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Generate plots and tables with SampCompDB

SampComp creates a python object database (shelve DBM) containing the statistical analysis results. The API directly returns a SampCompDB object wrapping the shelve DB. It is also possible to reload the SampCompDB latter using the db file path prefix. SampCompDB also need a FASTA file to get the corresponding reference id sequence and accept an optional BED file containing genomic annotations. SampCompDB provide a large selection of simple high level function to plot and export the results.

At the moment SampCompDB is only accessible through the python API.

Import the package

from nanocompore.SampCompDB import SampCompDB, jhelp

Load the database with SampCompDB

jhelp (SampCompDB)

SampCompDB (db_fn, fasta_fn, bed_fn, run_type, log_level)

Wrapper over the result shelve SampComp


  • db_fn (required) [str]

Path to a database file previously created with SampComp

  • fasta_fn (required) [str]

Path to a fasta file corresponding to the reference used for read alignemnt

  • bed_fn (default: None) [str]

Path to a BED file containing the annotation of the transcriptome used as reference when mapping

  • run_type (default: RNA) [str]

Define the run type model to import {RNA, DNA}

  • log_level (default: info) [str]

Set the log level. {warning,info,debug}"

Basic initialisation

# Load database
db = SampCompDB (
    db_fn = "results/simulated_SampComp.db",
    fasta_fn = "references/simulated/ref.fa")

# Print general metadata information
print (db)

# Prit list of references containing valid data
print (db.ref_id_list)
Loading SampCompDB
Calculate results


[SampCompDB]
    package_name: nanocompore
    package_version: 1.0.0rc3-dev
    timestamp: 2019-06-19 14:11:42.582069
    comparison_methods: ['GMM', 'KS']
    pvalue_tests: ['GMM_anova_pvalue', 'KS_dwell_pvalue', 'KS_intensity_pvalue']
    sequence_context: 0
    min_coverage: 30
    n_samples: 4
    Number of references: 5

['ref_0002', 'ref_0000', 'ref_0003', 'ref_0001', 'ref_0004']

Generate text reports

SampCompDB can generate 3 types of text reports: * Tabulated statistics => save_report * Tabulated intensity and dwell values per conditions => save_shift_stats * BED significant genomic positions => save_to_bed

In addition, we also conveniently wrapped all 3 methods in save_all.

save_report

jhelp(SampCompDB.save_report)

save_report (output_fn)

Saves a tabulated text dump of the database containing all the statistical results for all the positions


  • output_fn (default: None) [str]

Path to file where to write the data. If None, data is returned to the standard output.

# Reload DB
db = SampCompDB (db_fn = "results/simulated_SampComp.db", fasta_fn = "references/simulated/ref.fa", log_level="warning")

# Save report
db.save_report (output_fn="./results/simulated_report.tsv")

# Visualise first lines
!head "./results/simulated_report.tsv"
pos chr genomicPos  ref_id  strand  ref_kmer    GMM_anova_pvalue    KS_dwell_pvalue KS_intensity_pvalue GMM_cov_type    GMM_n_clust cluster_counts  Anova_delta_logit
0   NA  NA  ref_0002    NA  AAGGA   0.010390555392639545    3.7826176625121133e-28  7.372203704353806e-13   full    2   Modified_rep1:4/51__Modified_rep2:4/56__Unmodified_rep1:39/16__Unmodified_rep2:49/11    -3.5291008135000004
1   NA  NA  ref_0002    NA  AGGAC   0.018531016868604602    5.021033530354203e-38   2.6192924303287954e-32  full    2   Modified_rep1:49/6__Modified_rep2:54/6__Unmodified_rep1:0/55__Unmodified_rep2:3/57  5.363518116
2   NA  NA  ref_0002    NA  GGACT   0.016650130591381845    9.083535018319583e-23   1.9347009060981327e-19  full    2   Modified_rep1:48/7__Modified_rep2:53/7__Unmodified_rep1:13/42__Unmodified_rep2:7/53 3.3768032759999995
3   NA  NA  ref_0002    NA  GACTG   nan 1.5167780008574906e-20  5.038173683218352e-10   full    1   NC  nan
4   NA  NA  ref_0002    NA  ACTGC   nan 0.7103128384794648  0.2143969244232542  full    1   NC  nan
5   NA  NA  ref_0002    NA  CTGCG   nan 0.5427546298577994  0.9441367354284554  full    1   NC  nan
6   NA  NA  ref_0002    NA  TGCGC   nan 0.6283003014678024  0.2835540266938749  full    1   NC  nan
7   NA  NA  ref_0002    NA  GCGCC   nan 0.7818041839421846  0.2143969244232542  full    1   NC  nan
8   NA  NA  ref_0002    NA  CGCCG   nan 0.4544427448826311  0.8445913073368677  full    1   NC  nan

save_shift_stats

jhelp(SampCompDB.save_shift_stats)

save_shift_stats (output_fn)

Save the mean, median and sd intensity and dwell time for each condition and for each position. This can be used to evaluate the intensity of the shift for significant positions.


