Question: How are full length TCR/Ig transcripts assembled for a Single Cell V(D)J library?
Answer: Full length V(D)J sequences are obtained by assembling multiple short reads that share the same cell barcode against a reference. These short reads are randomly distributed and collectively span the entire T Cell Receptor (TCR) or immunoglobulin (Ig) sequence, including the 5' UTR.
The fragmentation step in the protocol ensures that library inserts representing different starting positions across the transcript are sequenced. The assembled contig sequences are therefore much longer than the individual reads and variants can be detected anywhere in the framework regions.