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Data Descriptor

Novel Molecular Resources to Facilitate Future Genetics Research on Freshwater Mussels (Bivalvia: Unionidae)

by
Nathan A. Johnson
1,* and
Chase H. Smith
2,3,*
1
Wetland and Aquatic Research Center, U.S. Geological Survey, 7920 NW 71st Street, Gainesville, FL 32653, USA
2
Biology Department, Baylor University, Waco, TX 76798, USA
3
Department of Integrative Biology, University of Texas at Austin, 2415 Speedway, Austin, TX 78712, USA
*
Authors to whom correspondence should be addressed.
Data 2020, 5(3), 65; https://0-doi-org.brum.beds.ac.uk/10.3390/data5030065
Submission received: 8 July 2020 / Revised: 27 July 2020 / Accepted: 28 July 2020 / Published: 30 July 2020
(This article belongs to the Section Computational Biology, Bioinformatics, and Biomedical Data Science)
Browse Figure
Versions Notes

Abstract

:
Molecular data have been an integral tool in the resolution of the evolutionary relationships and systematics of freshwater mussels, despite the limited number of nuclear markers available for Sanger sequencing. To facilitate future studies, we evaluated the phylogenetic informativeness of loci from the recently published anchored hybrid enrichment (AHE) probe set Unioverse and developed novel Sanger primer sets to amplify two protein-coding nuclear loci with high net phylogenetic informativeness scores: fem-1 homolog C (FEM1) and UbiA prenyltransferase domain-containing protein 1 (UbiA). We report the methods used for marker development, along with the primer sequences and optimized PCR and thermal cycling conditions. To demonstrate the utility of these markers, we provide haplotype networks, DNA alignments, and summary statistics regarding the sequence variation for the two protein-coding nuclear loci (FEM1 and UbiA). Additionally, we compare the DNA sequence variation of FEM1 and UbiA to three loci commonly used in freshwater mussel genetic studies: the mitochondrial genes cytochrome c oxidase subunit 1 (CO1) and NADH dehydrogenase subunit 1 (ND1), and the nuclear internal transcribed spacer 1 (ITS1). All five loci distinguish among the three focal species (Potamilus fragilis, Potamilus inflatus, and Potamilus purpuratus), and the sequence variation was highest for ND1, followed by CO1, ITS1, UbiA, and FEM1, respectively. The newly developed Sanger PCR primers and methodologies for extracting additional loci from AHE probe sets have great potential to facilitate molecular investigations targeting supraspecific relationships in freshwater mussels, but may be of limited utility at shallow taxonomic scales.
Dataset:https://0-doi-org.brum.beds.ac.uk/10.5066/P9Q3CFL5.
Dataset License: CC0. This work was authored as part of the Contributor’s official duties as an Employee of the United States Government and is therefore a work of the United States Government. In accordance with 17 U.S.C. 105, no copyright protection is available for such works under U.S. Law. This is an open access article that has been identified as being free of known restrictions under copyright law, including all related and neighboring rights (https://creativecommons.org/publicdomain/mark/1.0/). You can copy, modify, distribute, and perform the work, even for commercial purposes, all without asking permission.

