An efficient field and laboratory workflow for plant phylotranscriptomic projects.

PREMISE OF THE STUDY: We describe a field and laboratory workflow developed for plant phylotranscriptomic projects that involves cryogenic tissue collection in the field, RNA extraction and quality control, and library preparation. We also make recommendations for sample curation. METHODS AND
RESULTS: A total of 216 frozen tissue samples of Caryophyllales and other angiosperm taxa were collected from the field or botanical gardens. RNA was extracted, stranded mRNA libraries were prepared, and libraries were sequenced on Illumina HiSeq platforms. These included difficult mucilaginous tissues such as those of Cactaceae and Droseraceae.
CONCLUSIONS: Our workflow is not only cost effective (ca. $270 per sample, as of August 2016, from tissue to reads) and time efficient (less than 50 h for 10-12 samples including all laboratory work and sample curation), but also has proven robust for extraction of difficult samples such as tissues containing high levels of secondary compounds.

Phylotranscriptomics, or using transcriptome sequences to investigate phylogenetic relationships and gene family evolution in nonmodel plants, has gained popularity in recent years due to decreases in cost and improvements in analysis pipelines (Wickett et al., 2014; Edger et al., 2015; Li et al., 2015; Yang et al., 2015; McKain et al., 2016). It is often possible to recover at least 15,000 genes from the target species using de novo–assembled transcriptome data (Yang and Smith, 2013). Among these, approximately 5000 are shared among most species within an order (Yang et al., 2015), with the rest being tissue- and/or taxon-specific. Together they provide enormously rich data both for phylogenetic reconstruction and for investigating gene family evolution that underlies lineage-specific adaptations.

Generating plant phylotranscriptomic data has become much easier over the past few years due to improvements in sequencing and extraction protocols but may still be challenging for a variety of reasons. Previous literature on phylotranscriptomic methods has focused on RNA extraction and fragment analyses of those extracted RNA samples (Johnson et al., 2012; Yockteng et al., 2013; Jordon-Thaden et al., 2015) and sequence data analyses (Yang and Smith, 2013, 2014). However, as phylotranscriptomic studies expand to nonmodel systems that often require field sampling, the logistics of obtaining fresh tissues becomes a limiting factor. Likewise, some taxa such as cacti pose special challenges due to high levels of mucilage (Jordon-Thaden et al., 2015). Moving forward, the issues of long-term preservation and curation of cryogenic genetic materials will also be of the utmost importance for laboratories seeking to pursue these studies.

From 2012 to 2015, we conducted field expeditions to remote localities in both the southwestern United States and northern Mexico to support National Science Foundation–funded projects on the evolution of Caryophyllales and gypsum-endemic plants. Together with samples from living collections, we generated a transcriptome data set of 200 species of plants (Appendix 1). During the process we have developed an optimized workflow, which is described below. In addition, we discuss alternative procedures that we tested, as well as considerations for project planning.

METHODS AND RESULTS

Taxon sampling

The Caryophyllales phylotranscriptomics project emphasized a combination of broad taxon sampling across the order and in-depth sampling of lineages with key evolutionary transitions. These key transitions include the gain and loss of plant carnivory; the gain and loss of betalain pigmentation; transitions to saline, dry, or alpine habitats, and/or to specialized soil types; and transitions to C4 and CAM photosynthesis. Of the transcriptomes we have generated for the Caryophyllales phylotranscriptomic project, half were collected from the field, with the remaining half from living collections (Appendix 1). Additional transcriptomes and genomes were obtained from publicly available databases such as Phytozome (Goodstein et al., 2012), the National Center for Biotechnology Information (NCBI) Sequence Read Archive (SRA), and the 1000 Plants Initiative (1KP; Matasci et al., 2014).

Field collection

We timed our field trips to coincide with the beginning of the flowering season as much as possible to optimize the chance of obtaining young flower and leaf buds. Our experience has been that mature vegetative tissue is more difficult to work with due to its low concentration of nuclear RNA (Johnson et al., 2012) and high level of chloroplast RNA and secondary compounds compared to developing tissues. It is also important to emphasize that field conditions are more difficult to control than greenhouse conditions. While this may impose limitations for researchers wishing to study differential gene expression, this is less problematic for phylotranscriptomic studies.

Compared to tissue preservation using an RNA stabilization solution (such as RNAlater; Thermo Fisher Scientific, Waltham, Massachusetts, USA), tissue frozen in the field allows for biochemical analyses such as characterization of betalain and anthocyanin pigmentation, in addition to DNA and RNA sequencing, and hence this was our primary (and recommended) means of collection (Appendix 2). For all individuals frozen in liquid nitrogen, we also collected silica-preserved tissue from the same individual as a DNA backup, as well as herbarium specimens whenever possible. Because DNA may degrade relatively quickly for some groups in silica (e.g., Onagraceae), it is important to remove silica from the leaves once dried and place them in a −20°C freezer for long-term storage (Neubig et al., 2014).

RNA extraction (less than 6 h for six samples)

We tested five alternative RNA extraction protocols. These include TRIzol option 1 from Jordon-Thaden et al. (2015), the Aurum Total RNA Mini Kit (Bio-Rad Laboratories, Hercules, California, USA) following the manufacturer’s protocol, the QIAGEN RNeasy Mini Kit (QIAGEN, Hilden, Germany) following the manufacturer’s protocol, the PureLink protocol (Appendix 3; Yockteng et al., 2013), and the hot acid phenol-LiCl-RNeasy Mini Kit protocol (Appendix 4, modified from Protocol 12 of Johnson et al. [2012]). We had approximately 10–30% success rate (see below for quality control) with Bio-Rad, QIAGEN, and TRIzol protocols, whereas the PureLink protocol had close to 100% success rate and only failed when the sample itself was degraded or highly mucilaginous. Although more time consuming, the hot acid phenol-LiCl-RNeasy Mini Kit protocol had great success with tissues that are highly mucilaginous like cacti (Appendix 4).

Quality control and DNase digestion (less than 3 h for 12 samples)

For quality control of RNA, we used agarose gel for an initial assessment. If RNA was evident, removal of DNA was carried out following Jordon-Thaden et al. (2015) with minor modifications (Appendix 5). After that, we followed fig. 2 of Jordon-Thaden et al. (2015) for evaluating integrity of RNA on a 2100 Bioanalyzer (Agilent, Santa Clara, California, USA) or a Fragment Analyzer (Advanced Analytical Technologies, Ankeny, Iowa, USA). RNA concentration was measured with either a NanoDrop Spectrophotometer (Thermo Fisher Scientific) or a Qubit fluorometer (Thermo Fisher Scientific). We considered an RNA integrity number (RIN) of 6 or higher and concentration of 20 ng/μL or higher as successful. When RNA extraction failed, it was often due to either pellet loss (resulting in a completely empty gel with no DNA or RNA trace) or degradation (which shows up as smeared ribosomal RNA bands). RNA degradation can happen during collection, shipping, or in a suboptimal extraction, as for example with too much starting tissue. For difficult tissues that are mucilaginous, we reduced the amount of starting tissue by half.

