groupGenes {alakazam}

Group sequences by gene assignment

Description

groupGenes will group rows by shared V and J gene assignments, and optionally also by junction lengths. IGH:IGK/IGL, TRB:TRA, and TRD:TRG paired single-cell BCR/TCR sequencing and unpaired bulk sequencing (IGH, TRB, TRD chain only) are supported. In the case of ambiguous (multiple) gene assignments, the grouping may be specified to be a union across all ambiguous V and J gene pairs, analogous to single-linkage clustering (i.e., allowing for chaining).

Usage

groupGenes(
  data,
  v_call = "v_call",
  j_call = "j_call",
  junc_len = NULL,
  cell_id = NULL,
  locus = "locus",
  only_heavy = TRUE,
  first = FALSE
)

Arguments

data

data.frame containing sequence data.

v_call

name of the column containing the heavy/long chain V-segment allele calls.

j_call

name of the column containing the heavy/long chain J-segment allele calls.

junc_len

name of column containing the junction length. If NULL then 1-stage partitioning is perform considering only the V and J genes is performed. See Details for further clarification.

cell_id

name of the column containing cell identifiers or barcodes. If specified, grouping will be performed in single-cell mode with the behavior governed by the locus and only_heavy arguments. If set to NULL then the bulk sequencing data is assumed.

locus

name of the column containing locus information. Only applicable to single-cell data. Ignored if cell_id=NULL.

only_heavy

use only the IGH (BCR) or TRB/TRD (TCR) sequences for grouping. Only applicable to single-cell data. Ignored if cell_id=NULL.

first

if TRUE only the first call of the gene assignments is used. if FALSE the union of ambiguous gene assignments is used to group all sequences with any overlapping gene calls.

Details

To invoke single-cell mode the cell_id argument must be specified and the locus column must be correct. Otherwise, groupGenes will be run with bulk sequencing assumptions, using all input sequences regardless of the values in the locus column.

Values in the locus column must be one of c("IGH", "IGI", "IGK", "IGL") for BCR or c("TRA", "TRB", "TRD", "TRG") for TCR sequences. Otherwise, the function returns an error message and stops.

Under single-cell mode with paired chained sequences, there is a choice of whether grouping should be done by (a) using IGH (BCR) or TRB/TRD (TCR) sequences only or (b) using IGH plus IGK/IGL (BCR) or TRB/TRD plus TRA/TRG (TCR). This is governed by the only_heavy argument.

Specifying junc_len will force groupGenes to perform a 1-stage partitioning of the sequences/cells based on V gene, J gene, and junction length simultaneously. If junc_len=NULL (no column specified), then groupGenes performs only the first stage of a 2-stage partitioning in which sequences/cells are partitioned in the first stage based on V gene and J gene, and then in the second stage further splits the groups based on junction length (the second stage must be performed independently, as this only returns the first stage results).

In the input data, the v_call, j_call, cell_id, and locus columns, if present, must be of type character (as opposed to factor).

It is assumed that ambiguous gene assignments are separated by commas.

All rows containing NA values in any of the v_call, j_call, and junc_len (if junc_len != NULL) columns will be removed. A warning will be issued when a row containing an NA is removed.

Value

Returns a modified data.frame with disjoint union indices in a new vj_group column.

If junc_len is supplied, the grouping this vj_group will have been based on V, J, and junction length simultaneously. However, the output column name will remain vj_group.

The output v_call, j_call, cell_id, and locus columns will be converted to type character if they were of type factor in the input data.

Expectations for single-cell data

Single-cell paired chain data assumptions:

• every row represents a sequence (chain).

• heavy/long and light/short chains of the same cell are linked by cell_id.

• the value in locus column indicates whether the chain is the heavy/long or light/short chain.

• each cell possibly contains multiple heavy/long and/or light/short chains.

• every chain has its own V(D)J annotation, in which ambiguous V(D)J annotations, if any, are separated by a comma.

Single-cell example:

• A cell has 1 heavy chain and 2 light chains.

• There should be 3 rows corresponding to this cell.

• One of the light chains may have an ambiguous V annotation which looks like "Homsap IGKV1-39*01 F,Homsap IGKV1D-39*01 F".

Examples

# Group by genes
db <- groupGenes(ExampleDb)
head(db$vj_group)