1. Summary

Recent studies utilizing molecular tools have been integral in resolving the evolutionary history of freshwater mussels (Bivalvia: Unionida), despite a heavy reliance on the Sanger sequencing of mitochondrial DNA (mtDNA). Nearly all phylogenetic studies on freshwater mussels have largely relied on the commonly used marker internal transcribed spacer 1 (ITS1) to incorporate inference from the nuclear genome [1,2,3,4,5,6], which was first used in freshwater mussels nearly 20 years ago [7]. Even though researchers continue to utilize this locus, the number of studies reporting issues related to excessive heterozygosity, primarily due to length polymorphisms, continues to increase [1,2,6,8,9,10,11,12,13,14]. Other nuclear loci, such as histone H3 and 28S, have been utilized in freshwater mussel phylogenetic studies; however, these markers are well known to show limited diversity at shallow taxonomic scales and have primarily been used to resolve deep level phylogeny [15,16,17,18,19,20].
In recent years, the decreasing costs of next-generation sequencing platforms have significantly increased the ability to generate molecular supermatrices in non-model taxa [21,22], including freshwater mussels [23,24]. In particular, the recently developed anchored hybrid enrichment (AHE) probe set Unioverse [23] has drastically improved the ability to resolve phylogeny in freshwater mussels. The Unioverse probe set consists of 811 protein-coding loci derived from genomic and transcriptomic resources across Bivalvia that can be captured across all freshwater mussels to resolve phylogenetic relationships. Despite the decreasing costs of next-generation sequencing, the utilization of AHE probe sets can be cost-prohibitive for small-scale projects or molecular investigations that incorporate hundreds of individuals to investigate intra- or interspecific relationships. However, AHE probe sets offer opportunities for the development of primers for the amplification and Sanger sequencing of select protein-coding loci that can be used for small-scale projects.
Here, we evaluated the phylogenetic informativeness of loci in the Universe probe set and report the development of novel primer pairs for the amplification of two protein-coding nuclear genes fem-1 homolog C (FEM1) and UbiA prenyltransferase domain-containing protein 1 (UbiA). To demonstrate the utility of these markers and facilitate their use in future studies, we provide the PCR primer sequences, optimized PCR conditions and thermal cycling parameters, haplotype networks, DNA alignments, and summary statistics regarding sequence variation for the two protein-coding nuclear loci (FEM1 and UbiA) and three loci that are commonly used in studies in freshwater mussels: the mitochondrial genes cytochrome c oxidase subunit 1 (CO1) and NADH dehydrogenase subunit 1 (ND1), and the nuclear ITS1 locus. All five loci distinguish among the three focal species (Potamilus fragilis, Potamilus inflatus, and Potamilus purpuratus) and should be amplifiable across the subfamily Ambleminae. The observed sequence variation was highest for ND1, followed by CO1, ITS1, UbiA, and FEM1, respectively (Figure 1). We also provide the detailed methodology used in the marker selection to expedite the identification of additional candidate loci and primer development from available AHE data. The newly developed Sanger PCR primers and methodologies for extracting additional loci have great potential to facilitate molecular investigations targeting supraspecific relationships in freshwater mussels, but may be of limited utility at shallow taxonomic scales.

2. Data Description

2.1. Specimen Details

All the metadata related to the specimens used in this study, including the collection location, GPS coordinates, and museum catalog numbers, are provided (https://0-doi-org.brum.beds.ac.uk/10.5066/P9Q3CFL5) [25].

2.2. Molecular Data

We present the DNA sequence data from five markers: the mitochondrial genes CO1 and ND1, the nuclear non-coding marker ITS1, and the protein-coding nuclear genes FEM1 and UbiA. Our five-locus DNA alignment consisted of 3368 bp of mitochondrial and nuclear sequence data (CO1 = 657 bp; ND1 = 900 bp; FEM1 = 501 bp; UBiA = 765 bp; ITS1 = 545 bp). The number of loci sequenced for each individual varies from two to five loci, with all loci available for 28 individuals (Table 1). The specific sample sizes for each locus are as follows: CO1 (n = 102); ND1 (n = 103); FEM1 (n = 29); UBiA (n = 29); and ITS1 (n = 31). A subset of individuals was chosen for the additional nDNA loci due to the high prevalence of multiple copies at ITS1 and low genetic diversity at FEM1 and UbiA. All the DNA alignment files are available in Phylip format (.phy), with the first line indicating the number of taxa and number of nucleotides and subsequent lines containing a taxon identifier, catalog number, and GenBank Accession number in the first column (each separated by underscore), and the DNA sequence in the second column. The file names are as follows: CO1.phy; ND1.phy; FEM1.phy; UbiA.phy; ITS1.phy; and 5_locus.phy (https://0-doi-org.brum.beds.ac.uk/10.5066/P9Q3CFL5) [25].