RNA samples prepared at the Brockington Laboratory at the University of Cambridge, United Kingdom, were shipped on dry ice in cardboard freezer boxes to the University of Michigan for library preparation and sequencing. Dry ice shipments were sent on Monday or Tuesday to avoid delay over the weekend.

Library preparation (less than 20 h for 12 samples)

We tested four different library preparation protocols. In 2012, we started with Illumina TruSeq version 2 (Illumina, San Diego, California, USA), with and without additional strand-specific steps (see Supplementary Methods in Yang et al. [2015]). In 2013, we began using the newly released TruSeq Stranded mRNA Library Prep Kit (“the Illumina kit”; Illumina), which was more streamlined and produced much higher strand specificity than the previous stranded protocol. In 2014, we switched to the KAPA Stranded mRNA-Seq kit (“the KAPA kit”; KAPA Biosystems, Wilmington, Massachusetts, USA; Appendix 6), which is considerably cheaper than the Illumina kit with indistinguishable results in terms of both success rate and strand specificity. The KAPA kit is also more streamlined with fewer bead washing steps and required roughly 15% less time. The cost is ca. US$30 per sample for the KAPA kit itself plus ca. US$20 per sample for consumables (magnetic beads, tips, tubes, and additional chemicals; we used leftover adapters from the Illumina kit, which lasted through more than 150 additional libraries from one 48-sample Illumina kit). We modified the manufacturer’s protocol slightly to accommodate the increasing read length of newer Illumina platforms (125- or 150-bp paired-end; Appendix 6).

Quality control of the library was done at the University of Michigan DNA Sequencing Core using an Agilent 2100 Bioanalyzer followed by confirmation using qPCR. Although the minimal concentration of the library and percentage of adapter contamination allowed differ among sequencing platforms, we followed a few general rules. First, the peak of the library fragment size distribution should be approximately the read length plus adapter size. For example, for paired-end 125-bp sequencing on Illumina platforms, peak of library size distribution should be approximately 60 bp (adapter) + 125 bp (read) in each direction, making a total of 370 bp for the optimum library size (see Appendix 6 for modifications in library preparation to adjust library sizes). Second, although we do not quantify the library concentration in the laboratory, we visualized the library by loading 3 μL of library mixed with GelRed fluorescent stain (Biotium, Fremont, California, USA) onto a 1.5% agarose gel. As a rule of thumb, if the libraries were visible from the gel (even if only barely visible), they were sent to the DNA Sequencing Core for further quantification. Libraries were walked to the on-campus University of Michigan DNA Sequencing Core immediately in ambient temperature, or stored in −20°C for less than a month before walking to the sequencing core in ambient temperature.

Sample curation (less than 1 h per sample)

We store all RNAs in a −80°C freezer on standard storage racks. Ideally, they would be stored long-term in liquid nitrogen vapor freezers. To prevent freeze/thaw of sensitive samples, we placed samples into labeled cardboard freezer boxes and recorded the sample locations in a database that is properly backed up (Appendix 7).

CONCLUSIONS

We have developed an effective phylotranscriptomics workflow involving cryogenic tissue collection in the field, RNA extraction of diverse taxa with close to 100% success rate, library preparation for Illumina platforms, and sample storage and curation. Future efforts should focus on streamlining the workflow given specific laboratory and field settings and as sequencing technologies continue to evolve. In addition, it would be ideal to collaborate with major tissue and seed banks such as the Millennium Seed Bank (Royal Botanic Gardens, Kew) and the Global Genome Initiative (Smithsonian Institution) (Gostel et al., 2016) when designing phylotranscriptomic projects.

Appendix 1.

Voucher information for the accessions used in this study.

