Export¶

In order to export alignment results or clones from a binary file (.vdjca or .clns) to a human-readable text file one can use the exportAlignments and exportClones commands respectively. The syntax for these commands is:

mixcr exportAlignments [options] alignments.vdjca alignments.txt

mixcr exportClones [options] clones.clns clones.txt

The resulting tab-delimited text file will contain columns with different types of information. If no options are specified, the default set of columns - which is sufficient in most cases - will be exported. The possible columns include (see below for details): aligned sequences, qualities, all or just best hit for V, D, J and C genes, corresponding alignments, nucleotide and amino acid sequences of gene region present in sequence, etc. When exporting clones, the additional columns include: clone count, clone fraction etc.

One can customize the list of fields that will be exported by passing parameters to export commands. For example, in order to export just clone count, best hits for V and J genes with corresponding alignments and CDR3 amino acid sequence, one can do:

mixcr exportClones -count -vHit -jHit -vAlignment -jAlignment -aaFeature CDR3 clones.clns clones.txt

The columns in the resulting file will be exported in exactly the same order as parameters on the command line. The list of available fields will be reviewed in the next subsections. For convenience, MiXCR provides two predefined sets of fields for exporting: min (will export minimal required information about clones or alignments) and full (used by default); one can use these sets by specifying the --preset option:

mixcr exportClones --preset min clones.clns clones.txt

One can add additional columns to the preset in the following way:

mixcr exportClones --preset min -qFeature CDR2 clones.clns clones.txt

One can also put all specify export fields in a seperate file:

-vHits
-dHits
-feature CDR3
...

and pass this file to the export command:

mixcr exportClones --preset-file myFields.txt clones.clns clones.txt

Command line parameters¶

The following is a list of command line parameters for both exportAlignments and exportClones:

Option	Description
`-h`, `--help`	print help message
`-f`, `--fields`	list available fields that can be exported
`-p`, `--preset`	select a predefined set of fields to export (`full` or `min`)
`-pf`, `--preset-file`	load a file with a list of fields to export
`-lf`, `--list-fields`	list available fields that can be exported
`-s`, `--no-spaces`	output short versions of column headers to assist with analysis using Pandas, R/DataFrames or another data tables processing library

The following parameters only apply to exportClones:

`-c`, `--chains`	Limit output to specific locus (e.g. TRA or IGH). Clone fractions will be recalculated accordingly.
`-o`, `--filter-out-of-frames`	Exclude out of frames (fractions will be recalculated)
`-t`, `--filter-stops`	Exclude sequences containing stop codons (fractions will be recalculated)
`-m`, `--minimal-clone-count`	Filter clones by minimal read count.
`-q`, `--minimal-clone-fraction`	Filter clones by minimal clone fraction.

Available fields¶

The following fields can be exported both for alignments and clones:

