Strand invaders {CAGEr} | R Documentation

## Detect and remove strand invasion artefacts

### Description

`findStrandInvaders` detects strand invasion artefacts in the CTSS data. `removeStrandInvaders` removes them.

Strand invaders are artefacts produced by template switching reactions used in methods such as nanoCAGE and its derivatives (C1 CAGE, ...). They are described in details in Tang et al., 2013. Briefly, these artefacts create CAGE-like signal downstream of genome sequences highly similar to the tail of template-switching oligonucleotides, which is `TATAGGG` in recent (2017) nanoCAGE protocols. Since these artefacts represent truncated cDNAs, they do not indicate promoter regions. It is therefore advisable to remove these artefacts. Moreover, when a sample barcode is near the linker sequence (which is not the case in recent nanoCAGE protocols), the strand-invasion artefacts can produce sample-specific biases, which can be confounded with biological effects depending on how the barcode sequences were chosen. A `barcode` parameter is provided to incorporate this information.

### Usage

    findStrandInvaders(object, distance = 1, barcode = NULL,
      linker = "TATAGGG")

    removeStrandInvaders(object, distance = 1, barcode = NULL,
      linker = "TATAGGG")

    ## S4 method for signature 'CAGEexp'
    findStrandInvaders(object, distance = 1,
      barcode = NULL, linker = "TATAGGG")

    ## S4 method for signature 'CAGEexp'
    removeStrandInvaders(object, distance = 1,
      barcode = NULL, linker = "TATAGGG")

    ## S4 method for signature 'CTSS'
    findStrandInvaders(object, distance = 1,
      barcode = NULL, linker = "TATAGGG")

    ## S4 method for signature 'CTSS'
    removeStrandInvaders(object, distance = 1,
      barcode = NULL, linker = "TATAGGG")

### Arguments

`object` | A `CAGEexp` object object containing CTSS data and the name of a reference genome.
`distance` | The maximal edit distance between the genome and linker sequences. Regardless this parameter, only a single mismatch is allowed in the last three bases of the linker.
`barcode` | A vector of sample barcode sequences, or the name of a column metadata of the `CAGEexp` object containing this information. (Not implemented yet)
`linker` | The sequence of the tail of the template-switching oligonucleotide, that will be matched with the genome sequence (defaults to `TATAGGG`).

### Value

`findStrandInvaders` returns a logical-Rle vector indicating the position of the strand invaders in the input ranges. With CTSS objects as input `removeStrandInvaders` returns the object after removing the CTSS positions identified as strand invaders. In the case of `CAGEexp` objects, the input object is modified in place. Its sample metadata is also updated by creating a new `strandInvaders` column that indicates the number of molecule counts removed. This value is subtracted from the `counts` colum so that the total number of tags is still equal to `librarySizes`.

### References

Tang et al., “Suppression of artifacts and barcode bias in high-throughput transcriptome analyses utilizing template switching.” Nucleic Acids Res. 2013 Feb 1;41(3):e44. PubMed ID: 23180801, DOI: 10.1093/nar/gks112

### Examples

    # Note that these examples do not do much on the example data since it was
    # not constructed using a protocol based using the template-switching method.
    "BSgenome.Drerio.UCSC.danRer7"

    findStrandInvaders(exampleCAGEexp)
    removeStrandInvaders(exampleCAGEexp)

[Package CAGEr version 1.30.3 Index]