Error in using zumis to analyse transposon

[ROund-code]

Hi zumis teams:
Thank you very much for this amazing pipline!
I have generated smart3-seq data and try to use zumis to analysis diff mRNAs and transposons. for mRNAs, it went well. but I have troubles in transposon analysis. the only difference was the gtf I supplied in yaml file. it would be great if you could give me some suggestions!

YAML FILE:
sequence_files:
file1:
name: /media/nbfs/gongyy/3_smart_seq_rabbit/8_Project_gongyuanyuan_240322N/1_multiplexed_fastq_files/reads_for_zUMIs.R1.fastq.gz
base_definition:
- cDNA(23-150)
- UMI(12-19)
find_pattern: ATTGCGCAATG
file2:
name: /media/nbfs/gongyy/3_smart_seq_rabbit/8_Project_gongyuanyuan_240322N/1_multiplexed_fastq_files/reads_for_zUMIs.R2.fastq.gz
base_definition: cDNA(1-150)
file3:
name: /media/nbfs/gongyy/3_smart_seq_rabbit/8_Project_gongyuanyuan_240322N/1_multiplexed_fastq_files/reads_for_zUMIs.index.fastq.gz
base_definition: BC(1-8)
reference:
STAR_index: /media/nbfs/gongyy/3_smart_seq_rabbit/TE_STAR_index/index
GTF_file: /media/nbfs/gongyy/3_smart_seq_rabbit/TE_STAR_index/oryCun2.TE.gtf
additional_STAR_params: '--clip3pAdapterSeq CTGTCTCTTATACACATCT'
additional_files: ~
out_dir: /media/nbfs/gongyy/3_smart_seq_rabbit/8_Project_gongyuanyuan_240322N/3_TE_zUMIs
num_threads: 20
mem_limit: 50
filter_cutoffs:
BC_filter:
num_bases: 3
phred: 20
UMI_filter:
num_bases: 2
phred: 20
barcodes:
barcode_num: ~
barcode_file: /media/nbfs/gongyy/3_smart_seq_rabbit/8_Project_gongyuanyuan_240322N/1_multiplexed_fastq_files/reads_for_zUMIs.expected_barcodes.txt
automatic: no
BarcodeBinning: 0
nReadsperCell: 100
counting_opts:
introns: yes
downsampling: '0'
strand: 0
Ham_Dist: 1
velocyto: no
primaryHit: yes
twoPass: no
make_stats: yes
which_Stage: Filtering
Rscript_exec: Rscript
STAR_exec: STAR
pigz_exec: pigz
samtools_exec: samtools

output messages:
Good news! A newer version of zUMIs is available at https://github.com/sdparekh/zUMIs

---

Using miniconda environment for zUMIs!
note: internal executables will be used instead of those specified in the YAML file!

You provided these parameters:
YAML file: ./project8_TE.yaml
zUMIs directory: /media/nbfs/gongyy/3_smart_seq_rabbit/zUMIs
STAR executable STAR
samtools executable samtools
pigz executable pigz
Rscript executable Rscript
RAM limit: 50
zUMIs version 2.9.7e

Wed Apr 3 10:16:49 CST 2024
WARNING: The STAR version used for mapping is 2.7.3a and the STAR index was created using the ve rsion 2.7.1a. This may lead to an error while mapping. If you encounter any errors at the mappin g stage, please make sure to create the STAR index using STAR 2.7.3a.
Filtering...
Wed Apr 3 10:34:17 CST 2024
[1] " reads were assigned to barcodes that do not correspond to intact cells."
Mapping...
[1] "2024-04-03 10:34:19 CST"
Apr 03 10:34:19 ..... started STAR run
Apr 03 10:34:20 ..... loading genome
Apr 03 10:34:19 ..... started STAR run
Apr 03 10:34:20 ..... loading genome
Apr 03 10:35:44 ..... processing annotations GTF
Apr 03 10:36:10 ..... started mapping
Apr 03 10:36:40 ..... processing annotations GTF
Apr 03 10:37:06 ..... started mapping
Apr 03 10:57:18 ..... finished mapping
Apr 03 10:57:19 ..... finished successfully
Apr 03 11:03:54 ..... finished mapping
Apr 03 11:03:56 ..... finished successfully
Wed Apr 3 11:04:36 CST 2024
Counting...
[1] "2024-04-03 11:04:51 CST"
[1] "7.5e+07 Reads per chunk"
[1] "Loading reference annotation from:"
[1] "/media/nbfs/gongyy/3_smart_seq_rabbit/8_Project_gongyuanyuan_240322N/3_TE_zUMIs/project8_TE .final_annot.gtf"
Error: cannot allocate vector of size 6.6 Gb
In addition: Warning message:
as_quosure() requires an explicit environment as of rlang 0.3.0.
Please supply env.
This warning is displayed once per session.
Execution halted
Warning message:
system call failed: Cannot allocate memory
Wed Apr 3 12:39:59 CST 2024
Loading required package: yaml
Loading required package: Matrix
[1] "loomR found"
Error in gzfile(file, "rb") : cannot open the connection
Calls: rds_to_loom -> readRDS -> gzfile
In addition: Warning message:
In gzfile(file, "rb") :
cannot open compressed file '/media/nbfs/gongyy/3_smart_seq_rabbit/8_Project_gongyuanyuan_2403 22N/3_TE_zUMIs/zUMIs_output/expression/project8_TE.dgecounts.rds', probable reason 'No such file or directory'
Execution halted
Wed Apr 3 12:40:04 CST 2024
Descriptive statistics...
[1] "I am loading useful packages for plotting..."
[1] "2024-04-03 12:40:04 CST"
Error in gzfile(file, "rb") : cannot open the connection
Calls: readRDS -> gzfile
In addition: Warning message:
In gzfile(file, "rb") :
cannot open compressed file '/media/nbfs/gongyy/3_smart_seq_rabbit/8_Project_gongyuanyuan_2403 22N/3_TE_zUMIs/zUMIs_output/expression/project8_TE.dgecounts.rds', probable reason 'No such file or directory'
Execution halted
Wed Apr 3 12:40:12 CST 2024

It looks like the error occurred in the counting step, also the files are missing in expression and stats folder.

Best Regards,
Yuan