Field name	Description
`-chains`	Chains
`-topChains`	Chains of top hits
`-targets`	Number of targets
`-vHit`	Best V hit
`-dHit`	Best D hit
`-jHit`	Best J hit
`-cHit`	Best C hit
`-vGene`	Best V hit gene name (e.g. TRBV12-3 for TRBV12-3*00)
`-dGene`	Best D hit gene name (e.g. TRBV12-3 for TRBV12-3*00)
`-jGene`	Best J hit gene name (e.g. TRBV12-3 for TRBV12-3*00)
`-cGene`	Best C hit gene name (e.g. TRBV12-3 for TRBV12-3*00)
`-vFamily`	Best V hit family name (e.g. TRBV12 for TRBV12-3*00)
`-dFamily`	Best D hit family name (e.g. TRBV12 for TRBV12-3*00)
`-jFamily`	Best J hit family name (e.g. TRBV12 for TRBV12-3*00)
`-cFamily`	Best C hit family name (e.g. TRBV12 for TRBV12-3*00)
`-vHitScore`	Score for best V hit
`-dHitScore`	Score for best D hit
`-jHitScore`	Score for best J hit
`-cHitScore`	Score for best C hit
`-vHitsWithScore`	All V hits with score
`-dHitsWithScore`	All D hits with score
`-jHitsWithScore`	All J hits with score
`-cHitsWithScore`	All C hits with score
`-vHits`	All V hits
`-dHits`	All D hits
`-jHits`	All J hits
`-cHits`	All C hits
`-vGenes`	All V gene names (e.g. TRBV12-3 for TRBV12-3*00)
`-dGenes`	All D gene names (e.g. TRBV12-3 for TRBV12-3*00)
`-jGenes`	All J gene names (e.g. TRBV12-3 for TRBV12-3*00)
`-cGenes`	All C gene names (e.g. TRBV12-3 for TRBV12-3*00)
`-vFamilies`	All V gene family anmes (e.g. TRBV12 for TRBV12-3*00)
`-dFamilies`	All D gene family anmes (e.g. TRBV12 for TRBV12-3*00)
`-jFamilies`	All J gene family anmes (e.g. TRBV12 for TRBV12-3*00)
`-cFamilies`	All C gene family anmes (e.g. TRBV12 for TRBV12-3*00)
`-vAlignment`	Best V alignment
`-dAlignment`	Best D alignment
`-jAlignment`	Best J alignment
`-cAlignment`	Best C alignment
`-vAlignments`	All V alignments
`-dAlignments`	All D alignments
`-jAlignments`	All J alignments
`-cAlignments`	All C alignments
`-nFeature` `<gene_feature>`	Nucleotide sequence of specified gene feature
`-qFeature` `<gene_feature>`	Quality string of specified gene feature
`-aaFeature` `<gene_feature>`	Amino acid sequence of specified gene feature
`-minFeatureQuality` `<gene_feature>`	Minimal quality of specified gene feature
`-avrgFeatureQuality` `<gene_feature>`	Average quality of specified gene feature
`-lengthOf` `<gene_feature>`	S length of specified gene feature.
`-nMutations` `<gene_feature>`	Extract nucleotide mutations for specific gene feature; relative to germline sequence.
`-nMutationsRelative` `<gene_feature>` `<relative_to_gene_feature>`	Extract nucleotide mutations for specific gene feature relative to another feature.
`-aaMutations` `<gene_feature>`	Extract amino acid mutations for specific gene feature
`-aaMutationsRelative` `<gene_feature>` `<relative_to_gene_feature>`	Extract amino acid mutations for specific gene feature relative to another feature.
`-mutationsDetailed` `<gene_feature>`	Detailed list of nucleotide and corresponding amino acid mutations. Format <nt_mutation>:<aa_mutation_individual>:<aa_mutation_cumulative>, where <aa_mutation_individual> is an expected amino acid mutation given no other mutations have occurred, and <aa_mutation_cumulative> amino acid mutation is the observed amino acid mutation combining effect from all other
`-mutationsDetailedRelative` `<gene_feature>` `<relative_to_gene_feature>`	Detailed list of nucleotide and corresponding amino acid mutations written, positions relative to specified gene feature. Format <nt_mutation>:<aa_mutation_individual>:<aa_mutation_cumulative>, where <aa_mutation_individual> is an expected amino acid mutation given no other mutations have occurred, and <aa_mutation_cumulative> amino acid mutation is the observed amino acid mutation combining effect from all other
`-positionOf` `<reference_point>`	S position of specified reference point inside target sequences (clonal sequence / read sequence).
`-defaultAnchorPoints`	Outputs a list of default reference points (like CDR2Begin, FR4End, etc. see documentation for the full list and formatting)
`-vIdentityPercents`	V alignment identity percents
`-dIdentityPercents`	D alignment identity percents
`-jIdentityPercents`	J alignment identity percents
`-cIdentityPercents`	C alignment identity percents
`-vBestIdentityPercent`	Vbest alignment identity percent
`-dBestIdentityPercent`	Dbest alignment identity percent
`-jBestIdentityPercent`	Jbest alignment identity percent
`-cBestIdentityPercent`	Cbest alignment identity percent

The following fields are specific for alignments:

Field name	Description
`-readId`	Id of read corresponding to alignment
`-sequence`	Aligned sequence (initial read), or 2 sequences in case of paired-end reads
`-quality`	Initial read quality, or 2 qualities in case of paired-end reads
`-descrR1`	Description line from initial .fasta or .fastq file of the first read (only available if –save-description was used in align command)
`-descrR2`	Description line from initial .fasta or .fastq file of the second read (only available if –save-description was used in align command)
`-cloneId` `<index_file>`	To which clone alignment was attached.
`-cloneIdWithMappingType` `<index_file>`	To which clone alignment was attached with additional info on mapping type.

The following fields are specific for clones:

Field name	Description
`-cloneId`	Unique clone identifier
`-count`	Clone count
`-fraction`	Clone fraction
`-sequence`	Aligned sequence (initial read), or 2 sequences in case of paired-end reads
`-quality`	Initial read quality, or 2 qualities in case of paired-end reads
`-readIds` `<index_file>`	Read IDs aggregated by clone.

See this chapter for the translation rules used for options like: -aaFeature.