3. Methods

3.1. Taxon Sampling and DNA Extraction

We present molecular data on 103 specimens representing Potamilus fragilis (n = 22), Potamilus inflatus (n = 14), and Potamilus purpuratus (n = 67) used in Smith and Johnson [26] (Table 1). All specimens were collected from four Gulf of Mexico river drainages in the southeastern United States: Mobile, Pascagoula, Pearl, and Pontchartrain. Genomic DNA was extracted from mantle tissue clips from vouchered individuals using the Qiagen PureGene DNA extraction kit with the standard extraction protocol (Qiagen, Hilden, Germany).

3.2. Novel Primer Design and Gene Annotation

We compiled data from a recent study [24] utilizing the AHE probe set Unioverse to develop novel primer sets for amplifying protein-coding nuclear loci for use in the freshwater mussel genus Potamilus. To screen for loci in the dataset that were informative at shallow phylogenetic scales, we measured the net phylogenetic informativeness (PI) using an arbitrary time scale [27]. This methodology has been used in previous studies to calculate the power of individual loci in AHE datasets [28,29]. First, we reconstructed a phylogeny from a concatenated alignment of probe loci using IQ-TREE v 1.6.11 [30,31], and the consensus tree was arbitrarily dated with a molecular clock (i.e., tips = time 0, root = time 1) using the program PATHd8 [32]. A concatenated alignment partitioned by the probe and the ultrametric tree from PATHd8 were uploaded into the web application PhyDesign [33] (http://phydesign.townsend.yale.edu/) to estimate the PI using the HyPhy substitution rates algorithm with the GTR model of nucleotide evolution and empirical base frequencies [34]. We used the R script PhyDesign.r [29] to identify specific nucleotide positions in the alignment with unusually high substitution rates that could be contributing phylogenetic noise. Nucleotide positions with rate values higher than five were removed from the alignment manually and the filtered matrices were re-uploaded to PhyDesign as above for a final analysis.
Three nucleotides were removed from the dataset due to unusually high substitution rates (rate value > 5 = “phantom spikes”). In the filtered dataset, the probe regions with a 100% capture efficiency across Ambleminae had an average net PI of 4.62 and ranged from 0.32 to 23.09 (Table 2). Using the results from PhyDesign, we selected two candidate loci for primer development and PCR validation: locus 156 and locus 412. Locus 156 and locus 412 exhibited a 100% capture efficiency in our dataset, had suitable candidate primers that could be cross amplified across Potamilus, displayed high levels of average PI (9.22 and 11.61, respectively), and were able to discriminate our focal species. We were unable to develop compatible primers for the other candidate loci with high net PI scores (e.g., locus 70 and locus 413).
We used BLASTX [35] to annotate the gene and protein names for our candidate loci [36]. Briefly, the probe region sequences of both loci for P. inflatus were searched against the non-redundant protein database using BLASTX, which returned 172 and 118 BLAST hits for locus 156 and locus 412, respectively. Locus 156 was identified as UbiA prenyltransferase domain-containing protein 1, and the highest homology was to genes in the marine bivalves Crassostrea virginica (74.62%) and C. gigas (72.31%). Locus 412 was identified as a fem-1 homolog, and the highest homology was to genes in the unionid bivalves Hyriopsis schlegelii (99.44%) and H. cumingii (98.89%), and the marine bivalves Mizuhopecten yessoensis (87.22%), Pecten maximus (87.22%), C. virginica (86.11%), and C. gigas (86.11%). There were inconsistencies regarding whether the region was a fem-1 homolog A or fem-1 homolog C. All the blast hits except for H. cumingii and H. schlegelii indicated the sequence was representative of fem-1 homolog C; therefore, we annotated the locus as fem-1 homolog C.

3.3. PCR and Sequencing

PCRs were conducted using a 25 µL mixture of the following: molecular grade water (9.5 µL), MyTaqTM Red Mix (12.5 µL; Bioline, London, UK), primers (1.0 µL each), and DNA template (100 ng). The primers for all loci and thermal cycling conditions for CO1, ND1, and ITS1 are reported in Table 3. The thermal cycling conditions for FEM1 and UbiA were as follows: an initial denaturation at 95 °C for 3 min, followed by 35 cycles of 95 °C for 30 s, 51/60 °C (FEM1/UbiA) for 30 s, and 72 °C for 90 s. The products were sent to Molecular Cloning Laboratories (McLAB, South San Francisco, CA, USA) for bi-directional sequencing on an ABI 3730. Geneious v 10.2.3 [37] was used to assemble the contigs and edit chromatograms, and the sequences were aligned in Mesquite v 3.61 [38] using MAFFT v 7.311 [39]. The loci were aligned independently using the L-INS-i method in MAFFT and translated into amino acids to ensure the absence of stop codons and gaps.

