Error when running findalleles

[fookloy]

Hi guys, i came across the following errors after running cat sample.txt | while read id; do ( nohup mixcr findAlleles ${id}_assemble.clns --output-template {id}_allelic.clns --export-alleles-mutations {id}_allelestats.tsv & ); done with 4.5.0 version of mixcr, and the *assemble.clns were output by mixcr assemble --assemble-clonotypes-by "{FR1Begin:FR4End}" -ObadQualityThreshold=0 ${id}_align.vdjca ${id}_assemble.clns .

Error on searching alleles for IGKV1-9*00
Please copy the following information along with the stacktrace:
   Version: 4.5.0; built=Fri Sep 22 20:39:05 CST 2023; rev=cdb24b4fb7; lib=repseqio.v3.0.1
        OS: Linux
      Java: 11.0.21
  Cmd args: findAlleles H-HK_L2_assemble.clns --output-template H-HK_L2_allelic.clns --export-alleles-mutations H-HK_L2_allelestats.tsv
picocli.CommandLine$ExecutionException: Error while running command findAlleles java.lang.IllegalArgumentException
        at com.milaboratory.mixcr.cli.Main.registerExceptionHandlers$lambda-12(SourceFile:340)
        at picocli.CommandLine.execute(CommandLine.java:2088)
        at com.milaboratory.mixcr.cli.Main.main(SourceFile:98)
Caused by: java.lang.IllegalArgumentException
        at com.milaboratory.core.mutations.Mutations.combineWith(SourceFile:353)
        at com.milaboratory.o.gQ.a(SourceFile:26908)
        at com.milaboratory.o.gQ.a(SourceFile:511)
        at com.milaboratory.o.gQ.a(SourceFile:472)
        at com.milaboratory.o.gQ.a(SourceFile:30213)
        at com.milaboratory.o.hz.process(SourceFile:66)
        at cc.redberry.pipe.blocks.ParallelProcessor$Worker.run(ParallelProcessor.java:308)
        at java.base/java.lang.Thread.run(Thread.java:829)

Step 6 of 7: removing unnecessary genes: 99.7%  ETA: 00:00:00
Please copy the following information along with the stacktrace:
   Version: 4.5.0; built=Fri Sep 22 20:39:05 CST 2023; rev=cdb24b4fb7; lib=repseqio.v3.0.1
        OS: Linux
      Java: 11.0.21
  Cmd args: findAlleles HK-HK_L1_assemble.clns --output-template HK-HK_L1_allelic.clns --export-alleles-mutations HK-HK_L1_allelestats.tsv
picocli.CommandLine$ExecutionException: Error while running command findAlleles java.lang.IllegalStateException: Check failed.
        at com.milaboratory.mixcr.cli.Main.registerExceptionHandlers$lambda-12(SourceFile:340)
        at picocli.CommandLine.execute(CommandLine.java:2088)
        at com.milaboratory.mixcr.cli.Main.main(SourceFile:98)
Caused by: java.lang.IllegalStateException: Check failed.
        at com.milaboratory.mixcr.alleles.AlleleSearchResult.<init>(SourceFile:25)
        at com.milaboratory.mixcr.alleles.AlleleSearchResult.a(SourceFile:1000)
        at com.milaboratory.o.gQ.a(SourceFile:407)
        at com.milaboratory.mixcr.cli.CommandFindAlleles.run1(SourceFile:391)
        at com.milaboratory.mixcr.cli.MiXCRCommandWithOutputs.run0(SourceFile:69)
        at com.milaboratory.mixcr.cli.MiXCRCommand.run(SourceFile:37)
        at picocli.CommandLine.executeUserObject(CommandLine.java:1939)
        at picocli.CommandLine.access$1300(CommandLine.java:145)
        at picocli.CommandLine$RunLast.executeUserObjectOfLastSubcommandWithSameParent(CommandLine.java:2358)
        at picocli.CommandLine$RunLast.handle(CommandLine.java:2352)
        at picocli.CommandLine$RunLast.handle(CommandLine.java:2314)
        at picocli.CommandLine$AbstractParseResultHandler.execute(CommandLine.java:2179)
        at picocli.CommandLine$RunLast.execute(CommandLine.java:2316)
        at com.milaboratory.mixcr.cli.Main.registerLogger$lambda-26(SourceFile:447)
        at picocli.CommandLine.execute(CommandLine.java:2078)
        ... 1 more

For findalleles, 5 of 8 *allelic.clns files were completed, but the rest of them came across the problems up there i showed, it seems i met the same error in discussions/1286, but for different reason, any solution or advice? or could i just skip the error using any command? Thanks for help!

[mizraelson]

Hi, so the first error should be fixed in the latest develop version. Please give it a try and let me know. Also if the error is still there please try adding --dont-remove-unused-genes.

The second one is hard to debug without looking at the original data. You can try -OsearchMutationsInCDR3=null parameter, but then alleles search will skip variants in the CDR3 part (the very end of V gene). If you can share the raw data for this sample, I will try to give a more deeper insight.

[fookloy]

Sure, cat sample.txt | while read id; do ( nohup mixcr findAlleles ${id}_assemble.clns --output-template {id}_allelic.clns --export-alleles-mutations {id}_allelestats.tsv & ); done, and the *assemble.clns were output by mixcr assemble --assemble-clonotypes-by "{FR1Begin:FR4End}" -ObadQualityThreshold=0 ${id}_align.vdjca ${id}_assemble.clns .

[mizraelson]

What command was used to obtain .vdjca files from raw FASTQ files that you have shared?

[fookloy]

HI, the code is below.
java --jar mixcr.jar align --preset generic-amplicon --species human --rna --rigid-left-alignment-boundary --rigid-right-alignment-boundary C -OmergerParameters.minimalOverlap=10 -OmergerParameters.minimalIdentity=0.9 ${id}_1.fq.gz ${id}_2.fq.gz ${id}_align.vdjca

[mizraelson]

Please try the latest develop version. The issue should be fixed.

[fookloy]

It really works! Thanks for your help!