Prevalence of errors in lab-made plasmids across the globe 9243

Abstract Description Plasmids are an indispensable reagent in life sciences research and therapeutics development. Currently, most labs custom-build plasmids in a “do it yourself” (DIY) fashion at the bench. As yet, no systematic data on the quality of DIY plasmids exists. Here, we report a broad survey of plasmids made by hundreds of labs worldwide from both academia and industry. We found that nearly half of them contained design and/or sequence errors that could potentially compromise function. For adeno-associated virus (AAV) vectors, which are widely used in gene therapy, we found that about 40% of transfer plasmids used to package AAV carried mutations in the inverted terminal repeat (ITR) regions relative to the wildtype due to their inherent sequence instability, and such instability is strongly influenced by flanking GC content. We also list genes that are difficult to clone into plasmid or package into virus due to their toxicity to host cells. Our findings raise serious concerns over the trustworthiness of a ubiquitous research reagent, which parallels the widespread mycoplasma contamination and misidentified mammalian cell lines reported previously, and highlights the need for community-wide standards to uphold the quality of gene delivery vectors in research and medicine. Topic Categories Technological Innovations in Immunology (TECH)