Figure 3

Centromeric haplotype for NA06990. Panel A shows the haplotypes for the most centromeric repeats for NA06990. Constituent haplotypes are resolved using allele-specific primers, specific to either the T or C at rs4512398. The phase of sequence variants within the gene (numbers 2 to 8; see additional Table 1) are shown boxed, from variant number 2 on the left to number 8 on the right. Variant positions 5 and 6 are shown in bold. The corresponding base in the sequence trace is shown boxed. Examination of the sequence at variant position 1 (data not shown) illustrated that T at rs4512398 is on the same chromosome as a copy of DEFA1 and C at rs4512398 is in cis with a copy of DEFA3. Panel B illustrates some of the sequence information used to reconstruct the full haplotypes in this individual. For the centromeric emulsion product, allele-specific sequence using primer T at rs4512398 shows mixed positions at variant positions 2 and 3. At variant position 2, C is observed as the major peak, and A the minor peak and at variant position 3, C is the major peak with T as the minor peak. Sequencing of the entire fused product showed a C at variant position 1, which defines a copy of DEFA1 thereby excluding inappropriate fusion with Repeat 3 on the other haplotype, which carries a copy of DEFA3. Sequencing from the telomeric direction (telomeric emulsion product) showed precisely reciprocal mixed positions (with A at variant 2 being the major product and C the minor). This is consistent with a major contribution from Repeat 1 and a minor contribution from Repeat 2. This reciprocity allowed reconstruction of this 2-copy haplotype. DEFA1A3 repeat units are illustrated as arrows.