  • output_fn (default: None)

Path to file where to write the data. If None, data is returned to the standard output.

# Reload DB
db = SampCompDB (db_fn = "results/simulated_SampComp.db", fasta_fn = "references/simulated/ref.fa", log_level="warning")

# Save report
db.save_shift_stats (output_fn="./results/simulated_shift.tsv")

# Visualise first lines
!head "./results/simulated_shift.tsv"
red_if  pos c1_mean_intensity   c2_mean_intensity   c1_median_intensity c2_median_intensity c1_sd_intensity c2_sd_intensity c1_mean_dwell   c2_mean_dwell   c1_median_dwell c2_median_dwell c1_sd_dwell c2_sd_dwell
ref_0001    0   99.31924275131445   95.20865205128713   99.30450652082747   95.48506067828346   3.4542546529463602  3.2457394860292355  0.02892371350987724 0.011492822977360408    0.026110578053659444    0.008409393806440126    0.012309163786594407    0.010097740816656625
ref_0001    1   95.8472415436861    86.76320109231014   96.51658647295791   86.5897261563199    4.818212253968896   3.462616900325914   0.049904028745349535    0.014284063287085003    0.049430992173562574    0.00838888597355578 0.01891436186841055 0.01469932192592977
ref_0001    2   99.95956311201668   91.94076655256666   99.2359574515384    91.66515374416255   6.732308978957026   5.022072514781664   0.027605907187015038    0.013686251128416578    0.024211963436724607    0.009798007086063236    0.014206789096974112    0.01220646673010648
ref_0001    3   123.38251652061109  120.39646776876853  123.6421269382127   120.37875855935422  3.595296347867109   3.44338463383404    0.028760344887274834    0.013945564031975161    0.02351276462197541 0.010354002459941074    0.017565155869793847    0.011016717175902649
ref_0001    4   126.94163302104937  128.10426869865935  126.55839976875221  127.8992550363245   3.673183364723787   4.450043557112173   0.011798347544928565    0.013913244140038266    0.008085339548882277    0.009808706553017481    0.010235524821933415    0.012247487588286306
ref_0001    5   67.79566862215866   67.87066906855172   67.57337199388174   67.66864765166001   2.676099462286074   2.3271061496852905  0.011811464983494136    0.012975245722562987    0.007659091594070025    0.007913024971489328    0.01114312094701346 0.013957539542755816
ref_0001    6   66.155214143451 65.87786841257429   66.01760504576498   65.80903439895116   1.5942871182383656  1.9921259312963864  0.013708376752126073    0.012800371955813343    0.009551895684442788    0.008605404284103613    0.014921164088938836    0.011530961046002914
ref_0001    7   69.61169482565616   68.99766496300363   69.41928437670197   68.95292840911311   2.936936891666408   2.575355539206547   0.011149410753827556    0.011084327064933388    0.008581162093140242    0.007464155848405434    0.009824895741362088    0.01064339382049097
ref_0001    8   82.34611997642149   82.05089836082209   82.24692305018203   81.9154038674852    2.5845230611008763  2.4502360451089737  0.01267473742329643 0.012320988335592328    0.010373590682259867    0.009046996378582178    0.009070752556799426    0.00959451914969272

save_to_bed

jhelp(SampCompDB.save_to_bed)

save_to_bed (output_fn, bedgraph, pvalue_field, pvalue_thr, span, convert, assembly, title)

Save the position of significant positions in the genome space in BED6 or BEDGRAPH format. The resulting file can be used in a genome browser to visualise significant genomic locations. The option is only available if SampCompDB if initialised with a BED file containing genome annotations.


  • output_fn (default: None)

Path to file where to write the data

  • bedgraph (default: False)

save file in bedgraph format instead of bed

  • pvalue_field (default: None)

specifies what column to use as BED score (field 5, as -log10)

  • pvalue_thr (default: 0.01)

only report positions with pvalue<=thr

  • span (default: 5)

The size of each BED feature. If size=5 (default) features correspond to kmers. If size=1 features correspond to the first base of each kmer.

