Development and characterization of microsatellite primers for Triops granarius (Branchiopoda: Notostraca) using MiSeq technology

Background Next-generation sequencing technology has allowed for the rapid development of microsatellites, neutral polymorphic markers that can be used for the analysis of population structure. Methods and Results In this study, we performed whole-genome sequencing using the Illumina MiSeq system and de novo assembly to design microsatellite primers for Triops granarius populations in Qatar. The developed microsatellites are suitable for future studies of genetic structuring among geographically isolated freshwater pools. A total of 23 different primer pairs produced typical microsatellite results, with each pair successfully amplified in up to 40 individuals. Only five of the loci produced a significant departure from Hardy-Weinberg equilibrium. Conclusions Some of the underlying mechanisms regarding the few loci that deviated from HWE may be further investigated to determine the source of deviation. As T. granarius is the most widely distributed species of the family, the development of these molecular markers would be useful for conducting population genetics and biogeographical studies broadly.Other Information Published in: Molecular Biology Reports License: https://creativecommons.org/licenses/by/4.0See article on publisher's website: http://dx.doi.org/10.1007/s11033-022-07804-4</p

Distribution of total number of variants (SNPs and indels) upon step-wise addition of exomes, and the distribution of number of new variants added per exome.

<p>(Coefficient of determination (R²) >0.99 for all fitted curves).</p

Genome at Juncture of Early Human Migration: A Systematic Analysis of Two Whole Genomes and Thirteen Exomes from Kuwaiti Population Subgroup of Inferred Saudi Arabian Tribe Ancestry

<div><p>Population of the State of Kuwait is composed of three genetic subgroups of inferred Persian, Saudi Arabian tribe and Bedouin ancestry. The Saudi Arabian tribe subgroup traces its origin to the Najd region of Saudi Arabia. By sequencing two whole genomes and thirteen exomes from this subgroup at high coverage (>40X), we identify 4,950,724 Single Nucleotide Polymorphisms (SNPs), 515,802 indels and 39,762 structural variations. Of the identified variants, 10,098 (8.3%) exomic SNPs, 139,923 (2.9%) non-exomic SNPs, 5,256 (54.3%) exomic indels, and 374,959 (74.08%) non-exomic indels are ‘novel’. Up to 8,070 (79.9%) of the reported novel biallelic exomic SNPs are seen in low frequency (minor allele frequency <5%). We observe 5,462 known and 1,004 novel potentially deleterious nonsynonymous SNPs. Allele frequencies of common SNPs from the 15 exomes is significantly correlated with those from genotype data of a larger cohort of 48 individuals (Pearson correlation coefficient, 0.91; p <2.2×10⁻¹⁶). A set of 2,485 SNPs show significantly different allele frequencies when compared to populations from other continents. Two notable variants having risk alleles in high frequencies in this subgroup are: a nonsynonymous deleterious SNP (rs2108622 [19:g.15990431C>T] from CYP4F2 gene [MIM:*604426]) associated with warfarin dosage levels [MIM:#122700] required to elicit normal anticoagulant response; and a 3′ UTR SNP (rs6151429 [22:g.51063477T>C]) from ARSA gene [MIM:*607574]) associated with Metachromatic Leukodystrophy [MIM:#250100]. Hemoglobin Riyadh variant (identified for the first time in a Saudi Arabian woman) is observed in the exome data. The mitochondrial haplogroup profiles of the 15 individuals are consistent with the haplogroup diversity seen in Saudi Arabian natives, who are believed to have received substantial gene flow from Africa and eastern provenance. We present the first genome resource imperative for designing future genetic studies in Saudi Arabian tribe subgroup. The full-length genome sequences and the identified variants are available at <a href="ftp://dgr.dasmaninstitute.org" target="_blank">ftp://dgr.dasmaninstitute.org</a> and <a href="http://dgr.dasmaninstitute.org/DGR/gb.html" target="_blank">http://dgr.dasmaninstitute.org/DGR/gb.html</a>.</p></div

Markers denoting causal variants for OMIM diseases and showing significant differences in risk allele frequencies between the KWS and continental populations.

