EST and SAGE
üExpressed sequence tag (EST) sequencing are generated by randomly
picking clones from a cDNA
library and performing a single sequencing reaction to produce 300 to 500 bps
of sequence per clone. Differences in gene expression may be identified by
counting the number of times a particular sequence appears in EST libraries of
genes from different sources.
üSerial analysis of gene expression (SAGE) a technique for the analysis of gene expression that is essentially an accelerated version of EST sequencing.
SAGE: 10-14 bps tags created by type
IIS restriction endonuclease
Nla III.
Super SAGE: > 25 bps tag, created by type
III restriction enzyme
EcoP15I.
Advantages:
ØSAGE data are quantitative and
cumulative.
ØAccurate, quantitative transcript profiles describing the abundance of all genes expressed in a cell or tissue are generated by SAGE, if sufficient sequencing
is completed.