  • convert (default: None)

one of 'ensembl_to_ucsc' or 'ucsc_to_ensembl". Convert chromosome named between Ensembl and Ucsc conventions

  • assembly (default: None)

required if convert is used. One of "hg38" or "mm10"

  • title (default: None)
# Reload DB
db = SampCompDB (db_fn = "results/simulated_SampComp.db", fasta_fn = "references/simulated/ref.fa", bed_fn="references/simulated/annot.bed", log_level="warning")

# Save report
db.save_to_bed (output_fn="./results/simulated_sig_positions.bed")

# Visualise first lines
!head "./results/simulated_sig_positions.bed"

Generate plots

SampCompDB comes with a range of methods to visualise the data and explore candidates.

  • plot_pvalue: Plot the -log(10) of the pvalues obtained for all the statistical methods at reference level
  • plot_signal: Generate comparative plots of both median intensity and dwell time for each condition at read level
  • plot_coverage: Plot the read coverage over a reference for all samples analysed
  • plot_kmers_stats: Fancy version of plot_coverage that also report missing, mismatching and undefined kmers status from Nanopolish
  • plot_position: Allow to visualise the distribution of intensity and dwell time in 2D for a single position

Extra imports for the plotting library

Matplotlib is required to use the ploting methods in Jupyter

import matplotlib.pyplot as pl
%matplotlib inline

plot_pvalue

jhelp(SampCompDB.plot_pvalue)

plot_pvalue (ref_id, start, end, kind, threshold, figsize, palette, plot_style, tests)

Plot pvalues per position (by default plot all fields starting by "pvalue")


  • ref_id (required) [str]

Valid reference id name in the database

  • start (default: None) [int]

Start coordinate

  • end (default: None) [int]

End coordinate (included)

  • kind (default: lineplot) [str]

kind of plot to represent the data. {lineplot,barplot}

  • threshold (default: 0.01) [float]

  • figsize (default: (30, 10)) [tuple]

Length and heigh of the output plot

  • palette (default: Set2) [str]

Colormap. See https://matplotlib.org/users/colormaps.html, https://matplotlib.org/examples/color/named_colors.html

  • plot_style (default: ggplot) [str]

Matplotlib plotting style. See https://matplotlib.org/users/style_sheets.html

  • tests (default: None) [str]

Limit the pvalue methods shown in the plot. Either a list of methods or a string coresponding to a part of the name

Examples from simulated dataset

# Reload DB
db = SampCompDB (db_fn = "results/simulated_SampComp.db", fasta_fn = "references/simulated/ref.fa", log_level="warning")
# Plot
fig, ax = db.plot_pvalue ("ref_0000")

png

# Reload DB
db = SampCompDB (db_fn = "results/simulated_stats_SampComp.db", fasta_fn = "references/simulated/ref.fa", log_level="warning")
# Plot
fig, ax = db.plot_pvalue ("ref_0001", palette="Set1")

png

Example from real yeast dataset with extended sequence context

# Reload DB
db = SampCompDB (db_fn = "results/yeast_SampComp.db", fasta_fn = "references/yeast/Yeast_transcriptome.fa", log_level="warning")
# Plot
fig, ax = db.plot_pvalue ("YHR174W")

png

plot_signal

jhelp(SampCompDB.plot_signal)

plot_signal (ref_id, start, end, kind, split_samples, figsize, palette, plot_style)

Plot the dwell time and median intensity distribution position per position Pointless for more than 50 positions at once as it becomes hard to distinguish


  • ref_id (required) [str]

Valid reference id name in the database

  • start (default: None) [int]

Start coordinate

  • end (default: None) [int]

End coordinate (included)

  • kind (default: violinplot) [str]

Kind of plot {violinplot, boxenplot, swarmplot}

  • split_samples (default: False) [bool]

If samples for a same condition are represented separatly. If false they are merged per condition

  • figsize (default: (30, 10)) [tuple]

Length and heigh of the output plot

  • palette (default: Set2) [str]

Colormap. See https://matplotlib.org/users/colormaps.html, https://matplotlib.org/examples/color/named_colors.html

  • plot_style (default: ggplot) [str]

