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Fast, sensitive and accurate integration of single-cell data with Harmony
https://portals.broadinstitute.org/harmony/
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How do I download version 0.99.9 of harmony? #235

Closed Yuan-9 closed 9 months ago

Yuan-9 commented 9 months ago

How do I download version 0.99.9 of harmony? Many of the papers I need to reproduce use this version of harmony, but I can't find a way to install it anywhere.

pati-ni commented 9 months ago

This may be useful:

https://github.com/immunogenomics/harmony/releases

I would expect that v0.1 would reproduce most results. Version 1 is different.

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How do I download version 0.99.9 of harmony? Many of the papers I need to reproduce use this version of harmony, but I can't find a way to install it anywhere.

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Yuan-9 commented 9 months ago

Thanks for your very quick reply! I tried using the v0.1 version of harmony provided in the repository, but when I installed it it said the version was 1.0.

packageVersion('harmony') [1] ‘1.0’ remove.packages("harmony") Removing package from ‘/usr/local/lib/R/site-library’ (as ‘lib’ is unspecified) packageVersion('harmony') Error in packageVersion("harmony") : there is no package called ‘harmony’ devtools::install_version("harmony", version = "0.1") Warning: unable to access index for repository https://inwt-vmeh2.inwt.de/r-repo/src/contrib: cannot open URL 'https://inwt-vmeh2.inwt.de/r-repo/src/contrib/PACKAGES' Trying https://packagemanager.posit.co/cran/2020-04-23 Trying file:////r-repo/ Trying https://inwt-vmeh2.inwt.de/r-repo/ Trying https://inwtlab.github.io/drat/ Error in download_version_url(package, version, repos, type): couldn't find package 'harmony' install.packages("/home/1/harmony-0.1.tar.gz", repos = NULL, type = "source") Installing package into ‘/usr/local/lib/R/site-library’ (as ‘lib’ is unspecified)

Warning in untar2(tarfile, files, list, exdir, restore_times): skipping pax global extended headers

pati-ni commented 9 months ago

You need to install it through the release zip on the link I provided.

I believe that you can do that with the devtools::install_local($PATH_EXTRACTED_ZIP) or with the R CMD INSTALL

Yuan-9 commented 9 months ago

I found a 0.1 version of harmony on cran and after I ran: Error in UseMethod("RunHarmony") : no applicable method for 'RunHarmony' applied to an object of class "seurat"

traceback() 6: RunHarmony(object = b_seurat, group.by.vars = batch_label, theta = theta_harmony, plot_convergence = TRUE, nclust = numclust, max.iter.cluster = max_iter_cluster) at call_harmony.R#100 5: call_harmony(b_seurat, batch_label, celltype_label, npcs, plotout_dir = working_dir, saveout_dir = working_dir, outfilename_prefix = outfile_prefix, visualize = visualize, save_obj = save_obj) at run_harmony_02.R#73 4: eval(ei, envir) 3: eval(ei, envir) 2: withVisible(eval(ei, envir)) 1: source("run_harmony_02.R") packageVersion("harmony") [1] ‘0.1.0’

