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Biomedical subjects

Gabriele Schratzberger

Publications and source records attributed to Gabriele Schratzberger.

5 recordsLinked to original sources
PubMed2004

Reliability of t7-based mRNA linear amplification validated by gene expression analysis of human kidney cells using cDNA microarrays.

Genome wide gene expression analysis by cDNA microarrays is often limited by minute amounts of starting RNA. We therefore tested an optimized linear RNA amplification protocol using the RiboAmp amplification kit in the setting of cDNA microarrays. We isolated mRNA from a human kidney cell line (HK-2; ATCC) and from Universal Human Reference RNA (STR; Stratagene). After performing one and two rounds of linear RNA amplification, respectively, the amplified RNAs were co-hybridized to cDNA microarrays. Linearity and reproducibility of the individual experiments were then assessed by calculating the Pearson correlation. The intra-amplification consistency showed a correlation of 0.968 for the first round, 0.907 for the second round and 0.912 for two successive rounds of amplification. If the first round was compared to unamplified material, r was 0.925. The second round amplification yielded a correlation of 0.897 if compared to unamplified mRNA. Two rounds of amplification starting from 200 pg of mRNA compared to unamplified material resulted in a correlation of 0.868. These results indicate that linear amplification using RiboAmp kit yields amplified RNA with a high degree of linearity and reproducibility.

Cell Line↗
PubMed2002

Transcutaneous ultrasound augments naked DNA transfection of skeletal muscle.

This study was designed to test the hypothesis that transcutaneous ultrasound (US) exposure may augment the transfection efficiency and biological outcome associated with nonviral DNA gene transfer. Hindlimb muscles of New Zealand White rabbits were transfected with the reporter plasmid pCMV-beta, with or without US exposure. Optimization studies employed US exposure at various frequencies, mechanical indices, duty cycles, durations of exposure, and exposure time points. Based on these results, we explored the effect of US exposure on nonviral gene transfer of vascular endothelial growth factor (VEGF, phVEGF165) to promote neovascularization of ischemic hindlimbs. Ultrasound at 1 MHz, 100 W/cm(2), 6% duty cycle, and 5 minutes exposure time, applied immediately following DNA injection, was found to be the most effective among the settings tested, increasing beta-galactosidase expression approximately 20 fold. Compared with US exposure alone, or phVEGF165 only, phVEGF165 + US exposure yielded a statistically significant improvement in revascularization, as determined by calf blood pressure ratio, angiographic score, intravascular Doppler blood flow, and capillary/myocyte ratio. These data demonstrate that ultrasound, when applied directly after intramuscular gene transfer, significantly increases transfection efficiency in vivo. The biological significance of this finding was confirmed by augmented limb perfusion in response to US exposure and naked VEGF DNA.

Animals↗