RNABloom not creating transcriptome files

[dvirdi01]

I ran rnabloom on several of my input files individually. out of my 10 samples, it did not produce any transcriptome file for 2 of them. Why is that? It finished running but there are no output files. I ran the following command: rnabloom -long sample.fastq -t 48 -outdir .../.../sample -k 10 -e 5

These are my versions:

• RNA-Bloom v2.0.1
• openjdk 20.0.2-internal 2023-07-18

[kmnip]

Please remove the options -k 10 -e 5 and try again. If the issue persists, please report the entire log file.

[dvirdi01]

Hi, I have run my files with both -k 10 e 5 and with the default settings in the past. It produces the transcriptome file for the default setting command but not for -k 10 e 5. This is the message I get for the -k 10 -e5 one:

Parsing sample.fastq... Parsed 21,301,762 sequences in 2h 7m 13s DBG Bloom filter FPR: 1.04 % Counting Bloom filter FPR: 1.04 % Stage 1 completed in 2h 7m 13s

Stage 2: Correct long reads for "rnabloom" Parsing sample.fastq... Corrected Read Lengths Sampling Distribution (n=10000) min q1 med q3 max 10 193 364 715 2643 Parsed 21,301,762 sequences. Kept: 21,301,202 (100.0 %) Discarded: 560 (0.00263 %) Artifacts: 4,220,516 (19.81299%) Corrected reads in 1h 43m 33s Extracting seed sequences... strobemers: n=3, k=11, wMin=12, wMax=61, depth=3 Bloom filter FPR: 91.0 % before: 20,782,469 after: 15,730 (0.0757 %) too short: 0 Extraction completed in 43m 1s Stage 2 completed in 2h 26m 35s

Stage 3: Assemble long reads for "rnabloom" Overlapping sequences... Parsed 231,695 overlap records in 15.355s total reads: 15,730

• unique: 5,086 (32.3 %)
  • multi-seg: 1,193 Unique reads extracted in 0.638s Overlapping sequences... Parsed 44,253 overlap records in 2.313s contained reads: 1,697 dovetail reads: 1,746 G: |V|=3,493 |E|=4,118 G: |V|=3,337 |E|=3,470 before: 6,368 after: 4,136 Laid out paths in 0.232s Mapping sequences... Mapping completed in 1h 4m 25s Polishing sequences... Error running command! See log file at /.../.../../../sample/rnabloom.longreads.assembly4.pol.fa.log Polishing completed in 4m 16s ERROR: Error polishing reads to unitigs! ERROR: Error assembling long reads!

This is the log file: [racon::Polisher::initialize] loaded target sequences 0.043017 s [racon::Polisher::initialize] loaded sequences 174.185133 s Killed

I also tried giving it multiple files as input so it does a combined assembly with default settings. However, it gives me this error:

Exception: java.lang.OutOfMemoryError thrown from the UncaughtExceptionHandler in thread "Thread-836" Exception in thread "Thread-825" java.lang.OutOfMemoryError: Java heap space Exception in thread "Thread-830" java.lang.OutOfMemoryError: Java heap space Exception in thread "Thread-832" java.lang.OutOfMemoryError: Java heap space Exception in thread "Thread-854" java.lang.OutOfMemoryError: Java heap space Exception in thread "Thread-838" java.lang.OutOfMemoryError: Java heap space Exception in thread "Thread-828" java.lang.OutOfMemoryError: Java heap space Exception in thread "Thread-822" java.lang.OutOfMemoryError: Java heap space Exception in thread "Thread-844" java.lang.OutOfMemoryError: Java heap space Exception in thread "Thread-823" java.lang.OutOfMemoryError: Java heap space Exception in thread "Thread-816" java.lang.OutOfMemoryError: Java heap space

Exception: java.lang.OutOfMemoryError thrown from the UncaughtExceptionHandler in thread "Thread-853" Exception in thread "Thread-839" Exception in thread "Thread-850" java.lang.OutOfMemoryError: Java heap space java.lang.OutOfMemoryError: Java heap space Exception in thread "Thread-860" Exception in thread "Thread-852" java.lang.OutOfMemoryError: Java heap space java.lang.OutOfMemoryError: Java heap space Exception in thread "Thread-841" java.lang.OutOfMemoryError: Java heap space

Exception: java.lang.OutOfMemoryError thrown from the UncaughtExceptionHandler in thread "Thread-827" Exception in thread "Thread-846" java.lang.OutOfMemoryError: Java heap space Exception in thread "Thread-821" java.lang.OutOfMemoryError: Java heap space Exception in thread "Thread-831" java.lang.OutOfMemoryError: Java heap space Exception in thread "Thread-835" java.lang.OutOfMemoryError: Java heap space Exception in thread "Thread-849" java.lang.OutOfMemoryError: Java heap space Exception in thread "Thread-817" java.lang.OutOfMemoryError: Java heap space Exception in thread "Thread-859" java.lang.OutOfMemoryError: Java heap space Exception in thread "Thread-824" java.lang.OutOfMemoryError: Java heap space Exception in thread "Thread-843" java.lang.OutOfMemoryError: Java heap space Exception in thread "Thread-848" java.lang.OutOfMemoryError: Java heap space Exception in thread "Thread-863" java.lang.OutOfMemoryError: Java heap space Exception in thread "Thread-818" java.lang.OutOfMemoryError: Java heap space Exception in thread "Thread-851" java.lang.OutOfMemoryError: Java heap space: failed reallocation of scalar replaced objects Exception in thread "Thread-847" java.lang.OutOfMemoryError: Java heap space ERROR: Cannot throw exception because the return value of "rnabloom.RNABloom$LongReadCorrectionWorker.getExceptionCaught()" is null java.lang.NullPointerException: Cannot throw exception because the return value of "rnabloom.RNABloom$LongReadCorrectionWorker.getExceptionCaught()" is null at rnabloom.RNABloom.correctLongReadsMultithreaded(RNABloom.java:4006) at rnabloom.RNABloom.correctLongReads(RNABloom.java:5266) at rnabloom.RNABloom.main(RNABloom.java:7346)

[kmnip]

For the first run, the Racon command was killed for some unknown reason. I don't think it has to with memory or the -k or -e options.

The second run with multiple input files ran out of Java heap space. As mentioned previously, you can increase the max Java heap space if you have enough RAM.