4 ms·
> "It doesn't appear (to me) that selective lethality would account for this data." Please go into more detail about what selective lethality is not explaining
by nonbel 9y ago
> "It doesn't appear (to me) that selective lethality would account for this data."
Please go into more detail about what selective lethality is not explaining. Under that explanation
1) They started with ~35 wild type and ~35 mutant.
2) The treatment killed about half the mutants (but no/few wt) via inducing toxic double stranded breaks.
3) They end up with the original ~35 wt plus ~16 mutants, thus increasing the proportion of wt.
Regarding heterozygosity:
4) In some of the surviving mutants the cas9 damaged the DNA during the 2-cell phase or later, thus knocking out the mutant allele in some cells but not others.
As I said, I do not see what is difficult to understand. It seems to me guaranteed that a process like this accounts for at least some of the "editing" (ie, it is impossible to not be happening at all). Really, it is only a matter of what percent of the results are due to this.
- pcrh 9y agoIt isn't clear to me where you're getting these numbers from... where do you find 35 WT and 35 mutant? Could you point me to the data for WT vs mutant embryos in this experiment? Edit: in case you're referring to the table you linked to, the top line is unmanipulated embryos, and the second and third to experimental embryos, control and test. The survival rate of the second two lines is reported as similar.
- nonbel 9y agoThe man who donated the sperm was heterozygous, which would mean about half the sperm would contain the mutant. You can also see this in their figure 2: >"Sequencing of 83 individual blastomeres collected from 19 control embryos revealed that 9 (47.4%) were homozygous wild type (MYBPC3WT/WT) and 10 (52.6%) were heterozygous, carrying the wild-type maternal and mutant paternal alleles (MYBPC3WT/ΔGAGT) (Fig. 2a and Supplementary Table 2). This distribution was expected and confirms that the heterozygous patient sperm sample contained equal numbers of wild-type and mutant spermatozoa with similar motility and fertilization efficiency."
- nonbel 9y ago>"Edit: in case you're referring to the table you linked to, the top line is unmanipulated embryos, and the second and third to experimental embryos, control and test. The survival rate of the second two lines is reported as similar." Also, not completely correct. The second line is not controls. Those are the 70 - 16 = 54 cells for which data is shown in figure 2A. The third line is for additional experiments which I didn't really look at.