Family	Taxon name	Collection or accession no. (Herbarium)[a,b]	Collection locality	Field or cultivated?	RNA extraction	SRA accession no. (Publication)
Achatocarpaceae	Achatocarpus gracilis H. Walter	Michael J. Moore et al. 2704 (OC)	Jalisco, Mexico: La Huerta, Estacion Biologica Chamela, along Sendero Perico	Field	PureLink	Unpublished
Achatocarpaceae	Phaulothamnus spinescens A. Gray	Michael J. Moore 1677 (OC)	Texas, USA: Kleberg, along Kleberg County Rd. 1155 S, approx. 0.1 mi. N of jct. w/ FM 771.	Field	Bio-Rad	SRX998856 (Brockington et al., 2015)
Agdestidaceae	Agdestis clematidea Moc. & Sessé ex DC.	Michael J. Moore et al. 2669 (OC)	Veracruz, Mexico: San Andres Tuxtla, in thicket immediately adjacent to rd. in Montepio	Field	PureLink	Unpublished
Amaranthaceae	Froelichia latifolia R. A. McCauley	Michael J. Moore 1665 (OC)	Texas, USA: Caldwell, along FM 713 betw. McMahan and Delhi, just W of jct. w/ Taylorville Rd.	Field	Bio-Rad	SRX998855 (Brockington et al., 2015)
Amaranthaceae	Gomphrena decumbens Jacq.	Michael J. Moore et al. 2734 (OC)	Distrito Federal, Mexico: Coyoacan, at Instituto de Biologia at UNAM	Cultivated	PureLink	Unpublished
Amaranthaceae	Gossypianthus lanuginosus (Poir.) Moq.	Michael J. Moore 1807 (OC)	Texas, USA: Llano, along TX 71 at roadside historical marker, where Honey Creek passes under the hwy.	Field	Bio-Rad	Unpublished
Amaranthaceae	Guilleminea densa (Humb. & Bonpl. ex Schult.) Moq.	Michael J. Moore et al. 2445 (OC)	Chihuahua, Mexico: N end of Sierra de Fernando, on Rancho Puerto de Lobos	Field	Bio-Rad	Unpublished
Amaranthaceae	Iresine arbuscula Uline & W. L. Bray	Michael J. Moore et al. 2678 (OC)	Veracruz, Mexico: San Andres Tuxtla, Estacion Biologica Los Tuxtlas, along stream bed approx. 100 m upstream of path crossing	Field	PureLink	Unpublished
Amaranthaceae	Iresine rhizomatosa Standl.	Michael J. Moore & J. Lee 2943 (OC)	Cultivated at Missouri Botanical Garden	Cultivated	PureLink	Unpublished
Amaranthaceae	Nelsia quadrangula (Engl.) Schinz	Millennium Seed Bank accession 0468510	Cultivated at Cambridge University Botanic Garden	Cultivated	PureLink	Unpublished
Amaranthaceae	Nitrophila occidentalis (Moq.) S. Watson	Michael J. Moore et al. 3242 (OC)	California, USA: Inyo, along CA 190 approx. 1.3 mi. NE of jct. w/ US 395 in Olancha, just S of Owens Lake bed	Field	PureLink	Unpublished
Amaranthaceae	Polycnemum majus A. Braun	Gudrun Kadereit s.n.	Cultivated at Universitat Mainz	Cultivated	PureLink	Unpublished
Amaranthaceae	Tidestromia lanuginosa (Nutt.) Standl.	Michael J. Moore et al. 2259 (OC)	New Mexico, USA: Sierra, E side of Caballo Mountains, ca. 0.3 mi. N of jct. of Slater Rd. and Apache Gap Ranch entrance	Field	Bio-Rad	Unpublished
Anacampserotaceae	Grahamia kurtzii (Bacigalupo) G. D. Rowley	Sukkulenten-Sammlung Zürich accession 100046/0	Cultivated at Sukkulenten-Sammlung Zürich	Cultivated	Bio-Rad	Unpublished
Anacampserotaceae	Talinopsis frutescens A. Gray	Michael J. Moore et al. 2441 (OC)	Chihuahua, Mexico: N end of Sierra de Fernando, on Rancho Puerto de Lobos	Field	Trizol	Unpublished
Ancistrocladaceae	Ancistrocladus robertsoniorum J. Léonard	Michael J. Moore & J. Lee 2940 (OC)	Missouri Botanical Garden, cultivated in Climatron	Cultivated	PureLink	Unpublished
Apiaceae	Heracleum mantegazzianum Sommier & Levier	Hannah E. Marx 2014-016 (UI)	Provence-Alpes-Côte d’Azur, France: Hautes-Alpes, Villar-d’Arêne: Jardin Botanique Alpin du Lautaret: invaded plot	Field	PureLink	Unpublished
Apiaceae	Heracleum sphondylium L.	Hannah E. Marx 2014-010 (UI)	Provence-Alpes-Côte d’Azur, France: Hautes-Alpes, Villar-d’Arêne: Jardin Botanique Alpin du Lautaret: native plot	Field	PureLink	Unpublished
Apiaceae	Meum athamanticum Jacq.	Hannah E. Marx 2014-027 (UI)	Provence-Alpes-Côte d’Azur, France: Hautes-Alpes, Villar-d’Arêne: Jardin Botanique Alpin du Lautaret: native plot	Field	PureLink	Unpublished
Asteraceae	Centaurea uniflora Turra	Hannah E. Marx 2014-028 (UI)	Provence-Alpes-Côte d’Azur, France: Hautes-Alpes, Villar-d’Arêne: Jardin Botanique Alpin du Lautaret: native plot	Field	PureLink	Unpublished
Asteraceae	Gaillardia multiceps Greene	Michael J. Moore 1737 (OC)	Texas, USA: Winkler, along Winkler County Rd. 101, ca. 5 mi. N of jct. w/ TX 302	Field	Bio-Rad	Unpublished
Basellaceae	Anredera cordifolia (Ten.) Steenis	Cultivated at Cambridge University Botanic Garden 19770198	Cultivated at Cambridge University Botanic Garden	Cultivated	PureLink	Unpublished
Boraginaceae	Tiquilia hispidissima (Torr. & A. Gray) A. T. Richardson	Michael J. Moore 1736 (OC)	Texas, USA: Winkler, along Winkler County Rd. 101, ca. 5 mi. N of jct. w/ TX 302	Field	Bio-Rad	Unpublished
Brassicaceae	Nerisyrenia linearifolia (S. Watson) Greene	Michael J. Moore 1755 (OC)	Texas, USA: Culberson, along FM 652 ca. 25 mi. W of Orla	Field	Bio-Rad	Unpublished
Cactaceae	Opuntia arenaria Engelm.	Michael J. Moore et al. 2911 (OC)	New Mexico, USA: Dona Ana, cultivated at UTEP Botanical Garden; originally collected from Anapra, NM	Cultivated	PureLink	Unpublished
Cactaceae	Rhipsalis baccifera (Sol.) Stearn subsp. baccifera	Michael J. Moore 2938 (OC)	Cultivated at Oberlin College greenhouse	Cultivated	PureLink	Unpublished
Caryophyllaceae	Arenaria serpyllifolia L.	Michael J. Moore 1164 (OC)	Ohio, USA: Erie, in lawn behind my house (4910 State Route 113 E)	Field	Bio-Rad	Unpublished
Caryophyllaceae	Cerastium alpinum L. var. lanatum (Lam.) Hegetschw.	Alplains accession 07471.06	Cultivated at Matthaei Botanical Gardens, University of Michigan	Cultivated	PureLink	Unpublished
Caryophyllaceae	Cerastium arvense L.	Michael J. Moore 1767 (OC)	New Mexico, USA: Otero, along NM 130 a few mi. E of jct. w/ rd. to Sunspot	Field	Bio-Rad	SRX998858 (Brockington et al., 2015)