3.4. Sequence Variation and Haplotype Analysis

We created haplotype networks (Figure 1) and calculated the nucleotide diversity, number of haplotypes, number of segregating sites, and number of parsimony-informative sites (Table 4) to compare the amounts of sequence variation across all five loci used in this study. The TCS haplotype networks and sequence variation statistics were calculated using PopART 1.7 [42]. All five loci distinguish among the three focal species (Figure 1). The sequence variation was highest for ND1, followed by CO1, ITS1, UbiA, and FEM1, respectively (Table 4). Despite selecting loci from the AHE probe set with a high net PI, the level of sequence variation remains low when compared to mtDNA and ITS1, suggesting the limited utility of the probes at intraspecific levels.

4. User Notes

All the data and metadata described in this study are at https://0-doi-org.brum.beds.ac.uk/10.5066/P9Q3CFL5 [25], and all the novel GenBank accessions for this study were as follows: CO1: MT662002–MT662099; FEM1: MT669773–MT669799; ITS1: MT661766–MT661792; ND1: MT669647–MT669745; and UbiA: MT669746–MT669772 (Table 1).

Author Contributions

Conceptualization, N.A.J. and C.H.S.; methodology, N.A.J. and C.H.S.; validation, N.A.J. and C.H.S.; formal analysis, N.A.J. and C.H.S.; investigation, N.A.J. and C.H.S.; resources, N.A.J. and C.H.S.; data curation, N.A.J. and C.H.S.; writing—original draft preparation, N.A.J.; writing—review and editing, C.H.S.; visualization, N.A.J.; supervision, N.A.J.; project administration, N.A.J.; funding acquisition, N.A.J. All authors have read and agreed to the published version of the manuscript.

Funding

This research was funded by the U.S. Fish and Wildlife Service and U.S. Geological Survey.

Acknowledgments

The authors thank John Pfeiffer for providing preliminary data and advice during the marker development stage of this project and Matt Cannister for assistance with preparing the metadata file for sharing on ScienceBase. Special thanks to Jeff Powell for help obtaining funding, which was provided by the U.S. Fish and Wildlife Service and U.S. Geological Survey. The specimens utilized in this study were either from museum collections or collected under the U.S. Fish and Wildlife Service permit TE 697819-4. Any use of trade, firm, or product names is for descriptive purposes only and does not imply endorsement by the U.S. Government.

Conflicts of Interest

The authors declare no conflict of interest.

