We demonstrate how to simulate NGS data under various genotype distributions, then fit these data using `flexdog`

. The genotyping methods are described in Gerard et al. (2018).

Let’s suppose that we have 100 hexaploid individuals, with varying levels of read-depth.

```
set.seed(1)
library(updog)
<- 100
nind <- 6
ploidy <- round(stats::runif(n = nind,
sizevec min = 50,
max = 200))
```

We can simulate their read-counts under various genotype distributions, allele biases, overdispersions, and sequencing error rates using the `rgeno`

and `rflexdog`

functions.

Suppose these individuals are all siblings where the first parent has 4 copies of the reference allele and the second parent has 5 copies of the reference allele. Then the following code, using `rgeno`

, will simulate the individuals’ genotypes.

```
<- rgeno(n = nind,
true_geno ploidy = ploidy,
model = "f1",
p1geno = 4,
p2geno = 5)
```

Once we have their genotypes, we can simulate their read-counts using `rflexdog`

. Let’s suppose that there is a moderate level of allelic bias (0.7) and a small level of overdispersion (0.005). Generally, in the real data that I’ve seen, the bias will range between 0.5 and 2 and the overdispersion will range between 0 and 0.02, with only a few extremely overdispersed SNPs above 0.02.

```
<- rflexdog(sizevec = sizevec,
refvec geno = true_geno,
ploidy = ploidy,
seq = 0.001,
bias = 0.7,
od = 0.005)
```

When we plot the data, it looks realistic

```
plot_geno(refvec = refvec,
sizevec = sizevec,
ploidy = ploidy,
bias = 0.7,
seq = 0.001,
geno = true_geno)
```

We can test `flexdog`

on these data

```
<- flexdog(refvec = refvec,
fout sizevec = sizevec,
ploidy = ploidy,
model = "f1")
#> Fit: 1 of 5
#> Initial Bias: 0.3678794
#> Log-Likelihood: -363.9369
#> Keeping new fit.
#>
#> Fit: 2 of 5
#> Initial Bias: 0.6065307
#> Log-Likelihood: -363.937
#> Keeping old fit.
#>
#> Fit: 3 of 5
#> Initial Bias: 1
#> Log-Likelihood: -363.9369
#> Keeping new fit.
#>
#> Fit: 4 of 5
#> Initial Bias: 1.648721
#> Log-Likelihood: -381.6123
#> Keeping old fit.
#>
#> Fit: 5 of 5
#> Initial Bias: 2.718282
#> Log-Likelihood: -412.8604
#> Keeping old fit.
#>
#> Done!
```

`flexdog`

gives us reasonable genotyping, and it accurately estimates the proportion of individuals mis-genotyped.

`plot(fout)`

```
## Estimated proportion misgenotyped
$prop_mis
fout#> [1] 0.07011089
## Actual proportion misgenotyped
mean(fout$geno != true_geno)
#> [1] 0.05
```

Now run the same simulations assuming the individuals are in Hardy-Weinberg population with an allele frequency of 0.75.

```
<- rgeno(n = nind,
true_geno ploidy = ploidy,
model = "hw",
allele_freq = 0.75)
<- rflexdog(sizevec = sizevec,
refvec geno = true_geno,
ploidy = ploidy,
seq = 0.001,
bias = 0.7,
od = 0.005)
<- flexdog(refvec = refvec,
fout sizevec = sizevec,
ploidy = ploidy,
model = "hw")
#> Fit: 1 of 5
#> Initial Bias: 0.3678794
#> Log-Likelihood: -377.9226
#> Keeping new fit.
#>
#> Fit: 2 of 5
#> Initial Bias: 0.6065307
#> Log-Likelihood: -377.9226
#> Keeping old fit.
#>
#> Fit: 3 of 5
#> Initial Bias: 1
#> Log-Likelihood: -377.9226
#> Keeping old fit.
#>
#> Fit: 4 of 5
#> Initial Bias: 1.648721
#> Log-Likelihood: -377.9226
#> Keeping new fit.
#>
#> Fit: 5 of 5
#> Initial Bias: 2.718282
#> Log-Likelihood: -377.9226
#> Keeping old fit.
#>
#> Done!
plot(fout)
```

```
## Estimated proportion misgenotyped
$prop_mis
fout#> [1] 0.07625987
## Actual proportion misgenotyped
mean(fout$geno != true_geno)
#> [1] 0.07
## Estimated allele frequency close to true allele frequency
$par$alpha
fout#> [1] 0.7473264
```

Gerard, David, Luís Felipe Ventorim Ferrão, Antonio Augusto Franco Garcia, and Matthew Stephens. 2018. “Genotyping Polyploids from Messy Sequencing Data.” *Genetics* 210 (3). Genetics: 789–807. https://doi.org/10.1534/genetics.118.301468.