Figure 4

ATF 4.7 user interface. HLA-A genotype assignment showing the “master layer”. The forward and reverse sequence reads are aligned to exon 2, 3 and 4 with partial coverage of intronic regions. Read counts and sequencing direction are displayed in white bars. For instance, exon 4 is represented by two (allele 1 and 2) forward sequences with coverage of 90, respectively 81 sequence reads and two reverse sequences including 82 and 86 sequence reads. Underneath the exon map, the consensus sequence and the genotype assignment are shown. The phased sequences at the exon 2/intron 2 boundary are illustrated and propose the combination HLA-A*01:01:01:01, 24:02:01:01 with zero mismatches (master layer MM) compared to the database. The non- expressed A*01:01:01:02 N and the low expressed A*24:02:01:02 L are excluded by intron 2 variations as shown in the non coding (N-C) column. Mismatches in the phase layer 3 and 4 (MM3, MM4) demonstrate cis/trans ambiguities that would arise if the phase of SNPs in exon 3 and exon 2 cannot be defined (generic Sanger sequencing). The remaining alleles A*24:02:01:03 (variation in intron 3) and A*24:02:40 (synonymous substitution in exon 5) cannot be excluded and are therefore displayed likewise with zero mismatches.