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Data 05 00065 g001 550
Figure 1. TCS haplotype networks for (A) cytochrome c oxidase subunit 1 (CO1), (B) fem-1 homolog C (FEM1), (C) internal transcribed spacer 1 (ITS1), (D) NADH dehydrogenase subunit 1 (ND1), and (E) UbiA prenyltransferase domain-containing protein 1 (UbiA). Each colored circle represents a unique haplotype, the colors correspond to individual species, the black circles represent unsampled haplotypes, and the hash marks indicate nucleotide differences between haplotypes.
Figure 1. TCS haplotype networks for (A) cytochrome c oxidase subunit 1 (CO1), (B) fem-1 homolog C (FEM1), (C) internal transcribed spacer 1 (ITS1), (D) NADH dehydrogenase subunit 1 (ND1), and (E) UbiA prenyltransferase domain-containing protein 1 (UbiA). Each colored circle represents a unique haplotype, the colors correspond to individual species, the black circles represent unsampled haplotypes, and the hash marks indicate nucleotide differences between haplotypes.
Data 05 00065 g001
Table
Table 1. Collection information and GenBank or SRA accession numbers for all the specimens and loci analyzed in this study. Museum abbreviations are as follows: UA—Alabama Museum of Natural History; UF—Florida Museum.
Table 1. Collection information and GenBank or SRA accession numbers for all the specimens and loci analyzed in this study. Museum abbreviations are as follows: UA—Alabama Museum of Natural History; UF—Florida Museum.
TaxonIDDrainageSourceCO1ND1ITS1FEM1UbiA
Potamilus fragilisLfraAla001MobileUF438237MT662019MT669665MT661766MT669798MT669771
Potamilus fragilisLfraAmi040PontchartrainUF439330MT662020MT669666MT661773MT669778MT669751
Potamilus fragilisLfraAmi041PontchartrainUF439352MT662021MT669667
Potamilus fragilisLfraAmi042PontchartrainUF439352MT662022MT669668
Potamilus fragilisLfraPrl043PearlUF439332MT662023MT669669
Potamilus fragilisLfraPrl044PearlUF439332MT662024MT669670MT661780MT669785MT669758
Potamilus fragilisLfraPrl045PearlUF439365MT662025MT669671
Potamilus fragilisLfraPrl046PearlUF439343MT662026MT669672
Potamilus fragilisLfraPrl047PearlUF439343MT662027MT669673
Potamilus fragilisLfraPrl048PearlUF439343MT662028MT669674
Potamilus fragilisLfraAmi057PontchartrainUF439529MT662029MT669675
Potamilus fragilisLfraAmi058PontchartrainUF439529MT662030MT669676
Potamilus fragilisLfraAmi059PontchartrainUF439529MT662031MT669677
Potamilus fragilisLfraMob063MobileUF439528MT662033MT669679
Potamilus fragilisLfraMob064MobileUF439528MT662032MT669678
Potamilus fragilisLfraMob065MobileUncatalogedMT662034MT669680MT661792MT669797MT669770
Potamilus inflatusPinfMob001MobileUF439131MT662002MT669647MT661768MT669773MT669746
Potamilus inflatusPinfMob002MobileUF439131MK044952MK045103MK036203MT669774MT669747
Potamilus inflatusPinfMob003MobileUF439131MT662003MT669648MT661769MT669775MT669748
Potamilus inflatusPinfMob004MobileUF439131MK044953MK045104MK036204SRR10579071SRR10579071
Potamilus inflatusPinfMob005MobileUF439131MT662004MT669649MT661770MT669776MT669749
Potamilus inflatusPinfMob006MobileUF439131MT662005MT669650MT661771MT669777MT669750
Potamilus inflatusPinfAmi010PontchartrainUF439530MT662006MT669651MT661774MT669779MT669752
Potamilus inflatusPinfAmi011PontchartrainUF439530MT662007MT669652MT661775MT669780MT669753
Potamilus inflatusPinfAmi012PontchartrainUF439531MT662008MT669653MT661776MT669781MT669754
Potamilus inflatusPinfAmi013PontchartrainUF439532MT662009MT669654MT661777MT669782MT669755