Caryophyllaceae	Cerastium fontanum Baumg. subsp. vulgare (Hartm.) Greuter & Burdet	Michael J. Moore 1163 (OC)	Ohio, USA: Erie, in lawn behind my house (4910 State Route 113 E)	Field	Bio-Rad	Unpublished
Caryophyllaceae	Corrigiola litoralis L.	Cultivated at Cambridge University Botanic Garden SFB_221	Cultivated at Cambridge University Botanic Garden	Cultivated	PureLink	Unpublished
Caryophyllaceae	Drymaria cordata (L.) Willd. ex Schult.	Lucas C. Majure 3403 (FLAS)	Florida, USA: Alachua, Gainesville, University of Florida campus	Field	Bio-Rad	SRX998854 (Brockington et al., 2015)
Caryophyllaceae	Drymaria sp.	Michael J. Moore et al. 2679 (OC)	Veracruz, Mexico: Coscomatepec, along paved rd. from Fortin to Huatusco, a few km N of Coscomatepec	Field	PureLink	Unpublished
Caryophyllaceae	Drymaria subumbellata I. M. Johnst.	Michael J. Moore et al. 2503 (OC)	Durango, Mexico: on the W side of the Sierra de Tlahualilo	Field	Bio-Rad	Unpublished
Caryophyllaceae	Eremogone hookeri (Nutt. ex Torr. & A. Gray) W. A. Weber subsp. desertorum (Maguire) W. A. Weber	Alplains accession 35665.38	Cultivated at Matthaei Botanical Gardens, University of Michigan	Cultivated	Bio-Rad	Unpublished
Caryophyllaceae	Eremogone procera (Spreng.) Rchb.	Gudrun Kadereit s.n.	Cultivated at Cambridge University Botanic Garden	Cultivated	PureLink	Unpublished
Caryophyllaceae	Gypsophila repens L.	Cambridge University Botanic Garden 10007025, 19610163, 19860039	Cultivated at Cambridge University Botanic Garden	Cultivated	PureLink	Unpublished
Caryophyllaceae	Herniaria latifolia Lapeyr.	Cambridge University Botanic Garden 10005718	Cultivated at Cambridge University Botanic Garden	Cultivated	PureLink	Unpublished
Caryophyllaceae	Honckenya peploides (L.) Ehrh.	Millennium Seed Bank accession 0286981	Cultivated at Cambridge University Botanic Garden	Cultivated	PureLink	Unpublished
Caryophyllaceae	Illecebrum verticillatum L.	N/A	Cultivated at Cambridge University Botanic Garden	Cultivated	PureLink	Unpublished
Caryophyllaceae	Lepyrodiclis stellarioides Fisch. & C. A. Mey.	Millennium Seed Bank accession 0653842	Cultivated at Cambridge University Botanic Garden	Cultivated	PureLink	Unpublished
Caryophyllaceae	Paronychia drummondii Torr. & A. Gray	Michael J. Moore 1670 (OC)	Texas, USA: Caldwell, along FM 713 between McMahan and Delhi, just W of jct. w/ Taylorville Rd.	Field	PureLink	Unpublished
Caryophyllaceae	Paronychia jamesii Torr. & A. Gray	Michael J. Moore et al. 2931 (OC)	New Mexico, USA: Eddy, along gravel rd. leading W from US 285 along N edge of Seven Rivers Hills	Field	PureLink	Unpublished
Caryophyllaceae	Schiedea globosa H. Mann	Cambridge University Botanic Garden SFB_256	Cultivated at Cambridge University Botanic Garden	Cultivated	PureLink	Unpublished
Caryophyllaceae	Scleranthus polycarpos L.	Gudrun Kadereit s.n.	Cultivated at Cambridge University Botanic Garden	Cultivated	PureLink	Unpublished
Caryophyllaceae	Silene acaulis (L.) Jacq. subsp. acaulescens Hitchc. & Maquire	Alplains accession 01224.31	Cultivated at Matthaei Botanical Gardens, University of Michigan	Cultivated	Bio-Rad	Unpublished
Caryophyllaceae	Spergularia marina (L.) Besser	Michael J. Moore et al. 3185 (OC)	California, USA: Kern, Tejon Ranch: Amargo Springs	Field	PureLink	Unpublished
Caryophyllaceae	Telephium imperati L.	Cambridge University Botanic Garden 19910346	Cultivated at Cambridge University Botanic Garden	Cultivated	PureLink	Unpublished
Caryophyllaceae	Velezia rigida L.	Gudrun Kadereit s.n.	Cultivated at Cambridge University Botanic Garden	Cultivated	PureLink	Unpublished
Chenopodiaceae	Anabasis articulata (Forssk.) Moq.	LS168	Cultivated at Universitat Mainz	Cultivated	PureLink	Unpublished
Chenopodiaceae	Arthrocnemum macrostachyum (Moric.) K. Koch	LS33	Cultivated at Universitat Mainz	Cultivated	PureLink	Unpublished
Chenopodiaceae	Atriplex sp.	Michael J. Moore et al. 3295 (OC)	Utah, USA: Wayne, a few hundred yards W of Coal Mine Rd., approx. 3.2 mi. N of jct. w/ UT 24	Field	PureLink	Unpublished
Chenopodiaceae	Caroxylon vermiculatum (L.) Akhani & Roalson	LS178	Cultivated at Universitat Mainz	Cultivated	PureLink	Unpublished
Chenopodiaceae	Chenopodiastrum murale (L.) S. Fuentes, Uotila & Borsch	Michael J. Moore et al. 2991 (OC)	New Mexico, USA: Chaves, along gravel rd. just N of US 380 opposite rest area	Field	PureLink	Unpublished
Chenopodiaceae	Corispermum hyssopifolium L.	Millennium Seed Bank accession 0000170	Cultivated at Cambridge University Botanic Garden	Cultivated	PureLink	Unpublished
Chenopodiaceae	Eokochia saxicola (Guss.) Freitag & G. Kadereit	LS70	Cultivated at Universitat Mainz	Cultivated	PureLink	Unpublished
Chenopodiaceae	Extriplex californica Moq.	Michael J. Moore et al. 3214 (OC)	California, USA: San Luis Obispo, Morro Bay State Park/Morro Estuary Natural Preserve: just E of State Park Rd./Main St., approx. 0.7 mi. SW of jct. w/ S Bay Blvd.	Field	Bio-Rad	Unpublished
Chenopodiaceae	Grayia spinosa (Hook.) Moq.	Michael J. Moore et al. 3268 (OC)	Nevada, USA: Douglas, near two-track rd. that runs S from Mel Drive	Field	Bio-Rad	Unpublished
Chenopodiaceae	Kali collina (Pall.) Akhani & Roalson	Millennium Seed Bank accession 0496298	Cultivated at Cambridge University Botanic Garden	Cultivated	PureLink	Unpublished
Chenopodiaceae	Kalidium cuspidatum (Ung.-Sternb.) Grubov	LS97	Cultivated at Universitat Mainz	Cultivated	PureLink	Unpublished
Chenopodiaceae	Krascheninnikovia lanata (Pursh) A. Meeuse & A. Smit	Michael J. Moore et al. 2311 (OC)	New Mexico, USA: Sierra, along WSMR Rte. 6 just S of Big Gyp Mountain	Field	Trizol	Unpublished
Chenopodiaceae	Salsola sp.	N/A	Cultivated at Cambridge University Botanic Garden	Cultivated	PureLink	Unpublished
Chenopodiaceae	Sarcocornia pacifica (Standl.) A. J. Scott	Michael J. Moore et al. 3216 (OC)	California, USA: San Luis Obispo, Morro Bay State Park/Morro Estuary Natural Preserve: just E of State Park Rd./Main St., approx. 0.7 mi. SW of jct. w/ S Bay Blvd.	Field	PureLink	Unpublished