<p>Markers denoting causal variants for OMIM diseases and showing significant differences in risk allele frequencies between the KWS and continental populations.</p

Venn diagram depicting the number of SNPs having significant difference in allele frequencies between the KWS group and other continent populations from the 1000 Genomes Project (F_st >0.25 & q-value <0.05).

<p>Venn diagram depicting the number of SNPs having significant difference in allele frequencies between the KWS group and other continent populations from the 1000 Genomes Project (F_st >0.25 & q-value <0.05).</p

Phylogenetic tree of the observed HVS1 segments among the 15 participants together with those observed by Abu Amero [46] in Saudi Arabia natives.

<p>Kuwaiti samples are labeled as KWS. Green triangles denote sample from Central region of Saudi Arabia; Blue triangles denote samples from Southern region of Saudi Arabia; Red triangles denote samples from Western region of Saudi Arabia; Black triangles denote samples from Northern region of Saudi Arabia; Not Known [Cyan triangles].</p

Intergenome distances between the KWS genomes and individuals from continental populations.

<p>(a) Nearest neighbor tree based on variant positions shared between the KWS samples and individuals from intercontinental populations. (b). Intergenome comparisons based on variant positions associated with OMIM disease genes and are shared between the KWS samples and individuals from intercontinental populations.</p

Summary of analysis of genomes from Kuwait subgroup of Saudi Arabian tribe ancestry.

<p>Tracks (from outer to inner): Karyotype of Human Genome; Density (in every window of 1 Mb) of ‘known’ SNPs (i.e. annotated in dbSNP 137) from the UE data set; Density of ‘novel’ SNPs (i.e. not annotated in dbSNP137) from the UE data set; Density of ‘known’ indels from the UE data set; Density of ‘novel’ indels from the UE data set; Density of ‘known’ SNPs from the UW data set; Density of ‘novel’ SNPs from the UW data set; Density of ‘known’ indels from the UW data set; Density of ‘novel’ indels from the UW data set; Density of long Indels; Density of duplications, inversions and tandem duplications; Links representing intra- and inter-chromosomal translocations. The image was generated using Circos <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0099069#pone.0099069-Krzywinski1" target="_blank">[71]</a>.</p

Plot (Phylogenetic tree) of pairwise F_ST distances for the three Kuwaiti groups with a set of representative HGDP populations that cover Europe, Asia, and Middle East.

<p>Bootstrap confidence – Red: 70% to 80%; Blue -80% to 90%; Green -90% to 100%.</p

Genetic Substructure of Kuwaiti Population Reveals Migration History

<div><p>The State of Kuwait is characterized by settlers from Saudi Arabia, Iran, and other regions of the Arabian Peninsula. The settlements and subsequent admixtures have shaped the genetics of Kuwait. High prevalence of recessive disorders and metabolic syndromes (that increase risk of diabetes) is seen in the peninsula. Understanding the genetic structure of its population will aid studies designed to decipher the underlying causes of these disorders. In this study, we analyzed 572,366 SNP markers from 273 Kuwaiti natives genotyped using the illumina HumanOmniExpress BeadChip. Model-based clustering identified three genetic subgroups with different levels of admixture. A high level of concordance (Mantel test, p=0.0001 for 9999 repeats) was observed between the derived genetic clusters and the surname-based ancestries. Use of Human Genome Diversity Project (HGDP) data to understand admixtures in each group reveals the following: the first group (Kuwait P) is largely of West Asian ancestry, representing Persians with European admixture; the second group (Kuwait S) is predominantly of city-dwelling Saudi Arabian tribe ancestry, and the third group (Kuwait B) includes most of the tent-dwelling Bedouin surnames and is characterized by the presence of 17% African ancestry. Identity by Descent and Homozygosity analyses find Kuwait’s population to be heterogeneous (placed between populations that have large amount of ROH and the ones with low ROH) with Kuwait S as highly endogamous, and Kuwait B as diverse. Population differentiation F_ST estimates place Kuwait P near Asian populations, Kuwait S near Negev Bedouin tribes, and Kuwait B near the Mozabite population. F_ST distances between the groups are in the range of 0.005 to 0.008; distances of this magnitude are known to cause false positives in disease association studies. Results of analysis for genetic features such as linkage disequilibrium decay patterns conform to Kuwait’s geographical location at the nexus of Africa, Europe, and Asia.</p> </div