SIMBA on 1.3 million cells
[ ]:
[ ]:
[1]:
import os
import simba as si
si.__version__
[1]:
'1.1'
[3]:
workdir = 'result_simba_rnaseq'
si.settings.set_workdir(workdir)
Saving results in: result_simba_rnaseq
[4]:
si.settings.set_figure_params(dpi=80,
style='white',
fig_size=[5,5],
rc={'image.cmap': 'viridis'})
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[5]:
adata_CG = si.read_10x_h5('./data/1M_neurons_filtered_gene_bc_matrices_h5.h5')
Variable names are not unique. To make them unique, call `.var_names_make_unique`.
[6]:
adata_CG
[6]:
AnnData object with n_obs × n_vars = 1306127 × 27998
var: 'gene_ids'
[8]:
adata_CG.var_names_make_unique()
[9]:
df_metadata = pd.read_csv('./data/analysis/clustering/graphclust/clusters.csv', index_col=0)
df_metadata.index.name = None
df_metadata['Cluster'] = 'cluster_' + df_metadata['Cluster'].astype(str)
print(df_metadata.shape)
df_metadata.head()
(1306127, 1)
[9]:
| Cluster | |
|---|---|
| AAACCTGAGATAGGAG-1 | cluster_14 |
| AAACCTGAGCGGCTTC-1 | cluster_14 |
| AAACCTGAGGAATCGC-1 | cluster_37 |
| AAACCTGAGGACACCA-1 | cluster_4 |
| AAACCTGAGGCCCGTT-1 | cluster_20 |
[11]:
adata_CG.obs = df_metadata
[12]:
adata_CG
[12]:
AnnData object with n_obs × n_vars = 1306127 × 27998
obs: 'Cluster'
var: 'gene_ids'
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[14]:
si.pp.filter_genes(adata_CG,min_n_cells=3)
Before filtering:
1306127 cells, 27998 genes
Filter genes based on min_n_cells
After filtering out low-expressed genes:
1306127 cells, 22788 genes
[15]:
si.pp.cal_qc_rna(adata_CG)
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si.pl.violin(adata_CG,list_obs=['n_counts','n_genes','pct_mt'])
[17]:
si.pp.normalize(adata_CG,method='lib_size')
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si.pp.log_transform(adata_CG)
[19]:
si.pp.select_variable_genes(adata_CG, n_top_genes=2000)
si.pl.variable_genes(adata_CG,show_texts=True)
2000 variable genes are selected.
[20]:
si.tl.discretize(adata_CG,n_bins=5)
[21]:
si.pl.discretize(adata_CG)
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[22]:
si.tl.gen_graph(list_CG=[adata_CG],
copy=False,
use_highly_variable=True,
dirname='graph0')
relation0: source: C, destination: G
#edges: 84634858
relation1: source: C, destination: G
#edges: 74099104
relation2: source: C, destination: G
#edges: 40189514
relation3: source: C, destination: G
#edges: 14186556
relation4: source: C, destination: G
#edges: 3065315
Total number of edges: 216175347
Writing graph file "pbg_graph.txt" to "result_simba_rnaseq/pbg/graph0" ...
Finished.
[23]:
# modify parameters
dict_config = si.settings.pbg_params.copy()
dict_config['workers'] = 12 #The number of CPUs.
## start training
si.tl.pbg_train(pbg_params = dict_config, auto_wd=True, save_wd=True, output='model')
Auto-estimated weight decay is 0.000109
`.settings.pbg_params['wd']` has been updated to 0.000109
Converting input data ...
[2021-10-27 20:14:59.424864] Using the 5 relation types given in the config
[2021-10-27 20:14:59.425251] Searching for the entities in the edge files...
[2021-10-27 20:18:55.307204] Entity type C:
[2021-10-27 20:18:55.307964] - Found 1306127 entities
[2021-10-27 20:18:55.308346] - Removing the ones with fewer than 1 occurrences...
[2021-10-27 20:18:55.769544] - Left with 1306127 entities
[2021-10-27 20:18:55.770258] - Shuffling them...
[2021-10-27 20:18:56.854065] Entity type G:
[2021-10-27 20:18:56.854702] - Found 2000 entities
[2021-10-27 20:18:56.854991] - Removing the ones with fewer than 1 occurrences...
[2021-10-27 20:18:56.855821] - Left with 2000 entities
[2021-10-27 20:18:56.856118] - Shuffling them...
[2021-10-27 20:18:56.872202] Preparing counts and dictionaries for entities and relation types:
[2021-10-27 20:18:56.874087] - Writing count of entity type C and partition 0
[2021-10-27 20:18:57.477737] - Writing count of entity type G and partition 0
[2021-10-27 20:18:57.480468] Preparing edge path result_simba_rnaseq/pbg/graph0/input/edge, out of the edges found in result_simba_rnaseq/pbg/graph0/pbg_graph.txt
using fast version
[2021-10-27 20:18:57.481362] Taking the fast train!
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Starting training ...
