4 ms·

They have a response paper though : "Here we provide additional confirmatory data and clarifying discussion, including sequencing data showing extensive hetero

They have a response paper though : "Here we provide additional confirmatory data and clarifying discussion, including sequencing data showing extensive heterozygous mutations throughout the genome in the CRISPR treated mice, which are all progeny of inbred mice purchased from a commercial vendor (JAX). The heterozygosity in these cases cannot be parentally inherited. The summary statements in our Correspondence reflect observations of a secondary outcome following successful achievement of the primary outcome using CRISPR to treat blindness in Pde6b/rd1 mice. As the scientific community considers the role of WGS in off-target analysis, future in vivo studies are needed where the design and primary outcome focuses on CRISPR off-targeting. We agree that a range of WGS controls are needed that include parents, different gRNAs, different versions of Cas9, and different in vivo protocols. We look forward to the publication of such studies. Combined, these results will be essential to fully understand off-targeting and can be used to create better algorithms for off-target prediction. Overall, we are optimistic that some form of CRISPR therapy will be successfully engineered to treat blindness." http://www.biorxiv.org/content/early/2017/06/23/154450 http://www.biorxiv.org/content/early/2017/06/23/154450

nonbel 9y ago
Thanks, I was looking for this info earlier (https://news.ycombinator.com/item?id=14448371 https://news.ycombinator.com/item?id=14448371): >"In our case, 56 zygotes were harvested from six pregnant females bred to six stud males and injected with CRISPRCas9." http://www.biorxiv.org/content/early/2017/06/23/154450 http://www.biorxiv.org/content/early/2017/06/23/154450 So they started with 56 zygotes, of which 11 survived injection of Crispr/etc. Of these 11, five and two mice showed some evidence of containing NHEJ/HDR cells, respectively. In the case of NHEJ, they do not tell us what percent of cells were "edited" per mouse (or I missed it when I read that paper earlier). However for HDR it was only about 1/3 and 1/5 for the two mice they found. So this suggests the "editing" is occurring not at the zygote stage, but somewhat later (based on the percentages, perhaps at the 4 or 8 cell stages). Assuming the edit/mouse rate is similar for NHEJ and HDR (somewhat dubious), it looks like 4x56 = 224 to 8x56 = 448 cells were treated with crispr/etc and ~ 7 got "edited". This gives ~1-3% "edits" per treated cell.

nonbel 9y ago

Thanks, I was looking for this info earlier (https://news.ycombinator.com/item?id=14448371 https://news.ycombinator.com/item?id=14448371): >"In our case, 56 zygotes were harvested from six pregnant females bred to six stud males and injected with CRISPRCas9." http://www.biorxiv.org/content/early/2017/06/23/154450 http://www.biorxiv.org/content/early/2017/06/23/154450 So they started with 56 zygotes, of which 11 survived injection of Crispr/etc. Of these 11, five and two mice showed some evidence of containing NHEJ/HDR cells, respectively. In the case of NHEJ, they do not tell us what percent of cells were "edited" per mouse (or I missed it when I read that paper earlier). However for HDR it was only about 1/3 and 1/5 for the two mice they found. So this suggests the "editing" is occurring not at the zygote stage, but somewhat later (based on the percentages, perhaps at the 4 or 8 cell stages). Assuming the edit/mouse rate is similar for NHEJ and HDR (somewhat dubious), it looks like 4x56 = 224 to 8x56 = 448 cells were treated with crispr/etc and ~ 7 got "edited". This gives ~1-3% "edits" per treated cell.