RNA-Seq API
pyrpipe implements specialized classes for RNA-Seq processing. These classes are defined into different modules, each designed to capture steps integral to
analysis of RNA-Seq data –from downloading raw data to trimming, alignment and assembly or quantification.
Each of these module implements classes coressponding to a RNA-Seq tool. These classes extend the Runnable class.
Specialized functions are implemented such that analysis of RNA-Seq data is intuitive and easy to code.
The following table provide details about the pyrpipe’s RNA-Seq related modules.
| Module |
Class |
Purpose |
| assembly |
Assembly |
Abstract class to represent Assebler type |
| assembly |
Stringtie |
API to Stringtie |
| assembly |
Cufflinks |
API to Cufflinks |
| mapping |
Aligner |
Abstract class for Aligner type |
| mapping |
Star |
API to Star |
| mapping |
Bowtie2 |
API to Bowtie2 |
| mapping |
Hisat2 |
API to Hisat2 |
| qc |
RNASeqQC |
Abstract class for RNASeqQC type (quality control and trimming) |
| qc |
Trimgalore |
API to Trim Galore |
| qc |
BBmap |
API to bbduk.sh |
| quant |
Quant |
Abstract class for Quantification type |
| quant |
Salmon |
API to Salmon |
| quant |
Kallisto |
API to Kallisto |
| sra |
SRA |
Class to represent RNA-Seq data and API to NCBI SRA-Tools |
| tools |
Samtools |
API to Samtools and other commonly used tools |
The SRA class
The SRA class contained in the sra module captures RNA-Seq data.
It can automatically download RNA-Seq data from the NCBI SRA servers via the prefetch command.
The SRA constructor can take the SRR accession, path to fastq or sra file as arguments.
The main attributes and functions are defined the following table.
| Attribute |
Description |
| fastq_path |
Path to the fastq file. If single end this is the only fastq file. |
| fastq2_path |
Path to the second fastq file for paired end data. |
| sra_path |
Path to the sra file |
| srr_accession |
The SRR accession for RNA-Seq run |
| layout |
RNA-Seq layout, auto determined by SRA class. |
| bam_path |
Path to bam file after running the align() function |
| gtf |
Path to the gtf file after running assemble() function |
| Function |
Description |
| __init__() |
This is the constructor. It can take SRR accession, path to fastq files, or sra file as input. If accession if provided as input the files are downloaded via prefetch if they aren’t preset on disk. It will automatically handle single-end and paired-end data. |
| download_sra() |
This function downloads the sra file via prefetch. |
| download_fastq() |
This function runs fasterq-dump on the sra file downloaded via prefetch. |
| sra_exists() |
Check if fastq sra files are present |
| fastq_exists() |
Check if fastq file exists |
| delete_sra() |
Delete the sra file |
| delete_fastq() |
Delete the fastq files |
| trim() |
This function takes a RNASeqQC type object and performs trimming. The trimmed fastq files are then stored in fastq_path and fastq2_path. |
| align() |
This function takes an Aligner type object and performs read alignemnt. The BAM file returned is stored in bam_path attribute. |
| assemble() |
This function takes an Assembly type object and performs transcript assembly. The result is soted on the SRA object as gtf attributes |
| quant() |
This function takes a Quantification type object and performs quant. |
The RNASeqQC class
The RNASeqQC is an abstract class defined in the qc module. RNASeqQC class extends the Runnable class and thus has all the attributes as in the Runnable class.
Classes Trimgalore and BBmap extends RNASeqQC class and share following attributes and functions.
| Attribute |
Description |
| _category |
Represents the type: “RNASeqQC” |
| Function |
Description |
| __init__() |
The constructor function |
| perform_qc() |
Takes a SRA object, performs qc and returns path to resultant fastq files |
The Aligner class
The Aligner is an abstract class defined in the mapping module. Aligner class extends the Runnable class and thus has all the attributes as in the Runnable class.
Classes Star, Hisat2 and Bowtie2 extends the Aligner class and share following attributes and functions.
| Attribute |
Description |
| _category |
Represents the type: “Aligner” |
| index |
Index used by the aligner tool |
| genome |
Reference genome used by the tool |
| Function |
Description |
| __init__() |
The constructor function |
| build_index() |
Build an index for the aligner tool using the genome. |
| check_index() |
Checks if the index is valid |
| perform_alignment() |
Takes a sra object, performs alignemnt and returns path to the bam file |
The Assembly class
The Assembly is an abstract class defined in the assembly module. Assembly class extends the Runnable class and thus has all the attributes as in the Runnable class.
Classes Stringtie and Cufflinks extends the Assembly class and share following attributes and functions.
| Attribute |
Description |
| _category |
Represents the type: “Assembler” |
| Function |
Description |
| __init__() |
The constructor function |
| perform_assembly() |
Takes a SRA object, performs transcript assembly and returns path to resultant gtf/gff files |
The Quant class
The Quant is an abstract class defined in the quant module. Quant class extends the Runnable class and thus has all the attributes as in the Runnable class.
Classes Salmon and Kallisto extends the Quant class and share following attributes and functions.
| Attribute |
Description |
| _category |
Represents the type: “Quantification” |
| index |
Index used by the aligner tool |
| transcriptome |
Reference transcriptome used by the tool |
| Function |
Description |
| __init__() |
The constructor function |
| build_index() |
Build an index for the quantification tool using the transcriptome. |
| check_index() |
Checks if the index is valid |
| perform_quant() |
Takes a sra object, performs quantification and returns path to the quantification results file |