Sequencher file of processed and assembled ITS2 chromatograms
2016-10-31T11:40:40Z (GMT) by
In this Sequencher 4.1.4 file, the forward and reverse ITS2 chromatograms of each individual were assembled into contigs. For each homozygote, the contig was checked and cleaned then received the name of the individual sequenced. For length-variant heterozygotes, the double peaks were called either manually or using Sequencher's "Call Secondary Peak..." function, then the two haplotypes were reconstructed by combining the information carried by the forward and reverse chromatograms using the program Champuru (see http://dx.doi.org/10.1111/j.1471-8286.2006.01355.x, http://dx.doi.org/10.1111/j.1471-8286.2007.01857.x, available online at http://jfflot.mnhn.fr/champuru/ and http://seqphase.mpg.de/champuru); the contig and reconstructed haplotypes of each such individual received the name of this individual followed with "H", "a" and "b", respectively. For heterozygotes having two alleles of equal lengths, we used SeqPHASE (http://dx.doi.org/10.1111/j.1755-0998.2009.02732.x, available online at http://seqphase.mpg.de/seqphase) and PHASE to infer their haplotypes; the contig of each such individual received the name of this individual followed with "H" then the number of heterozygous SNPs observed (i.e., the number of double peaks in the contig), whereas its two inferred haplotypes received its name followed with "a" or "b", respectively. Finally, some individuals had double peaks but their chromatograms were two noisy to reconstruct accurately the two sequences: rather than risking to reconstruct one erroneous haplotype, for each such individual we only kept the haplotype that could be reconstructed unambiguously; these contigs were named by appending "h" to the names of the individuals concerned.