A script to filter erroneous CDR3 sequences produced due to hot-spot PCR and NGS errors. It can also use a hybrid error correction method that includes frequency-based filtering of singleton clonotypes (i.e. clonotypes represeted by a single MIG).
java -jar migec.jar FilterCdrBlastResultsBatch [options] cdrblast_batch_folder/ output_folder/
Perform hot-spot error filtration for data process with CdrBlastBatch. Options are the same as for manual version below.
java -jar migec.jar FilterCdrBlastResults [options] cdrblast_result_assembled_data cdrblast_result_raw_data output_file
java -jar migec.jar FilterCdrBlastResults cdrblast/S1_asm.cdrblast.txt cdrblast/S1_raw.cdrblast.txt final/S1.cdrblast.txt
-s option tells to filter CDR3s represented by single MIGs. The
rationale for this is that the deep repertoire profiling (at least with
our protocol) can generate spurious singletons that are associated with
reverse transcription errors and experimental artifacts. Filtering is a
non-greedy procedure and filters single-MIG clonotypes only if a 1- or
2-mismatch parent clonotype exists at ratio 1:20 and 1:400 respectively.
This is done to preserve diversity for samples with shallow sequencing,
e.g. ran on MiSeq.
-n- output non-coding clonotypes that contain either a stop codon or a frameshift within CDR3 sequence.
-c- include non canonical clonotypes that have a CDR3 region that does not start with conserved C residue, or end with a conserved F/W residue.
-r- sets the read accumulation threshold (default is
1.0) used for hot-spot error correction, see MiGEC paper for details.
Now the file S1.cdrblast.txt contains a filtered and sorted CDR3/V/J clonotype table.