Make sure you deleted all old version of dynamo and installed the newest one from github. The version I used to generate this tutorial is 0.0+89e5bb9.dirty (see below). dynamo is still under active development, but it will be submitted to PyPi in one month or so.

In [1]:
from IPython.core.display import display, HTML
display(HTML("<style>.container { width:90% !important; }</style>"))
%matplotlib inline

import dynamo as dyn
dyn.__version__

Out[1]:
'0.0+1ed7a32'

## Dynamo supports a variety of experimental setups¶

Experiment type Estimation method Description (has splicing data) Description (no splicing data) Parameters suitable to estimate
Kinetics:
multiple time point labeling;
lasts several hours
stochastic (moment equations) using first/second moments of $uu, ul, su, sl$ (unspliced unlabeled, unspliced labeled, spliced unlabled, spliced labeled) to estimate $\alpha, \beta, \gamma$ (transcription, splicing and degradataion rate) jointly. using first/second moments of $U (uu + su), L (ul + sl)$ (Unlabeled or labeled) to estimate $\alpha, \gamma$ jointly. $\beta, \gamma$
Kinetics:
multiple time point labeling;
lasts several hours
deterministic (ODEs) 1. use $uu$ over $t$ (time) to estimate $\beta$
2. use the estimated $\beta$ and $ul$ over $t$ to estimate $\alpha$. 3. use the estimated $\beta$ and $uu, su$ over $t$ to estimate $\gamma$
1. use $U$ over $t$ to estimate $\gamma$.
2. use the estimated $\gamma$ and $L$ over $t$ to estimate $\alpha$.
$\beta, \alpha$
one-shot:
single time point labeling;
short time
deterministic (ODEs) a. if $\beta, \gamma$ are already estimated by a degradation experiment, $\alpha$ is estimated by $ul$ over $t$ and $\beta$.
b. if $\beta, \gamma$ are not estimated, use $uu$ over $t$ to solve for $\beta$ first and then the solved $\beta$ and the $uu$ over $t$ to solve for $\alpha$.
a. if $\gamma$ are already estimated by a degradation experiment, $\alpha$ is estimated by $L$ over $t$ and the estimated $\gamma$.
b. if $\gamma$ are not estimated, use $U$ over $t$ to solve for $\gamma$ first and then the solved $\gamma$ and the $L$ over $t$ to solve for $\alpha$.
$\alpha$.
pulse-chase;
can chase more than 1 day
deterministic (ODEs) 1. use $ul$ over $t$ to estimate $\beta$, the estimated $\beta$ and $ul, sl$ over $t$ to estimate $\gamma$.
2. use the estimated $\beta$ and $uu$ over $t$ to estimate $\alpha$.
1. use $L$ over $t$ to estimate $\gamma$
2. use $U$ over $t$ to estimate $\alpha$
$\gamma$
no metabolic labeling:
regular scRNA-seq data
steady state assumption: $\frac{ds}{dt} = \beta u - \gamma s = 0$ assume $\beta$ is 1 and then estimate $\gamma$ by using extreme data points of $u$ (unspliced mRNA) and $s$ (spliced mRNA). None All parameters are relative
cite-seq/reap-seq, etc:
RNA/protein coassay
steady state assumption: $\frac{dp}{dt} = \eta s - \delta s = 0$ assume $\eta$ is 1 and then estimate $\delta$ by using extreme data points of $s$ (spliced mRNA) and $p$ (protein). None All parameters are relative

for details related to moment equations, see the full derivation here: https://github.com/aristoteleo/dynamo-notebooks/blob/master/full_derivation.pdf
for details related to others strategies, see the https://github.com/aristoteleo/dynamo-notebooks/blob/master/velocity_demo/dynamo_fitting_demo.ipynb

We used Gillespie simulation to simulate the cell fate bifurcation. Details can be refered to supplementary figure 1 of our preprint (https://www.biorxiv.org/content/10.1101/696724v1.full).

We first simulate cells to achieve a progenitor steady state. Then we simulate the bifurcation of cell fates into different terminal cell states because of drug treatment or external signals. Next we simulate a metabolic labeling experiment and synthesize kinetics experiment data at each checkpoint. Finally, we stimulate a degradation experiment at the begining or end of the cell differentiation experiment. This setup is typical for a labeling experiment.

In [2]:
# Gillespie simulation of a regular labeling experiment for a cell fate bifurcation process.


Iteration 48/50 finished.