Chenopodiaceae	Stutzia covillei (Torr. ex S. Watson) S. Watson	Michael J. Moore et al. 3228 (OC)	California, USA: Inyo, along Searles Dry Lake Rd., approx. 0.5 mi. E of jct. w/ Trona-Wildrose Rd.	Field	PureLink	Unpublished
Chenopodiaceae	Suaeda linearis (Elliott) Moq.	Michael J. Moore 1679 (OC)	Texas, USA: Kleberg, Riviera Beach, along beach just N of parking area at end of FM 771. GPS coordinates refer to parking area.	Field	Bio-Rad	Unpublished
Chenopodiaceae	Tecticornia pergranulata (J. M. Black) K. A. Sheph. & Paul G. Wilson	LS28	Cultivated at Universitat Mainz	Cultivated	PureLink	Unpublished
Cyperaceae	Carex capillaris L.	Hannah E. Marx 2014-030 (UI)	Provence-Alpes-Côte d’Azur, France: Hautes-Alpes, Villar-d’Arêne: Jardin Botanique Alpin du Lautaret: native plot	Field	PureLink	Unpublished
Cyperaceae	Carex nigra (L.) Reichard	Hannah E. Marx 2014-031 (UI)	Provence-Alpes-Côte d’Azur, France: Hautes-Alpes, Villar-d’Arêne: Jardin Botanique Alpin du Lautaret: native plot	Field	PureLink	Unpublished
Didiereaceae	Alluaudia dumosa (Drake) Drake	Desert Botanical Garden accession 1987-0301-0201	Cultivated at Desert Botanical Garden	Cultivated	PureLink	Unpublished
Didiereaceae	Alluaudia humbertii Choux	Desert Botanical Garden accession 1974-0229-01-1	Cultivated at Desert Botanical Garden	Cultivated	PureLink	Unpublished
Didiereaceae	Alluaudia procera (Drake) Drake	Desert Botanical Garden accession 1956-5744-02-3 G	Cultivated at Desert Botanical Garden	Cultivated	PureLink	Unpublished
Didiereaceae	Alluaudiopsis marnieriana Rauh	Sukkulenten-Sammlung Zürich accession 81 2213/0	Cultivated at Sukkulenten-Sammlung Zürich	Cultivated	Hot acid Phenol	Unpublished
Didiereaceae	Ceraria pygmaea (Pillans) G. D. Rowley	Sukkulenten-Sammlung Zürich accession 90 1893/b	Cultivated at Sukkulenten-Sammlung Zürich	Cultivated	Hot acid Phenol	Unpublished
Didiereaceae	Decarya madagascariensis Choux	Michael J. Moore & J. Lee 2944 (OC)	Missouri Botanical Garden, cultivated in nonpublic arid plant greenhouse	Cultivated	PureLink	Unpublished
Didiereaceae	Didierea madagascariensis Baill.	Desert Botanical Garden accession 1996-0312-01-1	Cultivated at Desert Botanical Garden	Cultivated	PureLink	Unpublished
Didiereaceae	Didierea trollii Capuron & Rauh	Desert Botanical Garden accession 1984-0024-0202 G	Cultivated at Desert Botanical Garden	Cultivated	PureLink	Unpublished
Didiereaceae	Portulacaria afra Jacq.	Desert Botanical Garden accession 1988-0583-02-1 G	Cultivated at Desert Botanical Garden	Cultivated	PureLink	Unpublished
Droseraceae	Aldrovanda vesiculosa L.	Michael J. Moore 1652 (OC)	Virginia, USA: Caroline, ponds at Meadowview Biological Station	Cultivated	Bio-Rad	SRX998847 (Brockington et al., 2015)
Droseraceae	Drosera binata Labill.	Cambridge University Botanic Garden	Cultivated at Cambridge University Botanic Garden	Cultivated	PureLink	Unpublished
Droseraceae	Drosera burmannii Vahl	Matthaei Botanical Gardens	Cultivated at Matthaei Botanical Gardens, University of Michigan	Cultivated	PureLink	Unpublished
Drosophyllaceae	Drosophyllum lusitanicum (L.) Link	Cambridge University Botanic Garden	Cultivated at Cambridge University Botanic Garden	Cultivated	PureLink	Unpublished
Fabaceae	Lotus corniculatus L.	Hannah E. Marx 2014-026 (UI)	Provence-Alpes-Côte d’Azur, France: Hautes-Alpes, Villar-d’Arêne: Jardin Botanique Alpin du Lautaret: native plot	Field	PureLink	Unpublished
Frankeniaceae	Frankenia salina (Molina) I. M. Johnst.	Michael J. Moore et al. 3209 (OC)	California, USA: San Luis Obispo, Morro Bay State Park/Morro Estuary Natural Preserve: just E of State Park Rd./Main St., approx. 0.7 mi. SW of jct. w/ S Bay Blvd.	Field	PureLink	Unpublished
Geraniaceae	Geranium sylvaticum L.	Hannah E. Marx 2014-014 (UI)	Provence-Alpes-Côte d’Azur, France: Hautes-Alpes, Villar-d’Arêne: Jardin Botanique Alpin du Lautaret: native plot	Field	PureLink	Unpublished
Gisekiaceae	Gisekia pharnaceoides L.	Millennium Seed Bank accession 0586315	Cultivated at Cambridge University Botanic Garden	Cultivated	PureLink	Unpublished
Hydrophyllaceae	Nama carnosa (Woot.) C. L. Hitchc.	Michael J. Moore 1750 (OC)	Texas, USA: Culberson, along FM 652 ca. 25 mi. W of Orla	Field	Bio-Rad	Unpublished
Kewaceae	Kewa bowkeriana (Sond.) Christenh.	Botanischen Gartens, Technische Universität Dresden accession 99 Bonn 931 (B:09964)	Cultivated at Cambridge University Botanic Garden	Cultivated	Hot acid Phenol	Unpublished
Lentibulariaceae	Pinguicula vulgaris L.	Hannah E. Marx 2014-021 (UI)	Provence-Alpes-Côte d’Azur, France: Hautes-Alpes, Villar-d’Arêne: Jardin Botanique Alpin du Lautaret: native plot	Field	PureLink	Unpublished
Limeaceae	Limeum aethiopicum Burm. f.	Cultivated at Cambridge University Botanic Garden s.n.	Cultivated at Cambridge University Botanic Garden	Cultivated	Hot acid Phenol	Unpublished
Loasaceae	Mentzelia humilis (Urb. & Gilg) J. Darl.	Michael J. Moore 1749 (OC)	Texas, USA: Culberson, along FM 652 ca. 25 mi. W of Orla	Field	Bio-Rad	Unpublished
Macarthuriaceae	Macarthuria australis Hügel ex Endl.	Kevin Thiele 5141 (UWA)	Western Australia, Australia: Perth. Cultivated at Cambridge University Botanic Garden.	Cultivated	PureLink	Unpublished
Melanthiaceae	Veratrum album L.	Hannah E. Marx 2014-025 (UI)	Provence-Alpes-Côte d’Azur, France: Hautes-Alpes, Villar-d’Arêne: Jardin Botanique Alpin du Lautaret: native plot	Field	PureLink	Unpublished
Molluginaceae	Glinus lotoides L. var. virens Fenzl	Millennium Seed Bank accession 0197698	Cultivated at Cambridge University Botanic Garden	Cultivated	PureLink	Unpublished
Molluginaceae	Pharnaceum exiguum Adamson	Millennium Seed Bank accession 0467649	Cultivated at Cambridge University Botanic Garden	Cultivated	PureLink	Unpublished
