SAGE analysis from 1 microg of total RNA.
Serial analysis of gene expression (SAGE) is a powerful transcription-profiling method that allows simultaneous expression analysis of thousands of transcripts, provides absolute digital readout of the expression level, and identifies new genes. A disadvantage of SAGE is the relatively high amount of input RNA required. Consequently, several techniques have been developed to overcome this limitation, so that SAGE can be applied to very limited amounts of starting material, e.g., small biological samples such as tissue biopsies or microdissected materials. Here we describe a modified version of the original microSAGE protocol, which requires only 1 to 2 microg total RNA. This method avoids PCR amplification of cDNA and reamplification of ditags, procedures that potentially compromise the quantitative nature of this technique.
Cai
Jerry
J
Johns Hopkins University, Baltimore, Maryland, USA.
Ash
David
D
Jabs
Ethylin Wang
EW
D000818:Animals; D018076:DNA, Complementary / Q000096:biosynthesis*; D012321:DNA-Directed RNA Polymerases / Q000737:chemistry*; D015723:Gene Library; D006801:Humans; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D012333:RNA, Messenger / Q000032:analysis* / Q000302:isolation & purification; D012680:Sensitivity and Specificity; D014764:Viral Proteins / Q000737:chemistry*
D018076:DNA, Complementary; D012333:RNA, Messenger; D014764:Viral Proteins; C062210:bacteriophage T7 RNA polymerase; D012321:DNA-Directed RNA Polymerases