Default anchor point positions¶

Positions of anchor points produced by the -defaultAnchorPoints option are outputted as a colon separated list. If an anchor point is not covered by the target sequence nothing is printed for it, but flanking colon symbols are preserved to maintain positions in array. See example:

:::::::::108:117:125:152:186:213:243:244:

If there are several target sequences (e.g. paired-end reads or multi-part clonal sequnce), an array is outputted for each target sequence. In this case arrays are separated by a comma:

2:61:107:107:118:::::::::::::,:::::::::103:112:120:147:181:208:238:239:

Even if there are no anchor points in one of the parts:

:::::::::::::::::,:::::::::108:117:125:152:186:213:243:244:

The following table shows the correspondence between anchor points and positions in the default anchor point array:

Anchors point	Zero-based position	One-based position
V5UTRBeginTrimmed	0	1
V5UTREnd / L1Begin	1	2
L1End / VIntronBegin	2	3
VIntronEnd / L2Begin	3	4
L2End / FR1Begin	4	5
FR1End / CDR1Begin	5	6
CDR1End / FR2Begin	6	7
FR2End / CDR2Begin	7	8
CDR2End / FR3Begin	8	9
FR3End / CDR3Begin	9	10
Number of 3’ V deletions (negative value), or length of 3’ V P-segment (positive value)	10	11
VEndTrimmed, next position after last aligned nucleotide of V gene	11	12
DBeginTrimmed, position of first aligned nucleotide of D gene	12	13
Number of 5’ D deletions (negative value), or length of 5’ D P-segment (positive value)	13	14
Number of 3’ D deletions (negative value), or length of 3’ D P-segment (positive value)	14	15
DEndTrimmed, next position after last aligned nucleotide of D gene	15	16
JBeginTrimmed, position of first aligned nucleotide of J gene	16	17
Number of 3’ J deletions (negative value), or length of 3’ J P-segment (positive value)	17	18
CDR3End / FR4Begin	18	19
FR4End	19	20
CBegin	20	21
CExon1End	21	22

The following regular expressions can be used to parse the contents of this field in Python:

for length analysis, or analysis of raw alignments:

^(?P<V5UTRBegin>-?[0-9]*):(?P<L1Begin>-?[0-9]*):(?P<VIntronBegin>-?[0-9]*):(?P<L2Begin>-?[0-9]*):(?P<FR1Begin>-?[0-9]*):(?P<CDR1Begin>-?[0-9]*):(?P<FR2Begin>-?[0-9]*):(?P<CDR2Begin>-?[0-9]*):(?P<FR3Begin>-?[0-9]*):(?P<CDR3Begin>-?[0-9]*):(?P<V3Deletion>-?[0-9]*):(?P<VEnd>-?[0-9]*):(?P<DBegin>-?[0-9]*):(?P<D5Deletion>-?[0-9]*):(?P<D3Deletion>-?[0-9]*):(?P<DEnd>-?[0-9]*):(?P<JBegin>-?[0-9]*):(?P<J5Deletion>-?[0-9]*):(?P<CDR3End>-?[0-9]*):(?P<CBegin>-?[0-9]*):(?P<CExon1End>-?[0-9]*)$

snipped for Pandas:

import pandas as pd
data = pd.read_table("exported.txt", low_memory=False)
anchorPointsRegex="^(?P<V5UTRBegin>-?[0-9]*):(?P<L1Begin>-?[0-9]*):(?P<VIntronBegin>-?[0-9]*):(?P<L2Begin>-?[0-9]*):(?P<FR1Begin>-?[0-9]*):(?P<CDR1Begin>-?[0-9]*):(?P<FR2Begin>-?[0-9]*):(?P<CDR2Begin>-?[0-9]*):(?P<FR3Begin>-?[0-9]*):(?P<CDR3Begin>-?[0-9]*):(?P<V3Deletion>-?[0-9]*):(?P<VEnd>-?[0-9]*):(?P<DBegin>-?[0-9]*):(?P<D5Deletion>-?[0-9]*):(?P<D3Deletion>-?[0-9]*):(?P<DEnd>-?[0-9]*):(?P<JBegin>-?[0-9]*):(?P<J5Deletion>-?[0-9]*):(?P<CDR3End>-?[0-9]*):(?P<CBegin>-?[0-9]*):(?P<CExon1End>-?[0-9]*)$"
data = pd.concat([data, d.refPoints.str.extract(anchorPointsRegex, expand=True).apply(pd.to_numeric)], axis=1)

A simplified regular expression with a smaller number of fields can be used for analysis of CDR3-assembled clonotypes:

