allVHitsWithScore allDHitsWithScore allJHitsWithScore allCHitsWithScore

[2061574124]

i run the code just as following:
mixcr align -Xmx15g -p rna-seq -s hsa -OallowPartialAlignments=true ./f1.rmrrna.fastp.fastq.1.gz ./f1.rmrrna.fastp.fastq.2.gz ./f1.vdjca;
mixcr assemblePartial -Xmx15g ./f1.vdjca ./f1.alignments_rescued_1.vdjca
mixcr assemblePartial -Xmx15g ./f1.alignments_rescued_1.vdjca ./f1.alignments_rescued_2.vdjca
mixcr extend -Xmx15g ./f1.alignments_rescued_2.vdjca ./f1.alignments_rescued_2_extended.vdjca
mixcr assemble -Xmx15g ./f1.alignments_rescued_2_extended.vdjca ./f1.clones.clns
mixcr exportClones -Xmx15g ./f1.clones.clns ./f1.clones.txt; done
mixcr exportClones -Xmx15g -c IGH ./f1.clones.clns ./f1.IGH.clones.txt

After that, f1.IGH.clones.txt results shown that some reads could mapping to multi-VDJ. is this is normal？

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cloneId	cloneCount	cloneFraction	targetSequences	targetQualities	allVHitsWithScore	allDHitsWithScore	allJHitsWithScore	allCHitsWithScore	allVAlignments	allDAlignments	allJAlignments	allCAlignments	nSeqFR1	minQualFR1	nSeqCDR1	minQualCDR1	nSeqFR2	minQualFR2	nSeqCDR2	minQualCDR2	nSeqFR3	minQualFR3	nSeqCDR3	minQualCDR3	nSeqFR4	minQualFR4	aaSeqFR1	aaSeqCDR1	aaSeqFR2	aaSeqCDR2	aaSeqFR3	aaSeqCDR3	aaSeqFR4	refPoints
3	28	0.019270475	TGTGGATCAGATGCCAGAGCACCAGACGGCAATGGTTATCACTATGCTTTTGCTATGTGG	NNNNNNNNNNNNNNNNNNNN????????????????????????????????????????	IGHV1-69-2*00(249.2)	IGHD3-2200(39),IGHD3-300(35),IGHD5-18*00(35)	IGHJ3*00(156.5)	IGHG100(459),IGHG300(402.8)	359|371|390|0|12|SC363GSA365T|28.0	49|60|93|32|43|ST56C|39.0;49|56|93|32|39||35.0;32|39|60|31|38||35.0	22|39|70|43|60|SA31CSC35G|53.0	;											TGTGGATCAGATGCCAGAGCACCAGACGGCAATGGTTATCACTATGCTTTTGCTATGTGG	30								CGSDARAPDGNGYHYAFAMW		:::::::::0:1:12:32:-18:-2:43:43:-2:60:::
14	12	0.008258775	TGTGCGAGAGTCGGGTACCGTGGTGGTGCCACACCTTTTGACTACTGG	????????????????????????????????????????????????	IGHV4-39*00(337.6)	IGHD2-2100(45),IGHD2-1500(40)	IGHJ4*00(200)	IGHM*00(372.3)	445|454|476|0|9||45.0	36|45|84|19|28||45.0;42|50|93|19|27||40.0	25|37|68|36|48||60.0												TGTGCGAGAGTCGGGTACCGTGGTGGTGCCACACCTTTTGACTACTGG	30								CARVGYRGGATPFDYW		:::::::::0:-2:9:19:-8:-11:28:36:-5:48:::
33	8	0.00550585	TGTGCGCGACATCGGGCAGTCCCGGAGTTCGGGGAGGGACTTGACTATTGG	???????????????????????????????????????????????????	IGHV1-4600(73.5),IGHV3-5300(72.3)	IGHD3-10*00(50)	IGHJ4*00(174)	IGHG2*00(832.8)	442|455|473|0|13|SA448CSG451C|33.0;439|452|470|0|13|SA445CSG448C|33.0	42|52|93|26|36||50.0	26|37|68|40|51|SC33T|39.0												TGTGCGCGACATCGGGCAGTCCCGGAGTTCGGGGAGGGACTTGACTATTGG	30								CARHRAVPEFGEGLDYW		:::::::::0:2:13:26:-11:-10:36:40:-6:51:::
34	8	0.00550585	TGTGCAAGAGATTCCGGAAACATTTATGGCGCGTACTACTTTGATTCCTGG	???????????????????????????????????????????????????	IGHV6-1*00(340.4)	IGHD2-2100(35),IGHD2-1500(30),IGHD4-23*00(30)	IGHJ4*00(200)	IGHG2*00(369.4)	454|466|485|0|12||60.0	53|60|84|12|19||35.0;59|65|93|12|18||30.0;35|41|57|12|18||30.0	18|37|68|32|51|SC30TSA32C|63.0												TGTGCAAGAGATTCCGGAAACATTTATGGCGCGTACTACTTTGATTCCTGG	30								CARDSGNIYGAYYFDSW		:::::::::0:1:12:12:-25:4:19:32:2:51:::
38	7	0.004817619	TGTGCGAGAGAATCGACTGAGACATGGCTACAAGTGTCCAACTATTTTGAACACTGG	?????????????????????????????????????????????????????????	IGHV1-2*00(287.3)	IGHD5-24*00(54)	IGHJ400(162.8),IGHJ500(145.8)	IGHA1*00(527.5)	442|453|473|0|11||55.0	22|36|60|19|33|SG25C|54.0	20|37|68|40|57|SC24TSC30AST31C|37.0;36|40|71|53|57||20.0												TGTGCGAGAGAATCGACTGAGACATGGCTACAAGTGTCCAACTATTTTGAACACTGG	30								CARESTETWLQVSNYFEHW		:::::::::0:0:11:19:-2:-4:33:40:0:57:::

[mizraelson]

Yes, the BCR gene segments often have very similar sequences. If your reads do not cover the entire gene sequence, there might be several candidates. In the brackets following the gene names, you see the alignment scores. The first gene in the list is the most probable one with the highest score.

[2061574124]

Thank you! May I ask if it is possible to add any parameters during analysis that strictly limit the acquisition of only one?

[2061574124]

Or what parameters can be added to only retain the highest score gene when exporting cloned. txt from the already generated .vdjca or .clns file? And if possib, merge genes with high similarity

[mizraelson]

You can add mixcr exportClones -vHit -dHit -jHit -cHit ... to add columns with the best score gene.

[2061574124]

You can add mixcr exportClones -vHit -dHit -jHit -cHit ... to add columns with the best score gene.

Thank you very much for your patience