Yuan-9 commented 9 months ago

I don't have this problem when running version 1.0 of harmony. When using version 1.0, I will get an error: Error in eval(ei, envir) : element-wise pow(): incompatible matrix dimensions: 50x2 and 2x1 traceback() 13: stop(list(message = "element-wise pow(): incompatible matrix dimensions: 50x2 and 2x1", call = eval(ei, envir), cppstack = NULL)) 12: .External(list(name = "CppMethod__invoke_notvoid", address = <pointer: 0x5613a16a03b0>, dll = list(name = "Rcpp", path = "/usr/local/lib/R/site-library/Rcpp/libs/Rcpp.so", dynamicLookup = TRUE, handle = <pointer: 0x5613a24d9230>, info = <pointer: 0x5613a029ae40>), numParameters = -1L), <pointer: 0x5613a5371390>, <pointer: 0x5613a27b3bc0>, .pointer) 11: harmonyObj$cluster_cpp() 10: cluster(harmonyObj) 9: harmonize(harmonyObj, max.iter.harmony, verbose) 8: HarmonyMatrix(pca_embedding, metavars_df, group.by.vars, FALSE, 0, theta, lambda, sigma, nclust, tau, block.size, max.iter.harmony, max.iter.cluster, epsilon.cluster, epsilon.harmony, plot_convergence, FALSE, verbose, reference_values) 7: RunHarmony.seurat(object = b_seurat, group.by.vars = batch_label, theta = theta_harmony, plot_convergence = TRUE, nclust = numclust, max.iter.cluster = max_iter_cluster) 6: RunHarmony(object = b_seurat, group.by.vars = batch_label, theta = theta_harmony, plot_convergence = TRUE, nclust = numclust, max.iter.cluster = max_iter_cluster) at call_harmony.R#100 5: call_harmony(b_seurat, batch_label, celltype_label, npcs, plotout_dir = working_dir, saveout_dir = working_dir, outfilename_prefix = outfile_prefix, visualize = visualize, save_obj = save_obj) at run_harmony_02.R#73 4: eval(ei, envir) 3: eval(ei, envir) 2: withVisible(eval(ei, envir)) 1: source("run_harmony_02.R")

Is there any information that can help me fix the bug?

pati-ni commented 9 months ago

I found a 0.1 version of harmony on cran and after I ran: Error in UseMethod("RunHarmony") : no applicable method for 'RunHarmony' applied to an object of class "seurat"

traceback() 6: RunHarmony(object = b_seurat, group.by.vars = batch_label, theta = theta_harmony, plot_convergence = TRUE, nclust = numclust, max.iter.cluster = max_iter_cluster) at call_harmony.R#100 5: call_harmony(b_seurat, batch_label, celltype_label, npcs, plotout_dir = working_dir, saveout_dir = working_dir, outfilename_prefix = outfile_prefix, visualize = visualize, save_obj = save_obj) at run_harmony_02.R#73 4: eval(ei, envir) 3: eval(ei, envir) 2: withVisible(eval(ei, envir)) 1: source("run_harmony_02.R") packageVersion("harmony") [1] ‘0.1.0’

What values of theta are you using? Also what is the batch_label? Is this harmony v1?

Yuan-9 commented 9 months ago

I found a 0.1 version of harmony on cran and after I ran: Error in UseMethod("RunHarmony") : no applicable method for 'RunHarmony' applied to an object of class "seurat"

traceback() 6: RunHarmony(object = b_seurat, group.by.vars = batch_label, theta = theta_harmony, plot_convergence = TRUE, nclust = numclust, max.iter.cluster = max_iter_cluster) at call_harmony.R#100 5: call_harmony(b_seurat, batch_label, celltype_label, npcs, plotout_dir = working_dir, saveout_dir = working_dir, outfilename_prefix = outfile_prefix, visualize = visualize, save_obj = save_obj) at run_harmony_02.R#73 4: eval(ei, envir) 3: eval(ei, envir) 2: withVisible(eval(ei, envir)) 1: source("run_harmony_02.R") packageVersion("harmony") [1] ‘0.1.0’

I call it like this:

Harmony settings

theta_harmony = 2 numclust = 50 max_iter_cluster = 100 b_seurat <- RunPCA(object = b_seurat, pcs.compute = npcs, pc.genes = b_seurat@var.genes, do.print = TRUE, pcs.print = 1:5, genes.print = 5)

b_seurat <- RunHarmony(object = b_seurat, group.by.vars =batch_label, theta = theta_harmony, plot_convergence = TRUE, nclust = numclust, max.iter.cluster = max_iter_cluster)

