Error in find_common_diploid(bulks, gamma = gamma, alpha = alpha, ncores = ncores) : Error in smooth_segs(., min_genes = min_genes) :

[whitneyt1]

`> #Run Numbat

out=run_numbat(count_mat = mat_clean,

• lambdas_ref = numbat_ref_N2535,
• df_allele = df_allele,
• genome = 'hg38',
• t=1e-5,
• max_entropy = 0.8,
• ncores=4,
• plot = TRUE,
• out_dir = './Numbat_Outs/outs/'
• )

Numbat version: 1.3.2.1 Scistreer version: 1.2.0 Running under parameters: t = 1e-05 alpha = 1e-04 gamma = 20 min_cells = 50 init_k = 3 max_cost = 1246.2 n_cut = 0 max_iter = 2 max_nni = 100 min_depth = 0 use_loh = auto segs_loh = None call_clonal_loh = FALSE segs_consensus_fix = None multi_allelic = TRUE min_LLR = 5 min_overlap = 0.45 max_entropy = 0.8 skip_nj = FALSE diploid_chroms = None ncores = 4 ncores_nni = 4 common_diploid = TRUE tau = 0.3 check_convergence = FALSE plot = TRUE genome = hg38 Input metrics: 4154 cells Mem used: 2.16Gb Approximating initial clusters using smoothed expression .. Mem used: 2.16Gb number of genes left: 12099 running hclust... ! # Invaild edge matrix for . A is returned.
! # Invaild edge matrix for . A is returned. Iteration 1 Mem used: 7.2Gb High SNP contamination detected (72.9%). Please make sure that cells from only one individual are included in genotyping step. Expression noise level (MSE): medium (1.4). Running HMMs on 5 cell groups.. Error in find_common_diploid(bulks, gamma = gamma, alpha = alpha, ncores = ncores) : Error in smooth_segs(., min_genes = min_genes) : No segments containing more than 10 genes for CHROM 13,21. In addition: Warning message: In mclapply(bulks %>% split(.$sample), mc.cores = ncores, function(bulk) { : scheduled cores 2, 1 encountered errors in user code, all values of the jobs will be affected`

Hello, I am getting this error once run_numbat begins trying to run HMMs.

I am using numbat on a series of 10X Visium lung cancer tissues. my normal reference is a normal lung tissues 10X visium reaction. I have ensured the gene names are in the same format between the reference and test.

This is the script i used to loop through reactions to perform the pileup step.

Thank you!

[whitneyt1]

Hello, I have been unable to address this issue still. Is there any additional information I can provide? @teng-gao

[teng-gao]

Hello, if your Visium data is probed based chemistry Numbat doesn’t support this

[whitneyt1]

hi @teng-gao it is not probe based, it is 3' sequencing on frozen sections. Thanks!

[teng-gao]

Seems that the SNP profile looks aberrant:

High SNP contamination detected (72.9%). Please make sure that cells from only one individual are included in genotyping step.

It's hard to tell what the issue is however only based on this message ..

[whitneyt1]

Thank you, @teng-gao these visium reactions are individual patient tumors. Is there any additional information I can provide that would be helpful to deduce the issue? thanks!

[whitneyt1]

I identified that I was following the mouse tutorial for human spatial data and did not phase my human data appropriately as per the "Getting Started" vignette. After running pileup_and_phase.R correctly, it resolved this issue.