Based on the Cleave-and-Rescue system of Oberhofer et. al. (doi:10.1101/2020.07.09.196253
),
this is a 3-locus Cas9-based toxin-antidote system. The first locus carries the
Cas9, the second locus carries the gRNAs, and a recoded copy of an essential gene.
The third locus is the targeted essential gene. This gene can be completely
haplosufficient (hSuf = 1) or completely haploinsufficient (hSuf = 0).
It is assumed that having 2 copies of the gene (be it wild-type at the second
locus or recoded at the first) confers complete viability. Additionally, loci
1 and 2 can be linked, given crM and crF, imitating the original
2-locus ClvR system.
For this construct, the first locus will have 2 alleles, the second will have 2
alleles, and the third will have 3 alleles:
Locus 1
W: Wild-type
C: Cas9
Locus 2
W: Wild-type
G: gRNAs and recoded essential gene
Locus 3
W: Wild-type
R: Functional resistant
B: Non-functional resistant
Usage
cubeClvR2(
cF = 1,
crF = 0,
ccF = cF,
ccrF = crF,
cM = 1,
crM = 0,
ccM = cM,
ccrM = crM,
dW = 0,
drW = 0,
ddW = dW,
ddrW = drW,
hSuf = 1,
crossF = 0.5,
crossM = 0.5,
eta = NULL,
phi = NULL,
omega = NULL,
xiF = NULL,
xiM = NULL,
s = NULL
)Arguments
- cF
Female cutting rate, one ClvR allele
- crF
Female functional resistance rate, one ClvR allele
- ccF
Female cutting rate, two ClvR alleles
- ccrF
Female functional resistance rate, two ClvR alleles
- cM
Male cutting rate, one ClvR allele
- crM
Male functional resistance rate, one ClvR allele
- ccM
Male cutting rate, two ClvR alleles
- ccrM
Male functional resistance rate, two ClvR alleles
- dW
Female deposition cutting rate
- drW
Female deposition functional resistance rate
- ddW
Female deposition (HH) cutting rate
- ddrW
Female deposition (HH) functional resistance rate
- hSuf
Haplosufficiency level, default is completely sufficient
- crossF
Crossover rate in females, 0 is completely linked, 0.5 is unlinked, 1.0 is complete divergence
- crossM
Crossover rate in males, 0 is completely linked, 0.5 is unlinked, 1.0 is complete divergence
- eta
Genotype-specific mating fitness
- phi
Genotype-specific sex ratio at emergence
- omega
Genotype-specific multiplicative modifier of adult mortality
- xiF
Genotype-specific female pupatory success
- xiM
Genotype-specific male pupatory success
- s
Genotype-specific fractional reduction(increase) in fertility
Value
Named list containing the inheritance cube, transition matrix, genotypes, wild-type allele, and all genotype-specific parameters.
Details
Female deposition is implemented incorrectly. Right now, it is performed on
male alleles prior to zygote formation - it should happen post-zygote formation.
Since this construct doesn't have HDR, this should be fine.
Additionally, it is assumed that deposition requries loaded Cas9-RNP complexes
from the mother, having Cas9 and no maternal gRNA, even in the presence of
paternal gRNA, will not result in maternal deposition mediated cleavage.
Copy-number dependent rates are based on Cas9, not gRNA. The assumption is that RNA is easier to produce, and therefore won't limit cleavage by Cas9.