Linear amplification scheme for genomic DNA. Step 1: Double stranded DNA starting material (shown with one strand in black and one strand in blue) is tailed on the 3' end of each strand to generate a 20–40 bp polyT tail with a terminal dideoxycytidine base. Step 2a: A T7-(A)18B anchored primer adaptor is annealed to the polyT tail of each template strand. Step 2b: During second strand synthesis Klenow fragment of DNA Polymerase I removes the excess bases from the tail overhang via its 3'-5' exonuclease activity, and extends from the primer to produce the second strand. This results in two double stranded DNAs identical to the original template, except that each has a T7 promoter at a different end. Step 3: The product of second strand synthesis is used as template in an in vitro transcription reaction. Step 4: To generate DNA probes for microarray analysis, amplified RNA is reverse transcribed.
Liu et al. BMC Genomics 2003 4:19 doi:10.1186/1471-2164-4-19