Potamilus inflatusPinfAmi014PontchartrainUF439532MT662010MT669655MT661778MT669783MT669756
Potamilus inflatusPinfAmi015PontchartrainUF439533MT662011MT669656MT661779MT669784MT669757
Potamilus inflatusPinfMob019MobileUF439514MT662012MT669657MT661783MT669788MT669761
Potamilus inflatusPinfMob020MobileUF439514MT662013MT669658MT661784MT669789MT669762
Potamilus inflatusPinfMob021MobileUF439514MT662014MT669659MT661785MT669790MT669763
Potamilus inflatusPinfMob022MobileUF439514MT662015MT669660MT661786MT669791MT669764
Potamilus inflatusPinfMob023MobileUF439514MT662016MT669661MT661787MT669792MT669765
Potamilus inflatusPinfMob017MobileUF439513MT662017MT669662MT661788MT669793MT669766
Potamilus inflatusPinfMob018MobileUF439513MT662018MT669663MT661789MT669794MT669767
Potamilus inflatusPinfMob016MobileUA2696 MT669664MT661781MT669786MT669759
Potamilus purpuratusPpurPas001PascagoulaUF438434MT662035MT669681
Potamilus purpuratusPpurPrl022PearlUF439145MT662036MT669682
Potamilus purpuratusPpurPrl023PearlUF439145MK044960MK045111MK036211MT669799MT669772
Potamilus purpuratusPpurPrl024PearlUF439145MK044961MK045112MK036212
Potamilus purpuratusPpurPrl025PearlUF439145MT662037MT669683
Potamilus purpuratusPpurPrl026PearlUF439145MT662038MT669684MT661767
Potamilus purpuratusPpurAmi038PontchartrainUF439452MT662039MT669685
Potamilus purpuratusPpurAmi039PontchartrainUF439452MT662040MT669686
Potamilus purpuratusPpurAmi040PontchartrainUF439452MT662041MT669687
Potamilus purpuratusPpurAmi041PontchartrainUF439452MT662042MT669688
Potamilus purpuratusPpurAmi042PontchartrainUF439452MT662043MT669689
Potamilus purpuratusPpurAmi043PontchartrainUF439453MT662044MT669690
Potamilus purpuratusPpurAmi044PontchartrainUF439453MT662045MT669691
Potamilus purpuratusPpurAmi045PontchartrainUF439453MT662046MT669692MT661772SRR10579081SRR10579081
Potamilus purpuratusPpurAmi046PontchartrainUF439453MT662047MT669693
Potamilus purpuratusPpurAmi047PontchartrainUF439453MT662048MT669694
Potamilus purpuratusPpurAmi048PontchartrainUF439454MT662049MT669695
Potamilus purpuratusPpurAmi049PontchartrainUF439454MT662050MT669696
Potamilus purpuratusPpurAmi050PontchartrainUF439454MT662051MT669697
Potamilus purpuratusPpurAmi051PontchartrainUF439454MT662052MT669698
Potamilus purpuratusPpurPrl052PearlUF439456MT662053MT669699
Potamilus purpuratusPpurPrl053PearlUF439456MT662054MT669700
Potamilus purpuratusPpurPrl054PearlUF439457MT662055MT669701
Potamilus purpuratusPpurPrl055PearlUF439457MT662056MT669702
Potamilus purpuratusPpurPrl056PearlUF439457MT662057MT669703
Potamilus purpuratusPpurPrl057PearlUF439457MT662058MT669704
Potamilus purpuratusPpurPrl058PearlUF439457MT662059MT669705
Potamilus purpuratusPpurPrl059PearlUF439456MT662060MT669706
Potamilus purpuratusPpurPrl060PearlUF439456MT662061MT669707
Potamilus purpuratusPpurPrl061PearlUF439456MT662062MT669708
Potamilus purpuratusPpurPrl062PearlUF439456MT662063MT669709
Potamilus purpuratusPpurPrl063PearlUF439456MT662064MT669710
Potamilus purpuratusPpurPrl064PearlUF439458MT662065MT669711
Potamilus purpuratusPpurPrl065PearlUF439459MT662066MT669712
Potamilus purpuratusPpurPrl066PearlUF439459MT662067MT669713
Potamilus purpuratusPpurPrl067PearlUF439459MT662068MT669714
Potamilus purpuratusPpurPrl068PearlUF439459MT662069MT669715
Potamilus purpuratusPpurPrl069PearlUF439459MT662070MT669716
Potamilus purpuratusPpurMob081MobileUA62MT662071MT669717
Potamilus purpuratusPpurMob082MobileUA2469MT662072MT669718
