Computational protocol: Mitochondrial DNA Diversity of Modern, Ancient and Wild Sheep (Ovis gmelinii anatolica) from Turkey: New Insights on the Evolutionary History of Sheep

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Protocol publication

[…] For the modern samples (domestic and O. g. anatolica), the lengths (75 bp/76 bp) of the repeat unit in CR (starting from the 15640th nucleotide of AF010406) as well as the number of repeat units for each sequence (n = 240) were examined.A neighbor-joining tree based on mtDNA CR sequences of the modern samples was constructed by MEGA 5 . The complete tandem repeat region was excluded from CR sequences for the tree construction. The Tamura-Nei model was applied to the sequences with 1000 bootstrap value. As out-groups O. vignei (AY091490, AY091491) and O. ammon (AF242347, AF242348) sequences and as a reference for each haplogroup HM236174–83 sequences were used.For the modern samples, nucleotide diversity (π) and haplotype diversity for each haplogroup, as well as the average number of nucleotide differences per site (Dxy) between haplogroups were calculated by DnaSP v.5 . For the calculation of Dxy between O. g. musimon and HPG B, O. g. musimon samples HM236184–85 , AY091487–88 , AF039579 from previous studies were used.Fu's FS test, Tajima's D test and mismatch distributions were calculated by Arlequin 3.11 for each haplogroup, except HPG D since it had a small sample size. The validity of the sudden population expansion model was tested using the sum of square deviations between the observed and expected mismatch values . For the rare HPG E, the analyses were repeated for three cases; E1: sequences (n = 11) were in 886 bp length and were the samples of the present study; E2: sequences (n = 15) were in 886 bp length and two sequences (HM236182–83) from Meadows et al. and another two sequences (AY829385 and AY829404) from Guo et al. were added to the sequences of E1; E3: sequences (n = 20) were in 665 bp length and DQ097468 , HM042760–61, HM042785, HM042838 sequences were added to the sequences of E2. The sequences used in neighbor-joining tree construction, genetic diversity calculations and mismatch distribution analyses are summarized in .The haplogroup frequencies obtained as a result of ND2-SSCP analysis (n = 628) were used to construct the pie charts to demonstrate the spatial distribution of haplogroups across Turkey. The haplogroup frequencies of O. g. anatolica and ancient samples based on mtDNA CR sequences are also given as pie charts.To examine the spatial distribution of haplogroups among the modern domestic sheep of Turkey, spatial autocorrelation coefficients (r) at each of 150 km distance classes based on the flock information of the samples (n = 628) were used (). The distance class was chosen under the assumption that it is the diameter of a circle which contains the sample collection sites for a breed. The correlogram was constructed with GENALEX 6 . The 95% CI belt for the spatial autocorrelation coefficients (r) under the null hypothesis of no genetic structure among the samples over the space was determined by 999 permutations. The 95% CI of r for each distance class was determined using 1000 bootstrap replicates.For the modern samples (domestic, wild and feral sheep), a Median-joining (MJ) network with partial mtDNA cytB region was constructed by NETWORK, nucleotide positions were weighted as was done by Olivieri et al. . The sequences retrieved for MJ network construction are listed in .The sequences of mtDNA CR of domestic sheep (n = 240), the mtDNA CR of O. g. anatolica (n = 30), the partial cytB of domestic sheep consisting of 2 from each haplogroup (n = 10) and the partial cytB of O. g. anatolica consisting of 2 from each haplotype (n = 4) that were obtained in the present study are available in GenBank accession no. KF677024–KF677307. […]

Pipeline specifications

Software tools MEGA, DnaSP, Arlequin, GenAlEx
Applications Phylogenetics, Population genetic analysis
Organisms Ovis aries, Meleagris gallopavo
Diseases Ataxia Telangiectasia