plasmid_concerns.md 1.1 KB
He is quoted to have said "without the translocation of the plasmid DNA into the nucleus, no gene expression or gene therapy can take place" (David Dean).
What this means is that there was no possibility for informed consent.
DNA contamination origin Two processes for manufacturing modified mRNA.
- Produced using PCR (used in clinical trials - 20k + unblinding)
- Upscaling method
- E coli-based system
Concerns
- DNA contamination
- Lipopolysaccharides (by product of using e coli system) At end of 5 step workflow, the product must be cleaned.
What happened?
- Hybrids of mRNA and DNA?
- Perhaps dnase couldn't completely break down unwanted components
- Lipo polysaccharide endotoxin
Questions
Can contaminant DNA get into the nucleus of transfected cells?
- Lipid Nanoparticles
- Trojan horses
Nucleus
SV40 promoter/enhancer
- Antibiotic resistance genes
- Strong promoters used in biotech
- Enhancers not necessary for transcription to ensue
- Enhancer's presence indicates it was put there to optimize transcription
- Why is enhancer there? Nuclear Location sequence (Fedex vehicle bringing things to nucleus).