Matplotlib plotting style. See https://matplotlib.org/users/style_sheets.html

Examples from simulated dataset

# Reload DB
db = SampCompDB (db_fn = "results/simulated_SampComp.db", fasta_fn = "references/simulated/ref.fa", log_level="warning")
# Plot
fig, ax = db.plot_signal ("ref_0000", start=75, end=100)

png

# Reload DB
db = SampCompDB (db_fn = "results/simulated_SampComp.db", fasta_fn = "references/simulated/ref.fa", log_level="warning")
# Plot
fig, ax = db.plot_signal ("ref_0001", start=100, end=125, kind="swarmplot")

png

Example from real yeast dataset

# Reload DB
db = SampCompDB (db_fn = "results/yeast_SampComp.db", fasta_fn = "references/yeast/Yeast_transcriptome.fa", log_level="warning")
# Plot
fig, ax = db.plot_signal ("YHR174W", start=665, end=700, kind="boxenplot")

png

plot_coverage

jhelp(SampCompDB.plot_coverage)

plot_coverage (ref_id, start, end, scale, split_samples, figsize, palette, plot_style)

Plot the read coverage over a reference for all samples analysed


  • ref_id (required) [str]

Valid reference id name in the database

  • start (default: None) [int]

Start coordinate

  • end (default: None) [int]

End coordinate (included)

  • scale (default: False) [bool]

  • split_samples (default: False) [bool]

  • figsize (default: (30, 5)) [tuple]

Length and heigh of the output plot

  • palette (default: Set2) [str]

Colormap. See https://matplotlib.org/users/colormaps.html, https://matplotlib.org/examples/color/named_colors.html

  • plot_style (default: ggplot) [str]

Matplotlib plotting style. See https://matplotlib.org/users/style_sheets.html

Example from real yeast dataset

# Reload DB
db = SampCompDB (db_fn = "results/yeast_SampComp.db", fasta_fn = "references/yeast/Yeast_transcriptome.fa", log_level="warning")
# Plot
fig, ax = db.plot_coverage ("YHR174W")

png

plot_kmers_stats

jhelp(SampCompDB.plot_kmers_stats)

plot_kmers_stats (ref_id, start, end, split_samples, figsize, palette, plot_style)

Fancy version of plot_coverage that also report missing, mismatching and undefined kmers status from Nanopolish


  • ref_id (required) [str]

Valid reference id name in the database

  • start (default: None) [int]

Start coordinate

  • end (default: None) [int]

End coordinate (included)

  • split_samples (default: False) [bool]

  • figsize (default: (30, 10)) [tuple]

Length and heigh of the output plot

  • palette (default: Accent) [str]

Colormap. See https://matplotlib.org/users/colormaps.html, https://matplotlib.org/examples/color/named_colors.html

  • plot_style (default: ggplot) [str]

Matplotlib plotting style. See https://matplotlib.org/users/style_sheets.html

Example from real yeast dataset

# Reload DB
db = SampCompDB (db_fn = "results/yeast_SampComp.db", fasta_fn = "references/yeast/Yeast_transcriptome.fa", log_level="warning")
# Plot
fig, ax = db.plot_kmers_stats ("YHR174W")

png

plot_position

jhelp(SampCompDB.plot_position)

plot_position (ref_id, pos, split_samples, figsize, palette, plot_style, xlim, ylim, alpha, pointSize, scatter, kde, model, gmm_levels)

Plot the dwell time and median intensity at the given position as a scatter plot.


  • ref_id (required) [str]

Valid reference id name in the database

  • pos (default: None) [int]

Position of interest

  • split_samples (default: False) [bool]

If True, samples for a same condition are represented separately. If False, they are merged per condition

  • figsize (default: (30, 10)) [tuple]

Length and heigh of the output plot

  • palette (default: Set2) [str]

Colormap. See https://matplotlib.org/users/colormaps.html, https://matplotlib.org/examples/color/named_colors.html

  • plot_style (default: ggplot) [str]

Matplotlib plotting style. See https://matplotlib.org/users/style_sheets.html

  • xlim (default: (None, None)) [tuple]

A tuple of explicit limits for the x axis

  • ylim (default: (None, None)) [tuple]

A tuple of explicit limits for the y axis

  • alpha (default: 0.3) [float]

  • pointSize (default: 20) [int]

int specifying the point size for the scatter plot

  • scatter (default: True) [bool]

if True, plot the individual data points

  • kde (default: True) [bool]

plot the KDE of the intensity/dwell bivarariate distributions in the two samples

  • model (default: False) [bool]

If true, plot the GMM density estimate

  • gmm_levels (default: 50) [int]

number of contour lines to use for the GMM countour plot

Example from simulated dataset

# Reload DB
db = SampCompDB (db_fn = "results/simulated_SampComp.db", fasta_fn = "references/simulated/ref.fa", log_level="warning")
# Plot
fig, ax = db.plot_position ("ref_0000", pos=82)

png