Molluginaceae	Suessenguthiella caespitosa Friedrich	Millennium Seed Bank accession 0467650	Cultivated at Cambridge University Botanic Garden	Cultivated	PureLink	Unpublished
Montiaceae	Calandrinia grandiflora	Cambridge University Botanic Garden 20070025	Cultivated at Cambridge University Botanic Garden	Cultivated	PureLink	Unpublished
Montiaceae	Calyptridium pygmaeum Parish ex Rydb.	Michael J. Moore et al. 3165 (OC)	California, USA: San Bernardino, approx. 0.2 mi. N of Forest Rd. 2N86, approx. 0.4 mi. NE of Bluff Lake	Field	PureLink	Unpublished
Montiaceae	Calyptridium umbellatum (Torr.) Hershkovitz	Michael J. Moore et al. 3142 (OC)	California, USA: San Bernardino, Mt. Baldy ski area	Field	Bio-Rad	Unpublished
Montiaceae	Cistanthe grandiflora (Lindl.) Schltdl.	Chileflora s.n.	Cultivated at Cambridge University Botanic Garden	Cultivated	PureLink	Unpublished
Montiaceae	Claytonia nevadensis S. Watson	Thomas R. Stoughton et al. 2027 (RSA)	California, USA: Alpine, about 250 yards W of Round Top Lake, near trail	Field	Bio-Rad	Unpublished
Montiaceae	Claytonia virginica L.	Michael J. Moore 1156 (OC)	Ohio, USA: Erie, in woods behind my house (4910 State Route 113 E)	Field	Bio-Rad	Unpublished
Montiaceae	Lewisia nevadensis (A. Gray) B. L. Rob.	Michael J. Moore et al. 3168 (OC)	California, USA: San Bernardino, near edge of meadow, along small creek that feeds into Bluff Lake	Field	PureLink	Unpublished
Montiaceae	Montia chamissoi (Ledeb. ex Spreng.) Greene	Michael J. Moore et al. 3167 (OC)	California, USA: San Bernardino, near edge of meadow, along small creek that feeds into Bluff Lake	Field	PureLink	Unpublished
Montiaceae	Phemeranthus parviflorus (Nutt.) Kiger	Michael J. Moore et al. 2214 (OC)	New Mexico, USA: Socorro, just E of Quebradas Backcountry Byway	Field	Trizol	Unpublished
Nepenthaceae	Nepenthes alata Blanco	Cambridge University Botanic Garden 20160588, 20160946	Cultivated at Cambridge University Botanic Garden	Cultivated	PureLink	Unpublished
Nepenthaceae	Nepenthes ventricosa Blanco	Cambridge University Botanic Garden 20050134	Cultivated at Cambridge University Botanic Garden	Cultivated	PureLink	Unpublished
Nyctaginaceae	Abronia bigelovii Heimerl	Michael J. Moore et al. 2189 (OC)	New Mexico, USA: Sandoval, just N of Cabezon Rd., N of White Mesa Bike Trails	Field	Bio-Rad	Unpublished
Nyctaginaceae	Abronia fragrans Nutt. ex Hook.	Michael J. Moore et al. 2992 (OC)	New Mexico, USA: Chaves, along gravel rd. just N of US 380 opposite rest area	Field	PureLink	Unpublished
Nyctaginaceae	Abronia glabrifolia Standl.	Michael J. Moore et al. 3318 (OC)	Colorado, USA: Mesa, along Mitchell Rd. approx. 1.0 mi. W of jct. w/ CO 139	Field	PureLink	Unpublished
Nyctaginaceae	Abronia latifolia Eschsch.	Michael J. Moore et al. 3204 (OC)	California, USA: San Luis Obispo, Montana de Oro State Park: along path from parking lot at end of Sand Spit Rd. to beach, just up the dunes from the beach	Field	PureLink	Unpublished
Nyctaginaceae	Abronia maritima Nutt. ex S. Watson	Michael J. Moore et al. 3217 (OC)	California, USA: San Luis Obispo, Morro Strand State Beach: just W of Hwy. 1 in Cayucos	Field	Trizol	Unpublished
Nyctaginaceae	Abronia nealleyi Standl.	Michael J. Moore 1751 (OC)	Texas, USA: Culberson, along FM 652 ca. 25 mi. W of Orla	Field	Bio-Rad	SRX998850 (Brockington et al., 2015)
Nyctaginaceae	Abronia umbellata Lam.	Michael J. Moore et al. 3203 (OC)	California, USA: San Luis Obispo, Montana de Oro State Park: along path from parking lot at end of Sand Spit Rd. to beach, approx. 100 yards from parking lot	Field	PureLink	Unpublished
Nyctaginaceae	Acleisanthes acatitensis M. J. Moore & E. Loke (ined.)	Michael J. Moore et al. 2505 (OC)	Durango, Mexico: on the W side of the Sierra de Tlahualilo	Field	PureLink	Unpublished
Nyctaginaceae	Acleisanthes acutifolia Standl.	Michael J. Moore et al. 2447 (OC)	Chihuahua, Mexico: N end of Sierra de Fernando, on Rancho Puerto de Lobos	Field	Bio-Rad	Unpublished
Nyctaginaceae	Acleisanthes chenopodioides (A. Gray) R. A. Levin	Michael J. Moore et al. 2246 (OC)	New Mexico, USA: Sierra, Armendaris Ranch, along rd. leading to highest point in Fra Cristobal Mountains	Field	Bio-Rad	Unpublished
Nyctaginaceae	Acleisanthes lanceolata (Wooton) R. A. Levin var. lanceolata	Michael J. Moore 1741 (OC)	Texas, USA: Winkler, along Winkler County Rd. 101, ca. 5 mi. N of jct. w/ TX 302	Field	Bio-Rad	SRX998849 (Brockington et al., 2015)
Nyctaginaceae	Acleisanthes obtusa (Choisy) Standl.	Michael J. Moore 1697 (OC)	Texas, USA: Zapata, immediately behind the Holiday Inn Express on the S side of Zapata, along US 83	Field	Bio-Rad	SRX998848 (Brockington et al., 2015)
Nyctaginaceae	Anulocaulis annulatus (Coville) Standl.	Michael J. Moore et al. 3237a (OC)	California, USA: Inyo, mouth of Surprise Canyon	Field	PureLink	Unpublished
Nyctaginaceae	Anulocaulis eriosolenus (A. Gray) Standl.	Michael J. Moore et al. 2362 (OC)	Chihuahua, Mexico: along MEX 16 in Sierra Peguis	Field	Bio-Rad	Unpublished
Nyctaginaceae	Anulocaulis leiosolenus (Torr.) Standl. var. gypsogenus (Waterf.) Spellenb. & T. Wootten	Michael J. Moore 1070 (OC)	Texas, USA: Culberson, along FM 652 25 mi. W of jct. w/ US 285 in Orla	Field	Bio-Rad	SRX717838 (Yang et al., 2015)
Nyctaginaceae	Boerhavia ciliata Brandegee	Michael J. Moore et al. 2760 (OC)	Nuevo Leon, Mexico: Mier y Noriega, large gypsum exposure about 35 km SSW of Doctor Arroyo	Field	PureLink	Unpublished
Nyctaginaceae	Boerhavia purpurascens A. Gray	Michael J. Moore et al. 2201 (OC)	New Mexico, USA: Socorro, along wash E of County Rd. 12	Field	Bio-Rad	Unpublished
Nyctaginaceae	Boerhavia torreyana (S. Watson) Standl.	Michael J. Moore et al. 2202 (OC)	New Mexico, USA: Socorro, along wash E of County Road 12	Field	Bio-Rad	Unpublished
Nyctaginaceae	Bougainvillea stipitata Griseb. var. grisebachiana Heimerl	Kew Living Collection #1986-4920	Cultivated at Royal Botanic Gardens, Kew	Cultivated	PureLink	SRX718672 (Yang et al., 2015)
Nyctaginaceae	Colignonia ovalifolia Heimerl	Crug Farms accession BSWJ10644	Cultivated at Cambridge University Botanic Garden	Cultivated	Hot acid Phenol	Unpublished
Nyctaginaceae	Commicarpus scandens (L.) Standl.	Michael J. Moore et al. 2726B (OC)	Jalisco, Mexico: Autlan de Navarro, along gravel rd. leading from the outskirts of El Grullo to El Chacalito	Field	PureLink	Unpublished
Nyctaginaceae	Cyphomeris gypsophiloides (M. Martens & Galeotti) Standl.	Michael J. Moore 1714 (OC)	Texas, USA: Val Verde, along US 90 W of Langtry	Field	Bio-Rad	SRX998857 (Brockington et al., 2015)
Nyctaginaceae	Guapira obtusata (Jacq.) Little	Kew Living Collection #2011-994	Cultivated at Royal Botanic Gardens, Kew	Cultivated	PureLink	SRX718384 (Yang et al., 2015)
Nyctaginaceae	Mirabilis multiflora (Torr.) A. Gray	Michael J. Moore 1771 (OC)	New Mexico, USA: Lincoln, along NM 55 approx. 0.5 mi. W of jct. w/ US 54	Field	Bio-Rad	SRX998851 (Brockington et al., 2015)
Nyctaginaceae	Mirabilis pringlei Weath.	Michael J. Moore et al. 2725 (OC)	Jalisco, Mexico: Autlan de Navarro, along gravel rd. leading from the outskirts of El Grullo to El Chacalito	Field	PureLink	Unpublished
Nyctaginaceae	Neea psychotrioides Donn. Sm.	Michael J. Moore et al. 2675 (OC)	Veracruz, Mexico: San Andres Tuxtla, Estacion Biologica Los Tuxtlas, near the collections building	Field	PureLink	Unpublished
Nyctaginaceae	Nyctaginia capitata Choisy	Michael J. Moore et al. 2585 (OC)	Coahuila, Mexico: in Valle Padilla along rd. leading W away from ranch house	Field	Bio-Rad	Unpublished
Nyctaginaceae	Okenia hypogaea Schltdl. & Cham.	Michael J. Moore et al. 2673 (OC)	Veracruz, Mexico: San Andres Tuxtla, along the face and near the base of large foredunes near the beach	Field	PureLink	Unpublished
Nyctaginaceae	Pisonia aculeata L.	Kew Living Collection #2011-448	Cultivated at Royal Botanic Gardens, Kew	Cultivated	PureLink	SRX718389 (Yang et al., 2015)
Nyctaginaceae	Pisonia umbellifera (J. R. Forst. & G. Forst.) Seem.	Kew Living Collection #1986-3623	Cultivated at Royal Botanic Gardens, Kew, United Kingdom	Cultivated	PureLink	SRX998852 (Brockington et al., 2015)
Nyctaginaceae	Salpianthus purpurascens (Cav. ex Lag.) Hook. & Arn.	Michael J. Moore et al. 2724 (OC)	Jalisco, Mexico: Autlan de Navarro, along arroyo bed across the street from the campus of CUCSUR in Autlan de Navarro	Field	PureLink	Unpublished
Nyctaginaceae	Tripterocalyx carneus (Greene) L. A. Galloway	Michael J. Moore et al. 3027 (OC)	New Mexico, USA: Socorro, along US 380 at picnic area a few mi. W of Bingham	Field	PureLink	Unpublished
Nyctaginaceae	Tripterocalyx crux-maltae (Kellogg) Standl.	Michael J. Moore et al. 3267 (OC)	Nevada, USA: Douglas, approx. 0.6 mi. S along two-track road that runs S from Mel Drive	Field	Bio-Rad	Unpublished
Orchidaceae	Dactylorhiza alpestris (Pugsley) Aver.	Hannah E. Marx 2014-019 (UI)	Provence-Alpes-Côte d’Azur, France: Hautes-Alpes, Villar-d’Arêne: Jardin Botanique Alpin du Lautaret: native plot	Field	PureLink	Unpublished
Orobanchaceae	Bartsia alpina L.	Hannah E. Marx 2014-020 (UI)	Provence-Alpes-Côte d’Azur, France: Hautes-Alpes, Villar-d’Arêne: Jardin Botanique Alpin du Lautaret: native plot	Field	PureLink	Unpublished
Petiveriaceae	Monococcus echinophorus F. Muell.	I. R. Telford & G. Butler 9043 (CANB)	Queensland, Australia: Moreton, Mount French, 6 km SW of Boonah. [Cultivated at Booderee National Park, Jervis Bay Territory.]	Cultivated	PureLink	Unpublished
Petiveriaceae	Rivina humilis L.	Michael J. Moore 1651 (OC)	Cultivated	Cultivated	Bio-Rad	SRX718277 (Yang et al., 2015)
Petiveriaceae	Seguieria aculeata Jacq.	Kew Living Collection #1991-169	Cultivated at Royal Botanic Gardens, Kew	Cultivated	PureLink	SRX718486 (Yang et al., 2015)
Petiveriaceae	Trichostigma octandrum (L.) H. Walter	Michael J. Moore et al. 3358 (OC)	Florida, USA: Miami-Dade, The Kampong: S fenceline immediately adjacent to Biscayne Bay	Cultivated	PureLink	Unpublished
Phytolaccaceae	Anisomeria littoralis (Poepp. & Endl.) Moq.	Chileflora s.n.	Cultivated at Cambridge University Botanic Garden	Cultivated	PureLink	Unpublished
Phytolaccaceae	Ercilla volubilis (Bertero) Moq.	Michael J. Moore 1649 (OC)	Oberlin greenhouse	Cultivated	Bio-Rad	SRX998846 (Brockington et al., 2015)
Phytolaccaceae	Phytolacca dioica L.	Kew Living Collection #1963-34101	Cultivated at Royal Botanic Gardens, Kew, United Kingdom	Cultivated	PureLink	SRX998853 (Brockington et al., 2015)
Plumbaginaceae	Acantholimon lycopodioides (Girard) Boiss.	Euroseeds s.n.; Cultivated at Cambridge University Botanic Garden 20150471	Cultivated at Cambridge University Botanic Garden	Cultivated	PureLink	Unpublished
Plumbaginaceae	Aegialitis annulata R. Br.	David Warmington s.n.	Queensland, Australia: near Cairns. Cultivated at Cambridge University Botanic Garden.	Cultivated	PureLink	Unpublished
Plumbaginaceae	Limonium californicum (Boiss.) A. Heller	Michael J. Moore et al. 3210 (OC)	California, USA: San Luis Obispo, Morro Bay State Park/Morro Estuary Natural Preserve: just E of State Park Rd./Main St., approx. 0.7 mi. SW of jct. w/ S Bay Blvd.	Field	PureLink	Unpublished
Plumbaginaceae	Plumbago auriculata Lam.	Michael J. Moore et al. 3360 (OC)	Florida, USA: Miami-Dade, The Kampong: betw. tennis court and main house	Cultivated	PureLink	Unpublished
Poaceae	Dactylis glomerata L.	Hannah E. Marx 2014-011 (UI)	Provence-Alpes-Côte d’Azur, France: Hautes-Alpes, Villar-d’Arêne: Jardin Botanique Alpin du Lautaret: invaded plot	Field	PureLink	Unpublished
Poaceae	Deschampsia cespitosa (L.) P. Beauv.	Hannah E. Marx 2014-012 (UI)	Provence-Alpes-Côte d’Azur, France: Hautes-Alpes, Villar-d’Arêne: Jardin Botanique Alpin du Lautaret: invaded plot	Field	PureLink	Unpublished
