Figure 1. Figure shows the methods used to narrow down the location of the linkage region. A: Segregation of the CCV phenotype is shown for a small branch of the large multigenerational Volkmann family [10,11]. The fine-mapped CCV linkage region includes eight loci at 1pter, and the disease haplotype is shown boxed and marked in red. The haplotypes for individuals I:1 and I:2 are inferred, and the recombination events in individuals II:3, II:4, II:5, and IV:8 are shown with an arrow. Symbols: square, male; circle, female; filled symbol, affected individual; dot in a symbol, healthy carrier. B: The chromosomal map of 1p36.33–36.32 shows the Volkmann cataract locus found by the initial genome-wide linkage analysis [11] together with the regions covered by direct Sanger sequencing of protein coding genes and with targeted next-generation sequencing (NGS) and whole genome sequencing (WGS). The key sequence-tagged sites (STSs) and single nucleotides (SNPs) are shown for the 2 Mb telomeric chromosome 1 regions together with the Sanger sequenced genes in the linkage region flanked by STS-22AC/ rs761317190 and rs549772338. C: The exon/intron gene structure for the lncRNA gene RP1–140A9.1 (1) includes the position of the CCV splice site mutation, the PCR primer pair (F1 and R1) positions, and the length and structure of the corresponding PCR amplicons (2–4). The CCV mutation is localized distal in exon 1. The cDNA (218 bp) PCR detected (2) from transformed B-leucocytes. The cDNA 324 bp PCR product detected (3) from two different mixed carcinoma cell preparations and from a lens cDNA library. A 500 bp band (4) from gDNA. The alternative splice sites shown in (2,3) are proposed by the NetGene2 predictor. The enhancer-promoter-associated histone mark H3K27ac (5) is active in the lncRNA gene RP1–140A9.1. The positions are shown for the four CAGCTG repeats (6) found in the region. The CAGCTG palindrome is a putative binding site for hsa-miR-1207-3p (seed sequence CAGCUG) and a binding site for the transcription factor AP-4 [35] proposed by TFSEARCH. The palindrome sequence at the exon/intron border is present only in the mutant sequence.