Potamilus purpuratusPpurMob083MobileUA2510MT662073MT669719
Potamilus purpuratusPpurMob084MobileUA2562MT662074MT669720
Potamilus purpuratusPpurMob085MobileUA2740MT662075MT669721MT661782MT669787MT669760
Potamilus purpuratusPpurMob086MobileUA3100MT662076MT669722
Potamilus purpuratusPpurMob087MobileUA3123MT662077MT669723
Potamilus purpuratusPpurMob088MobileUA3205MT662078MT669724
Potamilus purpuratusPpurMob089MobileUA3417MT662079MT669725
Potamilus purpuratusPpurMob090MobileUA3482MT662080MT669726
Potamilus purpuratusPpurPas097PascagoulaUF439510MT662081MT669727
Potamilus purpuratusPpurPas098PascagoulaUF439510MT662082MT669728
Potamilus purpuratusPpurPas099PascagoulaUF439510MT662083MT669729
Potamilus purpuratusPpurPas100PascagoulaUF439510MT662084MT669730
Potamilus purpuratusPpurPas101PascagoulaUF439510MT662085MT669731MT661790MT669795MT669768
Potamilus purpuratusPpurPas102PascagoulaUF439510MT662086MT669732
Potamilus purpuratusPpurPas103PascagoulaUF439510MT662087MT669733
Potamilus purpuratusPpurMob107MobileUF439527 MT662088MT669734MT661791MT669796MT669769
Potamilus purpuratusPpurMob108MobileUF439527 MT662089MT669735
Potamilus purpuratusPpurMob109MobileUF439527 MT662090MT669736
Potamilus purpuratusPpurMob110MobileUF439527 MT662091MT669737
Potamilus purpuratusPpurMob111MobileUF439527 MT662092MT669738
Potamilus purpuratusPpurMob112MobileUF439527 MT662093MT669739
Potamilus purpuratusPpurMob113MobileUF439527 MT662094MT669740
Potamilus purpuratusPpurMob114MobileUF439527 MT662095MT669741
Potamilus purpuratusPpurMob115MobileUF439527 MT662096MT669742
Potamilus purpuratusPpurMob116MobileUF439527 MT662097MT669743
Potamilus purpuratusPpurMob117MobileUF439527 MT662098MT669744
Potamilus purpuratusPpurMob118MobileUF439527 MT662099MT669745
Table
Table 2. Average, minimum, and maximum net phylogenetic informativeness (PI), and the time at maximum PI for the 55 loci with a 100% capture efficiency across Ambleminae. Time at max PI represents an arbitrary time scale, with values closer to zero providing the maximum phylogenetic signal at shallower taxonomic scales. Loci are ordered from the highest to lowest average net PI.
Table 2. Average, minimum, and maximum net phylogenetic informativeness (PI), and the time at maximum PI for the 55 loci with a 100% capture efficiency across Ambleminae. Time at max PI represents an arbitrary time scale, with values closer to zero providing the maximum phylogenetic signal at shallower taxonomic scales. Loci are ordered from the highest to lowest average net PI.
LocusAverage Net PIMin PIMax PITime at Max
L41323.095981.10365130.209210.99
L7015.901581.68189918.115040.63
L41211.607130.52760715.101240.93
L1629.2957250.44490411.7650.82
L1569.2201280.43083912.280360.99
L3698.0419720.28790611.836260.99
L5737.9602650.32674610.916730.99
L7477.6019160.30652710.456070.99
L976.8696370.308238.979960.94
L5646.5547540.3907077.8907580.73
L3706.3726690.2294179.1874680.99
L5936.1717680.3173277.6735080.78
L1135.6899080.301917.027460.77
L1085.6131720.2902177.1031360.91
L7765.5838520.1851928.4209680.99
L4265.4071490.2247377.2670660.99
L7455.2635460.1712658.1054920.99
L345.0712160.3758135.9098390.53
L5414.9956030.2224996.4974810.9
L164.8225840.1989336.4588980.96
L2224.8215830.1752826.9773830.99
L1844.8154540.168217.1140620.99
L6634.7542530.2799315.7310630.62
L3194.5094620.3474475.4286270.99
L3814.2605740.3822345.0042450.42
L4863.808630.1466945.44170.99
L6363.6765590.1268595.459360.99
L193.6542610.181874.570870.8