Observation names are not unique. To make them unique, call .obs_names_make_unique.
Observation names are not unique. To make them unique, call .obs_names_make_unique.

Iteration 50/50 finished.

In [3]:
# Now let get the kinetics experiment data at check point t_5, the degradation experiment at the beginning or the end of the differentiation process.


In [4]:
# Let us show Dynamo's fitting for the beginning of a degradation experiment


/home/xqiu/.local/lib/python3.6/site-packages/dynamo/tools/dynamics.py:159: UserWarning: protein layer exists but protein_names is not provided. No estimation will be performed for protein data.
'protein layer exists but protein_names is not provided. No estimation will be performed for protein data.')
/home/xqiu/.local/lib/python3.6/site-packages/scipy/sparse/_index.py:118: SparseEfficiencyWarning: Changing the sparsity structure of a csr_matrix is expensive. lil_matrix is more efficient.
self._set_arrayXarray_sparse(i, j, x)


from above, we can see that the fitting for ul and sl are very good but the fitting for uu and su are not as good. This is because degradation experiment is ideal for estimating $\gamma$ and $\beta$. Note that in real experiments, degradation experiment will chase for up to one day or more, while $\beta$ is often related to just a few minutes so in order to enable accurate $\beta$ estimation, the degradation experiment may need to include a fast labeling period at the beginning, for example including labeling period that separate by just a few minutes or so.

In [5]:
# moment model aims to learn all parameters jointly and also tries to estimate the RNA bursting related parameters, $a, b, \alpha_i, \alpha_a$. See  https://github.com/aristoteleo/dynamo-notebooks/blob/master/full_derivation.pdf.


Trying to set attribute .uns of view, making a copy.

In [6]:
# kinetics: the data for this mode is the same as the moment approach but it uses least sqaure fitting of the determinstic ODE solutions for fitting.


/home/xqiu/.local/lib/python3.6/site-packages/dynamo/tools/velocity.py:909: UserWarning: beta & gamma estimation: only works when there're at least 2 time points.
warn("beta & gamma estimation: only works when there're at least 2 time points.")


From above, we can see that the $uu, ul$ fitting are pretty good. This is because the kinetics experiment is good for estimating parameters $\beta, \alpha$. su fitting looks good here but in general kinetics experiment only involves at most a few hours, the degradation of mRNA can be up to one or two days, so in pratice the fitting su of certain genes in real data may be less optimal. Finally the sl fitting is not great, this is because it is involves all parameters $\alpha, \beta, \gamma$ to fit the data and any uncertainty from those parameters can affect the fitting.

In [7]:
# one_shot. One shot experiment is great for estimating $\alpha$ at different time points, especially if we combined with a degradation experiment and assume constant $\beta, \gamma$ through the entire period.

tmp = dyn.tl.dynamics(adata_dif_kin[adata_dif_kin.obs.time == 0.8, :], experiment_type='one_shot', filter_gene_mode='no', tkey='time')

/home/xqiu/.local/lib/python3.6/site-packages/scipy/sparse/_index.py:118: SparseEfficiencyWarning: Changing the sparsity structure of a csr_matrix is expensive. lil_matrix is more efficient.
self._set_arrayXarray_sparse(i, j, x)


# the following are similar analysis if we don't have splicing data¶

This is normally true because spliting U to uu, su and L to ul, sl may be difficult especially for some transcription which already have low expression. The discussion above will be generally applicable below but just avoid the consideration of splicing.

In [8]:
adata_dif_kin_nosplicing, adata_dif_deg_begin_nosplicing, adata_dif_deg_end_nosplicing = adata_dif_nosplicing[adata_dif_nosplicing.obs.experiment_type == 'kin_t_5', :], \

In [9]:
adata_dif_deg_begin_nosplicing

Out[9]:
View of AnnData object with n_obs Ã— n_vars = 250 Ã— 2
obs: 'trajectory', 'time', 'experiment_type'
obsm: 'protein'
layers: 'new', 'total'
In [10]:
# Degradation
import dynamo as dyn

/home/xqiu/.local/lib/python3.6/site-packages/dynamo/tools/dynamics.py:159: UserWarning: protein layer exists but protein_names is not provided. No estimation will be performed for protein data.
'protein layer exists but protein_names is not provided. No estimation will be performed for protein data.')
Trying to set attribute .var of view, making a copy.

In [11]:
# moment

Trying to set attribute .uns of view, making a copy.

In [12]:
# kinetics

# one_shot