Jun 12, · A team led by researchers from Broad Institute of MIT and Harvard, and the McGovern Institute for Brain Research at MIT, has characterized and engineered a new gene-editing system that can precisely and efficiently insert large DNA sequences into a genome. The system, harnessed from cyanobacteria and called CRISPR-associated transposase (CAST), May 17, · — Debate rages over whether SARS-CoV-2 can insert bits of its RNA into the human genome by Veronica Hackethal, MD, MSc, Enterprise & Investigative Writer, MedPage Today May 17, Insertion of multicopies of the particular gene of interest has become one very common approach in this field. This is usually done by two methods. Firstly, it can be done by inserting or integrating the multiple copies of the desired gene sequence at one or more locations in the chromosomes of the transformant
Four Ways to Insert Foreign DNA Into Cells | Education - Seattle PI
Respected experts in genetics have caused a stir on both social media and in scientific circles by suggesting that SARS-CoV-2 can integrate pieces of its genetic code into the human genome. If true, the results could explain why some people test positive for SARS-CoV-2 RNA sometimes months after recovering from their initial infection, even though no infectious virus can usually be grown.
But many scientists say they aren't convinced, citing concerns that the findings could simply be artifacts of lab preparation. Some have also raised concerns that the work could be misconstrued by the anti-vaccine community, which is reportedly stretching the results to apply to COVID vaccines -- even though the authors have stressed the work has no implications for the vaccines.
They also emphasized that there are no implications for human health at this time. The controversy began in Decemberwhen Rudolf Jaenisch, MD, inserting gene into genome founding member of the Whitehead Institute for Biomedical Research at MIT, and colleagues published a preprint on bioRxiv suggesting that SARS-CoV-2 RNA could be reverse transcribed and integrated into the human genome.
The results, which weren't peer-reviewed, set off a storm on ScienceTwitter. They were panned on an episode of This Week in Virology TWiVand garnered harsh criticism from commenters who said they could be used to bolster false claims about vaccines. This is now being used by some to spread doubts about the new vaccines.
Since then, at least two more preprints on the issue have been published on bioRxivin which researchers say they attempted to replicate inserting gene into genome results but could not.
The crux of their argument is that the results relied on published libraries of RNA sequencing data from SARS-CoV-2 infected inserting gene into genome cultures and organoids of the lung, heart, brain, and stomach, as well as COVID patient-derived cells.
Preparation of these RNA-sequencing libraries is error-prone and can also create chimeras. So the SARS-CoV-2 human chimeric transcripts could be lab artifacts created during the preparation of these libraries, not from natural infection, they argue. Jaenisch acknowledged to MedPage Today that "there were holes in the argument," that the evidence inserting gene into genome not complete," and that "there was reason for criticizing this paper. He added that the unnamed journal where he was seeking publication required coronavirus papers to be posted on the bioRxiv server upon submission.
I didn't want to do it, but I was sort of forced. We were under enormous pressure to make it known. Jaenisch and team subsequently performed additional analyses, which they published last month in PNASprompting another Twitter storm. The PNAS paper describing this 'finding' should never have been published as it is biologically irrelevant," tweeted Vincent Racaniello, PhDof Columbia University, and host of the TWiV podcast. Still, inserting gene into genome, Jaenisch contends that the newest paper provides evidence that confirms "totally without doubt" two key results: small fragments of SARS-CoV-2 can integrate into the human genome, and these integrated viral sequences can be detected by PCR.
No one can doubt this, based on seeing DNA copies of the virus in the genome of infected cells. And, these sequences in patients have to come from integrated sequences, based on the structure that you inserting gene into genome expect," he told MedPage Today.
Although they acknowledge that they cannot rule out artifacts, Jaenisch and colleagues addressed the issue by looking at the orientation of integrated viral sequences compared to human genes. They also found that the integrated viral sequences were flanked by genetic elements characteristic of long interspersed nuclear elements-1 LINE LINE-1 elements, also called selfish elements, can pop around the human genome and produce reverse transcriptase when activated.
Unlike retroviruses like HIV, SARS-CoV-2 does not carry a reverse transcriptase enzyme that helps it transcribe back into DNA, but these LINE-1 elements could be doing the job, according to Jaenisch.
Taken together, he says these results provide proof that the mechanism of integration involves these LINE-1 elements, inserting gene into genome. Viral integration has been described in many other viral infectionssuch as hepatitis C, influenza, inserting gene into genome, and measles, and has been explored for its role in oncogenesis as with the human papillomavirus.
But this study would be the first to show that a coronavirus can integrate into the human genome. One of the preprints that argued against this idea was just published in the peer-reviewed Journal of Virology. The group of researchers from Purdue University, the University of Michigan, and the NIH, maintains that these viral-human chimeric transcripts are laboratory artifacts created during RNA sequencing.
Cedric Feschotte, PhD, of Cornell University, an original critic of the work, also remains skeptical and says that more research is needed to prove viral integration. However, he now acknowledges that the hypothesis is plausible. But the "greatest weakness" lies in not quantifying the frequency of these reported SARS-CoV-2 integration events. The LINE-1 mechanism has already been found to be involved in integration for other viruses.
Scientists who are familiar with this process would expect that this could be the case, he added. The major question is the frequency: how much and how often. This study leaves us without a very clear picture of the frequency. It doesn't tell me much that I didn't know before," he said. On the other hand, Nobel laureate David Baltimore, PhD, inserting gene into genome, a professor emeritus at Caltech, said that the results "look pretty convincing.
There are always people who hold out against a new finding, and we have to live with that," Baltimore told MedPage Today. That may happen with the common cold or whatever other viruses infect us. While more researchers have come around to the updated results, the debate will continue to evolve, inserting gene into genome, and the scientific community will be watching closely.
Veronica Hackethal reports for MedPage Today's Enterprise and Investigative journalism team. share to facebook. share to twitter. share to linkedin. email article.
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, time: 6:30Can COVID Genes Integrate Into Human DNA? | MedPage Today
Adenovirus (Ad) vectors arguably represent the most efficient delivery systems of episomal DNA into eukaryotic cell nuclei. The most advanced recombinant Ads lack all adenoviral genes. This renders these so-called high-capacity (hc) Ad vectors less cytotoxic/immunogenic than those only deleted in early regions and creates space for the insertion of large/multiple blogger.com by: 21 Gene insertion is the addition of one or more genes into a DNA sequence, a technique that has been traditionally performed with plasmid DNA or integrating viral vectors. In the conventional gene insertion method, the insertion site cannot be controlled Sticking the vector and the gene together. The final step in cloning is to incorporate the DNA of interest into the vector. Scientists mix the gene and the opened vector together with a bacterial enzyme called DNA ligase. The ligase sticks DNA ends together to form a single circular molecule that includes both the vector and the gene
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