Pfizer and Moderna bivalent vaccines contain 20-35% expression vector and are transformation competent in E.coli

"Conclusions

Previous RNA-Seq based estimates of the double stranded DNA contamination in the vaccines significantly under reported the magnitude of the contamination. Using qPCR and electrophoresis, we demonstrate the dsDNA contamination levels are 100 fold higher and imply trillions of DNA molecules per dose. The Pfizer DNA contamination ranges from 8.19-11.3 ng/ul with 23-28ng/ul of mRNA. The Moderna DNA contamination ranges from 7.5 - 9.5ng/ul with 25.7ng/ul - 55.9ng/ul of mRNA. This averages to 9.1ng/ul mean DNA concentration versus 33.4ng/ul mean RNA concentration.

This equates to 27.3% (9.1/33.4) of the nucleic acid in each vaccine being expression vector. This is several orders of magnitude over the the EMAs limit of 330ng/mg.

An unknown portion of these dsDNA contaminants are replication competent plasmids that can transform E.coli with a simple 20 second 42C heat shock treatment. These plasmids provide antibiotic resistance on LB-Kan plates and can be isolated from E.coli cultures. It is unlikely these plasmids will express spike protein in non-laboratory modified E.coli as the ribosomal signals in the vaccine mRNA are designed for mammalian translation. The T7 promoter is known to leak in mammalian cell lines and some laboratory E.coli genotypes but is not expected to leak in wild type E.coli. This may enable mRNA to be expressed from these plasmids in mammalian cells but unless the plasmids are integrated into the human genome, they are unlikely to be replicated to high copy number.

While bacteria are unlikely to express this spike protein, bacteria can replicate this plasmid and serve as a bactofection source for introduction of these mammalian expression plasmids to human cells.

Given the near equimolar contamination, studies evaluating the reverse transcriptase capacity of LINE-1 should be reconsidered. If each injection provides trillions of dsDNA contaminants, LINE-1 RT activity is not a necessary step for genome integration. The critiques of Alden et al focused primarily on the fact that LINE-1 is predominantly expressed in cancer cells lines and that the LINE-1 observation shouldn’t be extrapolated to patients. The vaccines are providing trillions of dsDNAs containing a potentially leaky T7 promoter encoding a spike protein with a Kozak consensus sequence. With these levels of contamination, RT activity from LINE-1 is not a prerequisite for genome integration. These data cannot inform on genome integration and further IRB reviewed deep sequencing work is required to address rare mosaic integration events in patients. Regardless of these hypothetical concerns, the dsDNA contamination exceeds the EMA specifications by several orders of magnitude and further scrutiny should be applied to the endotoxin levels and dsRNA levels in these vaccines."