[24]:
si.pl.pbg_metrics(fig_ncol=1)
[25]:
dict_adata = si.read_embedding()
dict_adata
[25]:
{'C': AnnData object with n_obs × n_vars = 1306127 × 50,
'G': AnnData object with n_obs × n_vars = 2000 × 50}
[26]:
adata_C = dict_adata['C'] # embeddings for cells
adata_G = dict_adata['G'] # embeddings for genes
[27]:
## Add annotation of celltypes (optional)
adata_C.obs['Cluster'] = adata_CG[adata_C.obs_names,:].obs['Cluster'].copy()
adata_C
[27]:
AnnData object with n_obs × n_vars = 1306127 × 50
obs: 'Cluster'
[28]:
si.tl.umap(adata_C,n_neighbors=10,n_components=2, n_jobs=12)
[ ]:
[29]:
si.pl.umap(adata_C,color=['Cluster'],
fig_size=(10,8),
drawing_order='random',
size=0.1,
fig_legend_ncol=3,
)
[ ]:
[30]:
# embed cells and genes into the same space
adata_all = si.tl.embed(adata_ref=adata_C,list_adata_query=[adata_G])
Performing softmax transformation for query data 0;
[31]:
## add annotations of cells and genes
adata_all.obs['entity_anno'] = ""
adata_all.obs.loc[adata_C.obs_names, 'entity_anno'] = adata_all.obs.loc[adata_C.obs_names, 'Cluster']
adata_all.obs.loc[adata_G.obs_names, 'entity_anno'] = 'gene'
[32]:
si.tl.umap(adata_all,n_neighbors=10,n_components=2, n_jobs=12)
[33]:
palette_entity_anno = adata_C.uns['color']['Cluster_color'].copy()
palette_entity_anno['gene'] = "#607e95"
[ ]:
[34]:
si.pl.umap(adata_all,color=['id_dataset','entity_anno'],
dict_palette={'entity_anno': palette_entity_anno},
drawing_order='original',
fig_size=(10,8),
size=5,
fig_legend_ncol=3,)
[ ]:
[35]:
# find neighbor genes of given pins
query_genes = si.tl.query(adata_all,
pin=[6, 7],
use_radius=True, r=2,
obsm='X_umap',
anno_filter='entity_anno',
filters=['gene'])
print(query_genes.shape)
query_genes.head()
(153, 5)
[35]:
| Cluster | id_dataset | entity_anno | distance | query | |
|---|---|---|---|---|---|
| Pxdc1 | NaN | query_0 | gene | 0.007740 | 0 |
| Egln3 | NaN | query_0 | gene | 0.046162 | 0 |
| Pdpn | NaN | query_0 | gene | 0.075490 | 0 |
| Fgfbp3 | NaN | query_0 | gene | 0.076130 | 0 |
| Notch2 | NaN | query_0 | gene | 0.086966 | 0 |
[36]:
# show locations of pin points and its neighbor genes
si.pl.query(adata_all,
show_texts=False,
alpha=0.9,
alpha_bg=0.1,
fig_legend_ncol=1,
fig_size=(4,4))
[37]:
si.pl.umap(adata_all[::-1],
color=['entity_anno'],
dict_palette={'entity_anno': palette_entity_anno},
drawing_order='original',
show_texts=True,
texts=query_genes.index[:10],
text_expand=(1.2,1.4),
fig_size=(10,8),
size=0.1,
fig_legend_ncol=3)
[ ]:
[38]:
adata_CG.obsm['X_umap'] = adata_C[adata_CG.obs_names,].obsm['X_umap'].copy()
[ ]:
[39]:
si.pl.umap(adata_CG,
color=query_genes.index[:6],
drawing_order='sorted',
size=1,
alpha=0.5,
fig_ncol=3,
save_fig=False)
[ ]:
[40]:
# find neighbor genes of given pins
query_genes = si.tl.query(adata_all,
pin=[-7.5, 12.5],
use_radius=True, r=2,
obsm='X_umap',
anno_filter='entity_anno',
filters=['gene'])
print(query_genes.shape)
query_genes.head()
(52, 5)
[40]:
| Cluster | id_dataset | entity_anno | distance | query | |
|---|---|---|---|---|---|
| Reln | NaN | query_0 | gene | 0.035584 | 0 |
| Npnt | NaN | query_0 | gene | 0.041992 | 0 |
| Trp73 | NaN | query_0 | gene | 0.062758 | 0 |
| Ebf3 | NaN | query_0 | gene | 0.063638 | 0 |
| Lhx1 | NaN | query_0 | gene | 0.106870 | 0 |
[42]:
# show locations of pin points and its neighbor genes
si.pl.query(adata_all,
show_texts=False,
alpha=0.9,
alpha_bg=0.1,
fig_legend_ncol=1,
fig_size=(4,4))
[43]:
si.pl.umap(adata_all[::-1],
color=['entity_anno'],
dict_palette={'entity_anno': palette_entity_anno},
drawing_order='original',
show_texts=True,
texts=query_genes.index[:5],
text_expand=(1.2,1.4),
fig_size=(10,8),
size=0.1,
fig_legend_ncol=3)
[ ]:
[44]:
si.pl.umap(adata_CG,
color=query_genes.index[:3],
drawing_order='sorted',
size=1,
alpha=0.5,
fig_ncol=3,
save_fig=False)
[ ]:
[45]:
adata_CG.write(os.path.join(workdir, 'adata_CG.h5ad'))
adata_C.write(os.path.join(workdir, 'adata_C.h5ad'))
adata_G.write(os.path.join(workdir, 'adata_G.h5ad'))
adata_all.write(os.path.join(workdir, 'adata_all.h5ad'))
[ ]: