Criteria for effective design, construction, and gene knockdown by shRNA vectors

doi:10.1186/1472-6750-6-7

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BMC Biotechnol. 2006; 6: 7.

Published online 2006 Jan 24. doi: 10.1186/1472-6750-6-7

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Table 2

Evaluation of sequencing results of three DNA hairpin constructs. Average ratio of peak height after to before the hairpin region was determined as a measure of how well the sequence read through the hairpin structure. The greater the peak height ratio, the greater the ability to sequence through the hairpin. A value of 1 indicates no loss in peak height, and a value of zero indicates a complete stop in sequence after the hairpin region. All values are the averages of at least triplicate sequencing reactions.

		DNA Plasmid
		pHSPG-shTLR4	pHSPG-shmutTLR4	pHSPG-shmCNN3

Chemistry	DNA relaxing agent	Peak Height Ratio	Peak Height Ratio	Peak Height Ratio

BDv1.1	None	0.0	0.4	0.0
BDv1.1	5% DMSO	0.0	0.6	0.2
BDv1.1	0.83 M Betaine	0.3	0.9	0.7
BDv1.1	1 × PCRx Enhancer	0.1	0.7	0.3
BDv1.1	0.83 M Betaine & 1 × PCRx Enhancer	0.6	0.9	0.5
20:1 BD:dGTP	None	0.4	0.6	0.5
20:1 BD:dGTP	5% DMSO	0.6	0.7	0.6
20:1 BD:dGTP	0.83 M Betaine	0.7	1.0	0.8
20:1 BD:dGTP	1 × PCRx Enhancer	0.6	0.8	0.7
20:1 BD:dGTP	0.83 M Betaine & 1 × PCRx Enhancer	0.8	0.9	0.8
10:1 BD:dGTP	None	0.5	0.6	0.6
10:1 BD:dGTP	5% DMSO	0.7	0.8	0.7
10:1 BD:dGTP	0.83 M Betaine	0.8	0.9	0.9
10:1 BD:dGTP	1 × PCRx Enhancer	0.7	0.8	0.8
10:1 BD:dGTP	0.83 M Betaine & 1 × PCRx Enhancer	0.9	1.0	0.9
10:1 BD:dGTP	1 × ThermoFidelase I	0.1	0.2	0.2
5:1 BD:dGTP	None	0.6	0.7	0.7
3:1 BD:dGTP	None	0.6	0.6	0.6
dGTP only	None	0.7	0.7	0.8