L6673.4420980.1523754.4848310.9
L6973.3007290.3027093.8048310.43
L1553.2506090.1165824.7300850.99
L4202.9013640.1332873.7007770.82
L7722.8679330.0984124.2677940.99
L5762.7042470.229913.0729730.48
L4642.6617260.0923843.9337120.99
L5432.6478270.1064383.6373470.99
L2102.6363950.0866014.0134080.99
L6022.5647710.0899173.7991660.99
L3272.4265690.310263.1065790.29
L5862.3337850.0851073.3648780.99
L5392.3099950.116082.8823910.79
L4882.2703530.085393.2470810.99
L7282.241810.0716193.4564250.99
L2402.1310980.073673.1392560.99
L5682.102640.0799182.9406950.99
L2741.9817360.0655243.0119850.99
L3581.8725610.0590732.9006060.99
L6001.7351050.0584282.6164610.99
L2121.6595490.0630072.3303380.99
L6861.6438690.0562092.4619190.99
L2371.5089570.0493522.2995720.99
L7291.3682670.0433062.1181980.99
L1831.1425170.0363081.758230.99
L5440.8738770.024541.4453290.99
L5100.3171740.0092990.5117760.99
Table
Table 3. PCR primer sequences and cycling conditions used in this study.
Table 3. PCR primer sequences and cycling conditions used in this study.
LocusPrimersSourceConditions
CO1F: 5′-GTTCCACAAATCATAAGGATATTGG-3′
R: 5′-TACACCTCAGGGTGACCAAAAAACCA-3′		Campbell et al. (2005) [40]Johnson et al. 2018 [22]
ND1F: 5′-TGGCAGAAAAGTGCATCAGATTAAAGC-3′
R: 5′-CCTGCTTGGAAGGCAAGTGTACT-3′		Serb et al. (2003) [41]Serb et al. 2003 [41]
ITS1F: 5′-AAAAAGCTTCCGTAGGTGAACCTGCG-3′
R: 5′-AGCTTGCTGCGTTCTTCATCG-3′		King et al. (1999) [7]King et al. 1999 [7]
FEM1F: 5′- GTRATGGAGTATCGCAGTGT-3′
R: 5′-ACRCTYTTCCTGTTAACATC-3′		This studyThis study
UbiAF: 5′- TTTACTCCTGTTGCACTTGGGA-3′
R: 5′-AGCATCTGTCATGAAGACTCCAAC-3′		This studyThis study
Table
Table 4. Summary of diversity indices based on all five loci utilized in this study. Abbreviations and symbols are as follows: cytochrome c oxidase subunit 1 (CO1); fem-1 homolog C (FEM1); internal transcribed spacer 1 (ITS1); NADH dehydrogenase subunit 1 (ND1); UbiA prenyltransferase domain-containing protein 1 (UbiA); sample size (n); nucleotide diversity (π); number of haplotypes (nh); number of segregating sites (S); and number of parsimony-informative sites (P).
Table 4. Summary of diversity indices based on all five loci utilized in this study. Abbreviations and symbols are as follows: cytochrome c oxidase subunit 1 (CO1); fem-1 homolog C (FEM1); internal transcribed spacer 1 (ITS1); NADH dehydrogenase subunit 1 (ND1); UbiA prenyltransferase domain-containing protein 1 (UbiA); sample size (n); nucleotide diversity (π); number of haplotypes (nh); number of segregating sites (S); and number of parsimony-informative sites (P).
LocusnπnhSP
CO1280.047864148278
FEM1280.003431344
ITS1280.039169633331
ND1280.06211113146138
UbiA280.004002487

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Johnson, N.A.; Smith, C.H. Novel Molecular Resources to Facilitate Future Genetics Research on Freshwater Mussels (Bivalvia: Unionidae). Data 2020, 5, 65. https://0-doi-org.brum.beds.ac.uk/10.3390/data5030065

AMA Style

Johnson NA, Smith CH. Novel Molecular Resources to Facilitate Future Genetics Research on Freshwater Mussels (Bivalvia: Unionidae). Data. 2020; 5(3):65. https://0-doi-org.brum.beds.ac.uk/10.3390/data5030065

Chicago/Turabian Style

Johnson, Nathan A., and Chase H. Smith. 2020. "Novel Molecular Resources to Facilitate Future Genetics Research on Freshwater Mussels (Bivalvia: Unionidae)" Data 5, no. 3: 65. https://0-doi-org.brum.beds.ac.uk/10.3390/data5030065

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