Last updated: 2021-10-29

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Knit directory: mistyMBC/

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Rmd 5d2c9ad Jovan Tanevski 2021-09-22 add size filter of cells in codex, zoi paraview
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Rmd 125da67 Jovan Tanevski 2021-07-14 add slideseq cellcomm pipeline and outputs
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Rmd eb056ff Jovan Tanevski 2021-06-10 add simple misty analysis

Setup

library(reticulate)
library(mistyR)
library(future)
library(dplyr)
library(purrr)
library(stringr)

use_python("/usr/local/bin/python3")
plan(multisession)

Extract counts and geometry

Requires python3 and anndata installed (pip install anndata).

import anndata

def read_and_extract(datapath):
  adata = anndata.read(datapath)
  counts = adata.obsm["counts"]
  obs = adata.obs
  markers = adata.uns["counts_var"]
  return counts, obs, markers

Run misty

Define standard view composition and run MISTy on all h5ad files for modalities CODEX, MERFISH and ExSeq.

(list.files("data", ".h5ad", recursive = TRUE, full.names = TRUE) %>%
  keep(~ str_detect(.x, "(codex|merfish)"))) %>%
  walk(function(datapath) {
    data <- py$read_and_extract(datapath)
    
    if (str_detect(datapath, "codex")) {
      transformed <- asinh(as.matrix(data[[1]]))
    } else {
      counts  <- as.matrix(data[[1]])
      transformed <- log((counts/rowSums(counts))*1e5 + 1)
    }
    
    expr <- as.data.frame(transformed)
    colnames(expr) <- make.names(data[[3]])
    obs <- data[[2]]
    pos <- obs %>% select(x_orig, y_orig)

    unique(str_extract(rownames(pos), "-\\d$")) %>% walk(function(replicate) {
      output.folder <- paste0(
        str_replace(
          str_remove(datapath, ".h5ad"),
          "data", "output"
        ), replicate
      )

      output.folder.failed <- paste0(output.folder, "_failed")

      if (!(dir.exists(output.folder) | dir.exists(output.folder.failed))) {
        replicate.ind <- str_which(rownames(pos), paste0(replicate, "$"))

        # in codex filter cells with outlier sizes (Tukey's interquartile approach)
        if (str_detect(datapath, "codex")) {
          sizes <- obs %>%
            slice(replicate.ind) %>%
            pull("size")
          quartiles <- quantile(sizes, c(.25, .75))
          lower <- quartiles[1] - 1.5 * (quartiles[2] - quartiles[1])
          upper <- quartiles[2] + 1.5 * (quartiles[2] - quartiles[1])
          ind <- replicate.ind[which(sizes < upper & sizes > lower)]
        } else {
          ind <- replicate.ind
        }

        misty.views <- create_initial_view(expr[ind, ]) %>%
          add_juxtaview(pos[ind, ]) %>%
          add_paraview(pos[ind, ], l = 100, zoi = 15)

        tryCatch(
          run_misty(misty.views, results.folder = output.folder),
          error = function(e) file.rename(output.folder, output.folder.failed)
        )
      }
    })
  })

Computing triangulation

Generating juxtaview

Generating paraview

sessionInfo()
R version 4.1.1 (2021-08-10)
Platform: x86_64-apple-darwin17.0 (64-bit)
Running under: macOS Big Sur 10.16

Matrix products: default
BLAS:   /Library/Frameworks/R.framework/Versions/4.1/Resources/lib/libRblas.0.dylib
LAPACK: /Library/Frameworks/R.framework/Versions/4.1/Resources/lib/libRlapack.dylib

locale:
[1] en_US.UTF-8/en_US.UTF-8/en_US.UTF-8/C/en_US.UTF-8/en_US.UTF-8

attached base packages:
[1] stats     graphics  grDevices utils     datasets  methods   base     

other attached packages:
[1] stringr_1.4.0   purrr_0.3.4     dplyr_1.0.7     future_1.22.1  
[5] mistyR_1.1.14   reticulate_1.22 workflowr_1.6.2

loaded via a namespace (and not attached):
 [1] tidyselect_1.1.1  xfun_0.27         bslib_0.3.1       listenv_0.8.0    
 [5] distances_0.1.8   lattice_0.20-45   vctrs_0.3.8       generics_0.1.1   
 [9] htmltools_0.5.2   yaml_2.2.1        utf8_1.2.2        rlang_0.4.12     
[13] R.oo_1.24.0       jquerylib_0.1.4   later_1.3.0       pillar_1.6.4     
[17] glue_1.4.2        DBI_1.1.1         R.utils_2.11.0    lifecycle_1.0.1  
[21] R.methodsS3_1.8.1 codetools_0.2-18  evaluate_0.14     knitr_1.36       
[25] fastmap_1.1.0     httpuv_1.6.3      parallel_4.1.1    fansi_0.5.0      
[29] furrr_0.2.3       Rcpp_1.0.7        promises_1.2.0.1  jsonlite_1.7.2   
[33] deldir_1.0-6      parallelly_1.28.1 fs_1.5.0          png_0.1-7        
[37] digest_0.6.28     stringi_1.7.5     rlist_0.4.6.2     grid_4.1.1       
[41] rprojroot_2.0.2   tools_4.1.1       magrittr_2.0.1    sass_0.4.0       
[45] tibble_3.1.5      crayon_1.4.1      whisker_0.4       pkgconfig_2.0.3  
[49] ellipsis_0.3.2    Matrix_1.3-4      data.table_1.14.2 assertthat_0.2.1 
[53] rmarkdown_2.11    R6_2.5.1          globals_0.14.0    git2r_0.28.0     
[57] compiler_4.1.1