of transcripts. 1. The principle and experimental route of SAGE. 1.1 principles of SAGE SAGE has two main bases. First, a 9-10 base short nucleotide sequence tag contains enough information to uniquely identify a transcription. 9 a sequence of bases, for example, can distinguish 262144 different transcription (49), and the human genome is estimated only can encode 80000 kinds of transcription, so in theory every 9 base label to represent the characteristics of sequence a transcription. Second, if can be 9 base label focused on a cloning sequencing, and will receive the short sequences of nucleotide sequence in the form of a continuous data input in the computer for processing, can analyze the mRNA transcription of thousands. 1.2 the experimental route of SAGE. As shown in figure 1: (1) using biotinylated oligo (dT) to synthesize cDNA for the primers, in a restriction Enzyme (Anchoring Enzyme Anchoring, AE) Enzyme. The anchoring enzyme requires at least one enzyme cutting site on each transcriptional object, and the average 4 base restriction enzyme can meet this requirement, since most mrnas are longer than 256 bases (44). CDNA3 was collected by strepsin anti-biotin beads. For each mRNA, only fragments of its polyA tail and the nearest enzyme cutting site were collected. (2) the cDNA is divided into two parts, A and B, respectively. Each joint containing the tag Enzyme (Tagging Enzyme TE) Enzyme locus sequence (label Enzyme is a kind of Ⅱ restriction enzymes, it can in about 20 base position recognition site cutting double-stranded DNA). The structure of the joint is the primer A/B sequence + tag enzyme identification site + anchor enzyme recognition site. (3) A short cDNA fragment (about 9 ~ 10 base) with A connector