harmony version is 0.1.0, Seurat version is 2.3.3

Yuan-9 commented 9 months ago

I don't have this problem when running version 1.0 of harmony. When using version 1.0, I will get an error: Error in eval(ei, envir) : element-wise pow(): incompatible matrix dimensions: 50x2 and 2x1 traceback() 13: stop(list(message = "element-wise pow(): incompatible matrix dimensions: 50x2 and 2x1", call = eval(ei, envir), cppstack = NULL)) 12: .External(list(name = "CppMethod__invoke_notvoid", address = <pointer: 0x5613a16a03b0>, dll = list(name = "Rcpp", path = "/usr/local/lib/R/site-library/Rcpp/libs/Rcpp.so", dynamicLookup = TRUE, handle = <pointer: 0x5613a24d9230>, info = <pointer: 0x5613a029ae40>), numParameters = -1L), <pointer: 0x5613a5371390>, <pointer: 0x5613a27b3bc0>, .pointer) 11: harmonyObj$cluster_cpp() 10: cluster(harmonyObj) 9: harmonize(harmonyObj, max.iter.harmony, verbose) 8: HarmonyMatrix(pca_embedding, metavars_df, group.by.vars, FALSE, 0, theta, lambda, sigma, nclust, tau, block.size, max.iter.harmony, max.iter.cluster, epsilon.cluster, epsilon.harmony, plot_convergence, FALSE, verbose, reference_values) 7: RunHarmony.seurat(object = b_seurat, group.by.vars = batch_label, theta = theta_harmony, plot_convergence = TRUE, nclust = numclust, max.iter.cluster = max_iter_cluster) 6: RunHarmony(object = b_seurat, group.by.vars = batch_label, theta = theta_harmony, plot_convergence = TRUE, nclust = numclust, max.iter.cluster = max_iter_cluster) at call_harmony.R#100 5: call_harmony(b_seurat, batch_label, celltype_label, npcs, plotout_dir = working_dir, saveout_dir = working_dir, outfilename_prefix = outfile_prefix, visualize = visualize, save_obj = save_obj) at run_harmony_02.R#73 4: eval(ei, envir) 3: eval(ei, envir) 2: withVisible(eval(ei, envir)) 1: source("run_harmony_02.R")

Is there any information that can help me fix the bug?

The harmony version of this is 1.0

Yuan-9 commented 9 months ago

batch_label is "batch"

b_seurat@meta.data nGene nUMI batch T-cell_Thymus_1.GCGTCCGGTACACCGCTA 525 672 1 T-cell_Thymus_2.CGAGTATAGTCGGTCGGT 885 1859 1

pati-ni commented 9 months ago

Can you check whether the meta.data$batch is a factor? If not, cast it as such.

Yuan-9 commented 9 months ago

I changed it to this:

b_seurat <- RunPCA(object = b_seurat, pcs.compute = npcs, pc.genes = b_seurat@var.genes, do.print = TRUE, pcs.print = 1:5, genes.print = 5) if (!is.factor(b_seurat@meta.data$batch)) { print("meta.data$batch is not a factor") b_seurat@meta.data$batch <- as.factor(b_seurat@meta.data$batch) } b_seurat <- RunHarmony(object = b_seurat, group.by.vars =batch_label, theta = theta_harmony, plot_convergence = TRUE, nclust = numclust, max.iter.cluster = max_iter_cluster)

It still has the original problem: [1] "meta.data$batch is not a factor" Error in UseMethod("RunHarmony") : no applicable method for 'RunHarmony' applied to an object of class "seurat"

traceback() 6: RunHarmony(object = b_seurat, group.by.vars = batch_label, theta = theta_harmony, plot_convergence = TRUE, nclust = numclust, max.iter.cluster = max_iter_cluster) at call_harmony.R#103 5: call_harmony(b_seurat, batch_label, celltype_label, npcs, plotout_dir = working_dir, saveout_dir = working_dir, outfilename_prefix = outfile_prefix, visualize = visualize, save_obj = save_obj) at run_harmony_02.R#73 4: eval(ei, envir) 3: eval(ei, envir) 2: withVisible(eval(ei, envir)) 1: source("run_harmony_02.R")

pati-ni commented 9 months ago

Please provide a minimal reproducible example

Yuan-9 commented 9 months ago

I packaged my working environment as a docker at https://hub.docker.com/r/jxbsx/bug. You can start it with the following command: docker run -it --name try_idea -p 9001:8787 jxbsx/bug:1.1 R The running file path is /home/2/run_harmony_02.R You need to execute setwd("/home/2") first