^(?:-?[0-9]*:){8}(?:-?[0-9]*):(?P<CDR3Begin>-?[0-9]*):(?P<V3Deletion>-?[0-9]*):(?P<VEnd>-?[0-9]*):(?P<DBegin>-?[0-9]*):(?P<D5Deletion>-?[0-9]*):(?P<D3Deletion>-?[0-9]*):(?P<DEnd>-?[0-9]*):(?P<JBegin>-?[0-9]*):(?P<J5Deletion>-?[0-9]*):(?P<CDR3End>-?[0-9]*):(?:-?[0-9]*:){2}(?:-?[0-9]*)$

snipped for Pandas:

import pandas as pd
data = pd.read_table("exported.txt", low_memory=False)
anchorPointsRegex="^^(?:-?[0-9]*:){8}(?:-?[0-9]*):(?P<CDR3Begin>-?[0-9]*):(?P<V3Deletion>-?[0-9]*):(?P<VEnd>-?[0-9]*):(?P<DBegin>-?[0-9]*):(?P<D5Deletion>-?[0-9]*):(?P<D3Deletion>-?[0-9]*):(?P<DEnd>-?[0-9]*):(?P<JBegin>-?[0-9]*):(?P<J5Deletion>-?[0-9]*):(?P<CDR3End>-?[0-9]*):(?:-?[0-9]*:){2}(?:-?[0-9]*)$"
data = pd.concat([data, d.refPoints.str.extract(anchorPointsRegex, expand=True).apply(pd.to_numeric)], axis=1)

Examples¶

Export only the best V, D, J hits and the best V hit alignment from a .vdjca file:

mixcr exportAlignments -vHit -dHit -jHit -vAlignment input.vdjca test.txt

Best V hit	Best D hit	Best J hit	Best V alignment
IGHV4-34*00		IGHJ4*00	`\|262\|452\|453\|47\|237\|SC268GSC271ASC275G\|956.1,58\|303\|450\|` `56\|301\|SG72TSA73CSG136TSA144CSA158CSG171T\|331.0\|`
IGHV2-23*00	IGHD2*21	IGHJ6*00	`\|262\|452\|453\|47\|237\|SC268GSC271ASC275G\|956.1,58\|303\|450\|` `56\|301\|SG72TSA73CSG136TSA144CSA158CSG171T\|331.0\|`

The syntax of alignment is described in appendix.

Exporting well formatted alignments for manual inspection¶

MiXCR is able to export alignments create with the align step as pretty formatted text (human readable) for manual analysis. This can be used both to inspect alignments and to facilitate optimization of analysis parameters and library preparation protocol. To export pretty formatted alignments use the exportAlignmentsPretty command:

mixcr exportAlignmentsPretty --skip 1000 --limit 10 input.vdjca test.txt

this will export 10 results after skipping the first 1000 records, then place the results into the file test.txt. Skipping earlier records is often useful because the first sequences in a fastq file may have lower than average read quality. Omitting the last parameter (output file name) will print results directly to the standard output stream (to console), like this:

mixcr exportAlignmentsPretty --skip 1000 --limit 10 input.vdjca

Here is a summary of the command line options:

Option	Description
`-h`, `--help`	print help message
`-n`, `--limit`	limit number of alignments; no more than provided number of results will be outputted
`-s`, `--skip`	number of results to skip
`-t`, `--top`	output only top hits for V, D, J nad C genes
`--cdr3-contains`	output only those alignments in which CDR3 contains specified nucleotides (e.g. `--cdr3-contains TTCAGAGGAGC`)
`--read-contains`	output only those alignments for which the corresonding reads contain specified nucleotides e.g. `--read-contains ATGCTTGCGCGCT`)
`--verbose`	use a more verbose format for alignments (see below for example)

Results produced by this command have the following structure:


  >>> Read id: 1

                                                      5'UTR><L1
   Quality    88888888888888888888888887888888888888888888888888888888888888888888888887888878
   Target0  0 AAGGCCTTTCCACTTGGTGATCAGCACTGAGCACAGAGGACTCACCATGGAGTTGGGGCTGAGCTGGGTTTTCCTTGTTG 79
IGHV3-7*00 54 aaggcctttccacttggtgatcagcactgagcacagaggactcaccatggaAttggggctgagctgggttttccttgttg 133

                        L1><L2     L2><FR1
   Quality     88888888887888888888888888888889989989989889999997999999989999999999999999999899
   Target0  80 CTATTTTAGAAGGTGTCCAGTGTGAGGTGAAGTTGGTGGAGTCTGGGGGAGGCCTGGTCCAGCCTGGGGGGTCCCTGAGA 159
IGHV3-7*00 134 ctattttagaaggtgtccagtgtgaggtgCagCtggtggagtctgggggaggcTtggtccagcctggggggtccctgaga 213

                             FR1><CDR1              CDR1><FR2
   Quality     999999999999999999999999999999999999999999999 9999999999999999999999999999999999
   Target0 160 CTCTCCTGTGAAGCCTCCGGATTCACCTTTAGTAGTTATTGGATG-GCATGGGTCCGCCAGGGTCCAGGGCAGGGGCTGG 238
IGHV3-7*00 214 ctctcctgtgCagcctcTggattcacctttagtagCtattggatgAgc-tgggtccgccaggCtccagggAaggggctgg 292

                         FR2><CDR2              CDR2><FR3
   Quality     99999999999999999999999999999999999799999999999999999999999999998999899898999999
   Target0 239 AATGGGTGGGCAACATAAGGCCGGATGGAAGTGAGAGTTGGTACTTGGAGTCTGTGATGGGGCGATTCATGATATCTAGA 318
IGHV3-7*00 293 aGtgggtggCcaacataaAgcAAgatggaagtgagaAAtACtaTGtggaCtctgtgaAgggCcgattcaCCatCtcCaga 372

                                                                                 FR3><CDR3
    Quality     99899899999999988989999889979988888888878878788888888878888888778788888888878888
    Target0 319 GACAACGCCAAGAAGTCACTTTATCTGCAAATGGACAGCCTGAGAGTCGAGGACACGGCCGTCTATTATTGTGCGACTTC 398
 IGHV3-7*00 373 gacaacgccaagaaCtcactGtatctgcaaatgAacagcctgagagCcgaggacacggcTgtGtattaCtgtgcga     448
IGHD3-10*00  12                                                                              ttc 14

                                 CDR3><FR4
    Quality     88888788888888888888888787788777887787777877777877787787877878788788777767778788
    Target0 399 GGAGGAGCCGGAGGACTACTGGGGCCAGGGAGCCCTGGTCACCGTCTCCTCGGCTTCCACCAAGGGCCCATCGGTCTTCC 478
IGHD3-10*00  15 gg-ggag                                                                          20
   IGHJ4*00   8              gactactggggccagggaAccctggtcaccgtctcctc                              45
   IGHG4*00   0                                                      cttccaccaagggcccatcggtcttcc 26
   IGHG3*00   0                                                      cttccaccaagggcccatcggtcttcc 26
   IGHG2*00   0                                                      cCtccaccaagggcccatcggtcttcc 26
   IGHG1*00   0                                                      cCtccaccaagggcccatcggtcttcc 26
   IGHGP*00 194                                                    AgcCtccaccaagggcccatcggtcttcc 222


 Quality     87370
 Target0 479 CCTTG 483
IGHG4*00  27 ccCtg 31
IGHG3*00  27 ccCtg 31
IGHG2*00  27 ccCtg 31
IGHG1*00  27 ccCtg 31
IGHGP*00 223 ccCtg 227

Usage of the --verbose option will produce alignments in a slightly different format:

>>> Read id: 12343    <--- Index of analysed read in input file

>>> Target sequences (input sequences):

Sequence0:   <--- Read 1 from paired-end read
Contains features: CDR1, VRegionTrimmed, L2, L, Intron, VLIntronL, FR1, Exon1,              <--- Gene features
VExon2Trimmed                                                                                    found in read 1

     0 TCTTGGGGGATTCGGTGATCAGCACTGAACACAGAGGACTCACCATGGAGTTTGGGCTGAACTGGGTTTTCCTCGTTGCT 79  <--- Sequyence & quality 
       FGGEGGGGGDG8F78CFC6CEFF<,CFG9EED,6,CFCC<EEGFG,CE:CCAFFGGC87CEF?A?FBC@FGGFG>B,FC9          of read 1

    80 CTATTAAGAGGTGTCCAGTGTCAGGTGCAGCTGGTGGAGTCTGGGGGTGGCGTGTTCCAGCCTGGGGGGTCCGTGAGACT 159
       F9,A,95AFE,B?,E,C,9AC<FGA<EE5??,A,A<:=:E,=B8C7+++8,++@+,885=D7:@8E+:5*1**11**++<
   160 CTCCTGTGCAGCGTCGGGATGCACATCATGGAGCTATGGCCAGCCCTGGGTACGCCAGGCTACAGGCCACGGGCTGGAGG 239
       <++*++0++2A:ECE5EC5**2@C+:++++++22*2:+29+*2***25/79*0299))*/)*0*0*.75)7:)1)1/)))

   240 GGGTGCGTGGTAGATGGGAA 259
       )9:.)))*1)12***-/).)

Sequence1:   <--- Read 2 from paired-end read
Contains features: JCDR3Part, DCDR3Part, DJJunction, CDR2, JRegionTrimmed, CDR3, VDJunction,
VJJunction, VCDR3Part, ShortCDR3, FR4, FR3

     0 CGAGGCAAGAGGCTGGTGTGGGTGGCGGTTATATGGTATGGTGGAAGTAATAAACACTATGCAGACCCCGTGAAGGGCCG 79
       **0*0**)2**/**5D7<15*9<5:1+*0:GF:=C>6A52++*:2+++FF>>3<++++++302**:**/<+**;:/**2+

    80 ATTCACCATCGCCAGAGACAATTCCAAGAACACGCTGTATCTGCAAATGAAGAGCCTGAGAGCCGAGGACACGGCTTTGT 159
       +++<0***C:2+9GGFB?,5,4,+,2F<>FC=*,,C:>,=,@,,;3<@=,3,,<3,CF?=**<>@,?3,<<:3,CC,E,@

   160 ATTACTGTGCGAGAGGTCAACAGGGTGACTATGTCTACGGTAGGGACGTCGGGGGCCAAGGGACCACGGTCACCGTCTCC 239
       ,@;FCF@+F@FGGF9FD,F>>+B:=,,=><GFCGGCFEGFF?+=B+7EF>+FFA,8F<E:,5+GDFFE,@F?,,7GGDFE

   240 TCAGGGAGTGCATCCGCCCCAACCCTTTTCCCCCTCTCTGCGTTGATACCACTGGCAGCTC 300
       C,FGGGEFCCGEEGGCFCC:8FGEGGGE@DFB-GFGGGGF@GFGFE<,GFCCFCAGC@CCC

>>> Gene features that can be extracted from this (paired-)read:                         <--- For paired-end reads
JCDR3Part, CDR1, VRegionTrimmed, L2, DCDR3Part, VDJTranscriptWithout5UTR, Exon2, L,           some gene features
DJJunction, Intron, FR2, CDR2, VDJRegion, JRegionTrimmed, CDR3, VDJunction, VJJunction,       can be extracted by
VLIntronL, FR1, VCDR3Part, ShortCDR3, Exon1, FR4, VExon2Trimmed, FR3                          merging sequence
                                                                                              information

>>> Alignments with V gene:

IGHV3-33*00 (total score = 1638.0) <--- Alignment of both reads with IGHV3-33
Alignment of Sequence0 (score = 899.0):   <--- Alignment of IGHV3-33 with read 1 from paired-end read
     65 ATTCGGTGATCAGCACTGAACACAGAGGACTCACCATGGAGTTTGGGCTGAGCTGGGTTTTCCTCGTTGCTCTTTTAAGA 144 <--- Germline
        ||||||||||||||||||||||||||||||||||||||||||||||||||| ||||||||||||||||||||| ||||||
      9 ATTCGGTGATCAGCACTGAACACAGAGGACTCACCATGGAGTTTGGGCTGAACTGGGTTTTCCTCGTTGCTCTATTAAGA 88  <--- Read
        DG8F78CFC6CEFF<,CFG9EED,6,CFCC<EEGFG,CE:CCAFFGGC87CEF?A?FBC@FGGFG>B,FC9F9,A,95AF     <--- Quality score

    145 GGTGTCCAGTGTCAGGTGCAGCTGGTGGAGTCTGGGGGAGGCGTGGTCCAGCCTGGGAGGTCCCTGAGACTCTCCTGTGC 224
        |||||||||||||||||||||||||||||||||||||| |||||| ||||||||||| ||||| ||||||||||||||||
     89 GGTGTCCAGTGTCAGGTGCAGCTGGTGGAGTCTGGGGGTGGCGTGTTCCAGCCTGGGGGGTCCGTGAGACTCTCCTGTGC 168
        E,B?,E,C,9AC<FGA<EE5??,A,A<:=:E,=B8C7+++8,++@+,885=D7:@8E+:5*1**11**++<<++*++0++

    225 AGCGTCTGGATTCACCTTCA-GTAGCTATGGCATGCACTGGGTCCGCCAGGCTCCAGGCAAGGGGCTGGAGTGGGTG 300
        |||||| |||| || | ||| | |||||||||  || |||||| ||||||||| ||||| | ||||||||| |||||
    169 AGCGTCGGGATGCA-CATCATGGAGCTATGGCCAGCCCTGGGTACGCCAGGCTACAGGCCACGGGCTGGAGGGGGTG 244
        2A:ECE5EC5**2@ C+:++++++22*2:+29+*2***25/79*0299))*/)*0*0*.75)7:)1)1/))))9:.)

Alignment of Sequence1 (score = 739.0):   <--- Alignment of IGHV3-33 with read 2 from paired-end read
    279 AGGCAAGGGGCTGGAGTGGGTGGCAGTTATATGGTATGATGGAAGTAATAAATACTATGCAGACTCCGTGAAGGGCCGAT 358
        ||||||| |||||| ||||||||| ||||||||||||| ||||||||||||| ||||||||||| |||||||||||||||
      2 AGGCAAGAGGCTGGTGTGGGTGGCGGTTATATGGTATGGTGGAAGTAATAAACACTATGCAGACCCCGTGAAGGGCCGAT 81
        0*0**)2**/**5D7<15*9<5:1+*0:GF:=C>6A52++*:2+++FF>>3<++++++302**:**/<+**;:/**2+++

    359 TCACCATCTCCAGAGACAATTCCAAGAACACGCTGTATCTGCAAATGAACAGCCTGAGAGCCGAGGACACGGCTGTGTAT 438
        |||||||| |||||||||||||||||||||||||||||||||||||||| |||||||||||||||||||||||| |||||
     82 TCACCATCGCCAGAGACAATTCCAAGAACACGCTGTATCTGCAAATGAAGAGCCTGAGAGCCGAGGACACGGCTTTGTAT 161
        +<0***C:2+9GGFB?,5,4,+,2F<>FC=*,,C:>,=,@,,;3<@=,3,,<3,CF?=**<>@,?3,<<:3,CC,E,@,@

    439 TACTGTGCGAGAG 451
        |||||||||||||
    162 TACTGTGCGAGAG 174
        ;FCF@+F@FGGF9

IGHV3-30*00 (total score = 1582.0)  <--- Alternative hit for V gene
Alignment of Sequence0 (score = 885.0):
     65 ATTCGGTGATCAGCACTGAACACAGAGGACTCACCATGGAGTTTGGGCTGAGCTGGGTTTTCCTCGTTGCTCTTTTAAGA 144
        ||||||||||||||||||||||||||||||||||||||||||||||||||| ||||||||||||||||||||| ||||||
      9 ATTCGGTGATCAGCACTGAACACAGAGGACTCACCATGGAGTTTGGGCTGAACTGGGTTTTCCTCGTTGCTCTATTAAGA 88
        DG8F78CFC6CEFF<,CFG9EED,6,CFCC<EEGFG,CE:CCAFFGGC87CEF?A?FBC@FGGFG>B,FC9F9,A,95AF

    145 GGTGTCCAGTGTCAGGTGCAGCTGGTGGAGTCTGGGGGAGGCGTGGTCCAGCCTGGGAGGTCCCTGAGACTCTCCTGTGC 224
        |||||||||||||||||||||||||||||||||||||| |||||| ||||||||||| ||||| ||||||||||||||||
     89 GGTGTCCAGTGTCAGGTGCAGCTGGTGGAGTCTGGGGGTGGCGTGTTCCAGCCTGGGGGGTCCGTGAGACTCTCCTGTGC 168
        E,B?,E,C,9AC<FGA<EE5??,A,A<:=:E,=B8C7+++8,++@+,885=D7:@8E+:5*1**11**++<<++*++0++

    225 AGCCTCTGGATTCACCTTCA-GTAGCTATGGCATGCACTGGGTCCGCCAGGCTCCAGGCAAGGGGCTGGAGTGGGTG 300
        ||| || |||| || | ||| | |||||||||  || |||||| ||||||||| ||||| | ||||||||| |||||
    169 AGCGTCGGGATGCA-CATCATGGAGCTATGGCCAGCCCTGGGTACGCCAGGCTACAGGCCACGGGCTGGAGGGGGTG 244
        2A:ECE5EC5**2@ C+:++++++22*2:+29+*2***25/79*0299))*/)*0*0*.75)7:)1)1/))))9:.)

Alignment of Sequence1 (score = 697.0):
    279 AGGCAAGGGGCTGGAGTGGGTGGCAGTTATATCATATGATGGAAGTAATAAATACTATGCAGACTCCGTGAAGGGCCGAT 358
        ||||||| |||||| ||||||||| |||||||  |||| ||||||||||||| ||||||||||| |||||||||||||||
      2 AGGCAAGAGGCTGGTGTGGGTGGCGGTTATATGGTATGGTGGAAGTAATAAACACTATGCAGACCCCGTGAAGGGCCGAT 81
        0*0**)2**/**5D7<15*9<5:1+*0:GF:=C>6A52++*:2+++FF>>3<++++++302**:**/<+**;:/**2+++

    359 TCACCATCTCCAGAGACAATTCCAAGAACACGCTGTATCTGCAAATGAACAGCCTGAGAGCTGAGGACACGGCTGTGTAT 438
        |||||||| |||||||||||||||||||||||||||||||||||||||| ||||||||||| |||||||||||| |||||
     82 TCACCATCGCCAGAGACAATTCCAAGAACACGCTGTATCTGCAAATGAAGAGCCTGAGAGCCGAGGACACGGCTTTGTAT 161
        +<0***C:2+9GGFB?,5,4,+,2F<>FC=*,,C:>,=,@,,;3<@=,3,,<3,CF?=**<>@,?3,<<:3,CC,E,@,@

    439 TACTGTGCGAGAG 451
        |||||||||||||
    162 TACTGTGCGAGAG 174
        ;FCF@+F@FGGF9

>>> Alignments with D gene:

IGHD4-17*00 (total score = 40.0)
Alignment of Sequence1 (score = 40.0):
      7 GGTGACTA 14
        ||||||||
    183 GGTGACTA 190
        :=,,=><G

IGHD4-23*00 (total score = 36.0)
Alignment of Sequence1 (score = 36.0):
      0 TGACTACGGT 9
        || |||||||
    191 TGTCTACGGT 200
        FCGGCFEGFF

IGHD2-21*00 (total score = 35.0)
Alignment of Sequence1 (score = 35.0):
     13 GGTGACT 19
        |||||||
    183 GGTGACT 189
        :=,,=><

>>> Alignments with J gene:

IGHJ6*00 (total score = 172.0)
Alignment of Sequence1 (score = 172.0):
     22 GGACGTCTGGGGCAAAGGGACCACGGTCACCGTCTCCTCA 61
        ||||||| ||||| ||||||||||||||||||||||||||
    203 GGACGTCGGGGGCCAAGGGACCACGGTCACCGTCTCCTCA 242
        =B+7EF>+FFA,8F<E:,5+GDFFE,@F?,,7GGDFEC,F

>>> Alignments with C gene:

No hits.

Exporting reads aggregated by clones¶

MiXCR can preserve the mapping between initial reads and final clonotypes. There are several options for accessing this information.

To enable this behaviour, specify the option --index for the assemble command:

mixcr assemble --index index_file alignments.vdjca output.clns

This will tell MiXCR to store an index mapping in the file index_file (actually two files will be created: index_file and index_file.p both of which are used to store the index; in further options one should specify only index_file without .p extension and MiXCR will automatically read both required files). It is then possible to use this index_file in order to map clones to initial reads. For example using the -cloneId option for the exportAlignments command:

mixcr exportAlignments -p min -cloneId index_file alignments.vdjca alignments.txt

will print an additional column with the id of the clone which matches the corresponding alignment:

Best V hit	Best D hit	…	CloneId
IGHV4-34*00		…	321
IGHV2-23*00	IGHD2*21	…
IGHV4-34*00	IGHD2*21	…	22143
…	…	…	…

For more information it’s also possible to export the mapping type:

mixcr exportAlignments -p min -cloneIdWithMappingType index_file alignments.vdjca alignments.txt

Will give you something like:

Best V hit	Best D hit	…	Clone mapping
IGHV4-34*00		…	321:core
IGHV2-23*00	IGHD2*21	…	dropped
IGHV4-34*00	IGHD2*21	…	22143:clustered
IGHV4-34*00	IGHD2*21	…	23:mapped
…	…	…	…

It’s also possible to export all read IDs that were aggregated by each clone, using the -readIds export options for exportClones:

mixcr exportClones -c IGH -p min -readIds index_file clones.clns clones.txt

This will add a column with a full enumeration of all reads that were clustered into a particular clone:

Clone ID	Clone count	Best V hit	…	Reads
0	7213	IGHV4-34*00	…	56,74,92,96,101,119,169,183…
1	2951	IGHV2-23*00	…	46,145,194,226,382,451,464…
2	2269	IGHV4-34*00	…	58,85,90,103,113,116,122,123…
3	124	IGHV4-34*00	…	240,376,496,617,715,783,813…
…		…	…	…

Note, the resulting txt file may be _ very large _ since all read numbers that were successfully assembled will be printed.

Finally, you can export reads aggregated by clone into separate .fastq files. To support this behaviour the align command needs to be run with the additional -g option:

mixcr align -g input.fastq alignments.vdjca.gz

With this option MiXCR will store the original reads in the .vdjca file. Then one can export reads corresponding to a particular clone with the exportReadsForClones command. For example, export all reads that were assembled into the first clone (clone with cloneId = 0):

mixcr exportReadsForClones index_file alignments.vdjca.gz 0 reads.fastq.gz

This will create the file reads_clns0.fastq.gz (or two files reads_clns0_R1.fastq.gz and reads_clns0_R2.fastq.gz if the original data were paired) with all reads that were aggregated by the first clone. One can export reads for several clones at a time:

mixcr exportReadsForClones index_file alignments.vdjca.gz 0 1 2 33 54 reads.fastq.gz

This will create several files (reads_clns0.fastq.gz, reads_clns1.fastq.gz etc.) for each clone with cloneId equal to 0, 1, 2, 33 and 54 respectively.