Evaluation of methods for amplification of picogram amounts of total RNA for whole genome expression profiling

BMC Genomics

Table 2 Comparison of hybridisation quality metrics of the evaluated RNA inputs and protocols.

Protocol	RNA input	Background	Present Call	β-Actin 3'/5' ratio	GAPDH 3'/5' ratio
Ambion	500 pg	58.4	42%	139.1	11.2
		32.0	48%	235.5	6.5
	250 pg	58.4	38%	215.6	13.5
		32.4	44%	123.3	7.6
Arcturus	500 pg	54.0	29%	96.3	25.2
		52.4	30%	70.5	15.0
	250 pg	32.7	37%	36.5	14.2
		43.9	41%	33.3	9.4
Epicentre	500 pg	58.4	43%	30.9	11.4
		-	-	-	-
	250 pg	-	-	-	-
		-	-	-	-
Nugen	500 pg	33.5	45%	6.6	1.1
		35.7	49%	7.2	1.2
	250 pg	31.5	39%	6.9	0.8
		32.8	45%	10.6	1.3
Affymetrix, One-round amplification protocol	2 μg	44.9 44.7	50% 49%	1.5 2.6	1.0 1.2
Affymetrix, Two-rounds amplification protocol	100 ng	66.7 54.7	52% 47%	12.2 12.8	1.3 1.8

'-': no hybridisation performed due to poor yield after amplification.
Targets were synthesised, in each laboratory (see Table 1), with the five protocols and hybridised to HG U133 Plus 2.0 array. A rapid analysis was performed with Expression Console (Affymetrix, MAS 5 normalisation).

ISSN: 1471-2164