Yuan-9 commented 9 months ago

Additionally, I have a very simple test code running in my environment:

Load necessary packages

library(Seurat) library(harmony)

Load the pbmc_small dataset

data("pbmc_small")

Add a batch variable

batch_data <- data.frame(batch = sample(c("batch1", "batch2"), size = nrow(pbmc_small@meta.data), replace = TRUE)) pbmc_small <- AddMetaData(pbmc_small, metadata = batch_data)

Normalize the data

pbmc_small <- NormalizeData(pbmc_small)

Find variable genes

pbmc_small <- FindVariableGenes(pbmc_small)

RunPCA

pbmc_small <- RunPCA(pbmc_small, pcs.print = 1:5, genes.print = 5)

RunHarmony

pbmc_small <- RunHarmony(pbmc_small, group.by.vars = "batch")

The same problem occurred: Error in UseMethod("RunHarmony") : no applicable method for 'RunHarmony' applied to an object of class "seurat"

pati-ni commented 9 months ago

Can you please share a sessionInfo() for your R environment?

With Seurat 4.9.9 I get this using your code.

data("pbmc_small")

#Add a batch variable
batch_data <- data.frame(batch = sample(c("batch1", "batch2"), size = nrow(pbmc_small@meta.data), replace = TRUE))
pbmc_small <- AddMetaData(pbmc_small, metadata = batch_data)
Error: No cell overlap between new meta data and Seurat object
sessionInfo()
R version 4.3.2 (2023-10-31)
Platform: x86_64-pc-linux-gnu (64-bit)
Running under: Arch Linux

Matrix products: default
BLAS/LAPACK: /usr/lib/libopenblas.so.0.3;  LAPACK version 3.11.0

locale:
 [1] LC_CTYPE=en_US.UTF-8       LC_NUMERIC=C
 [3] LC_TIME=en_US.UTF-8        LC_COLLATE=en_US.UTF-8
 [5] LC_MONETARY=en_US.UTF-8    LC_MESSAGES=en_US.UTF-8
 [7] LC_PAPER=en_US.UTF-8       LC_NAME=C
 [9] LC_ADDRESS=C               LC_TELEPHONE=C
[11] LC_MEASUREMENT=en_US.UTF-8 LC_IDENTIFICATION=C

time zone: America/New_York
tzcode source: system (glibc)

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

other attached packages:
[1] Seurat_4.9.9.9067       SeuratObject_4.9.9.9091 sp_1.6-0
Yuan-9 commented 9 months ago

sessionInfo() R version 3.6.3 (2020-02-29) Platform: x86_64-pc-linux-gnu (64-bit) Running under: Ubuntu 20.04.6 LTS

Matrix products: default BLAS/LAPACK: /usr/lib/x86_64-linux-gnu/openblas-pthread/libopenblasp-r0.3.8.so

locale: [1] LC_CTYPE=en_US.UTF-8 LC_NUMERIC=C
[3] LC_TIME=en_US.UTF-8 LC_COLLATE=en_US.UTF-8
[5] LC_MONETARY=en_US.UTF-8 LC_MESSAGES=C
[7] LC_PAPER=en_US.UTF-8 LC_NAME=C
[9] LC_ADDRESS=C LC_TELEPHONE=C
[11] LC_MEASUREMENT=en_US.UTF-8 LC_IDENTIFICATION=C

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

other attached packages: [1] harmony_0.1.0 Rcpp_1.0.11 Seurat_2.3.3 Matrix_1.2-18 cowplot_1.1.2 [6] ggplot2_3.4.4

pati-ni commented 9 months ago

Seurat v2 was released on 2016, 3 years before harmony was published. Upgrade this package to get proper support

Yuan-9 commented 9 months ago

Since the environment used in the original paper was Seurat v2, I could not upgrade it when reproducing it. Thank you for your help.

pati-ni commented 9 months ago

In that case, use the HarmonyMatrix (in v0) instead of the Seurat wrapper.

Yuan-9 commented 9 months ago

Okay, I'm going to try to use this function. Thank you for your help.