Polemoniaceae	Polemonium caeruleum L.	Hannah E. Marx 2014-004 (UI)	Provence-Alpes-Côte d’Azur, France: Hautes-Alpes, Villar-d’Arêne: Jardin Botanique Alpin du Lautaret: invaded plot	Field	PureLink	Unpublished
Polygonaceae	Antigonon leptopus Hook. & Arn.	Michael J. Moore 1811 (OC)	Texas, USA: Travis, in overgrown lot on N side of house at 3104 Tom Green St., Austin	Field	Bio-Rad	SRX998859 (Brockington et al., 2015)
Polygonaceae	Bistorta bistortoides (Pursh) Small	Michael J. Moore et al. 3333 (OC)	Colorado, USA: Larimer, approx. 0.25 mi. E of Forest Rd. 69, S of Red Feather Lakes	Field	PureLink	Unpublished
Polygonaceae	Chorizanthe angustifolia Nutt.	Michael J. Moore et al. 3201 (OC)	California, USA: San Luis Obispo, Montana de Oro State Park: along path from parking lot at end of Sand Spit Rd. to beach, approx. 100 yards from parking lot	Field	PureLink	Unpublished
Polygonaceae	Coccoloba pubescens L.	Michael J. Moore & J. Lee 2942 (OC)	Missouri Botanical Garden, cultivated in Climatron	Cultivated	PureLink	Unpublished
Polygonaceae	Coccoloba uvifera (L.) L.	Michael J. Moore 2665 (OC)	Oberlin greenhouse	Cultivated	PureLink	Unpublished
Polygonaceae	Dedeckera eurekensis Reveal & J. T. Howell	Michael J. Moore et al. 3138 (OC)	California, USA: Los Angeles. Cultivated at Rancho Santa Ana Botanic Garden.	Cultivated	PureLink	Unpublished
Polygonaceae	Emex spinosa (L.) Campd.	Millennium Seed Bank accession 0210373	Cultivated at Cambridge University Botanic Garden	Cultivated	PureLink	Unpublished
Polygonaceae	Eriogonum arcuatum Greene var. arcuatum	Michael J. Moore et al. 2215 (OC)	New Mexico, USA: Socorro, just E of Quebradas Backcountry Byway	Field	PureLink	Unpublished
Polygonaceae	Eriogonum callistum Reveal	Michael J. Moore et al. 3179 (OC)	California, USA: Kern, Tejon Ranch: Tehachapi Mtns., on calcareous outcrop near crest	Field	PureLink	Unpublished
Polygonaceae	Eriogonum deflexum Torr.	Lucas C. Majure and Michael J. Moore 5367 (DES)	Arizona, USA: Maricopa, Desert Botanical Garden	Field	PureLink	Unpublished
Polygonaceae	Eriogonum inflatum Torr. subsp. inflatum	Michael J. Moore et al. 3227 (OC)	California, USA: Kern, along Short Canyon Rd., approx. 2.6 mi. W of Bradys	Field	PureLink	Unpublished
Polygonaceae	Eriogonum longifolium Nutt. var. longifolium	Michael J. Moore et al. 2918 (OC)	New Mexico, USA: Eddy, Yeso Hills, on long WNW/ENE trending ridge of gypsum, approx 1.0 mi. along gravel rd. leading E from US 62/180	Field	PureLink	Unpublished
Polygonaceae	Eriogonum rotundifolium Benth.	Michael J. Moore 1769 (OC)	New Mexico, USA: Otero, next to parking lot at Holiday Inn Express, near US 54/70 jct. in southern end of Alamogordo	Field	PureLink	Unpublished
Polygonaceae	Muehlenbeckia platyclada (F. J. Müll.) Meisn.	Michael J. Moore 1170 (OC)	Cultivated at Oberlin greenhouse	Cultivated	Bio-Rad	Unpublished
Polygonaceae	Oxytheca perfoliata Torr. & A. Gray	Millennium Seed Bank accession 0266864	Cultivated at Cambridge University Botanic Garden	Cultivated	PureLink	Unpublished
Polygonaceae	Persicaria virginiana (L.) Gaertn.	Michael J. Moore 1162 (OC)	Ohio, USA: Erie, along the bank of the creek behind my house	Field	Bio-Rad	Unpublished
Polygonaceae	Podopterus cordifolius Rose & Standl.	Michael J. Moore et al. 2714 (OC)	Jalisco, Mexico: La Huerta, behind farmhouse approx. 0.5 km landward from main coastal hwy.	Field	PureLink	Unpublished
Polygonaceae	Podopterus mexicanus Humb. & Bonpl.	Michael J. Moore et al. 2721 (OC)	Jalisco, Mexico: La Huerta, Estacion Biologica Chamela, along Arroyo Colorado near the end of the Eje Central	Field	PureLink	Unpublished
Polygonaceae	Polygonum sp.	Michael J. Moore et al. 3219 (OC)	California, USA: San Luis Obispo, Morro Strand State Beach: just W of Hwy. 1 in Cayucos	Field	PureLink	Unpublished
Polygonaceae	Polygonum sp.	Michael J. Moore et al. 3263 (OC)	California, USA: alpine, along Round Top Lake Trail	Field	PureLink	Unpublished
Polygonaceae	Polygonum aviculare L.	Michael J. Moore 2667 (OC)	Ohio, USA: Lorain, adjacent to Woodland St. parking lot on campus of Oberlin College	Field	Bio-Rad	Unpublished
Polygonaceae	Polygonum dentoceras T. M. Schust. & Reveal	Michael J. Moore et al. 3353 (OC)	Florida, USA: Highlands, just S of the Denny’s parking lot on the E side of US 27, approx. 0.6 mi. N of jct. w/ Schumacher Rd./Sebring Pkwy.	Field	PureLink	Unpublished
Polygonaceae	Pterostegia drymarioides Fisch. & C. A. Mey.	Millennium Seed Bank accession 0496863	Cultivated at Cambridge University Botanic Garden	Cultivated	PureLink	Unpublished
Polygonaceae	Rumex alpinus L.	Hannah E. Marx 2014-013 (UI)	Provence-Alpes-Côte d’Azur, France: Hautes-Alpes, Villar-d’Arêne: Jardin Botanique Alpin du Lautaret: native plot	Field	PureLink	Unpublished
Polygonaceae	Rumex arifolius Aiton	Hannah E. Marx 2014-015 (UI)	Provence-Alpes-Côte d’Azur, France: Hautes-Alpes, Villar-d’Arêne: Jardin Botanique Alpin du Lautaret: native plot	Field	PureLink	Unpublished
Polygonaceae	Ruprechtia coriacea (H. Karst.) S. F. Blake	Michael J. Moore et al. 3364 (OC)	Florida, USA: Miami-Dade, The Kampong: N side of Bay Breeze Ave. near main entrance to property. Area B1. Original collecting locality: Venezuela, Sangre de Toro, Biscochito	Cultivated	PureLink	Unpublished
Polygonaceae	Ruprechtia salicifolia (Cham. & Schltdl.) C. A. Mey.	Sunshine Seeds s.n.	Cultivated at Cambridge University Botanic Garden	Cultivated	PureLink	Unpublished
Polygonaceae	Sidotheca caryophylloides (Parry) Reveal	Michael J. Moore et al. 3164 (OC)	California, USA: San Bernardino, just S of CA 18 approx. 80 yards E of jct. w/ Rim of the World Dr.	Field	PureLink	Unpublished
Polygonaceae	Stenogonum salsuginosum Nutt.	Michael J. Moore et al. 3060 (OC)	New Mexico, USA: San Juan, along Rd. 6893 approx. 2.4 mi. N of jct. w/ US 64	Field	PureLink	Unpublished
Polygonaceae	Triplaris weigeltiana (Rchb.) Kuntze	Michael J. Moore & J. Lee 2941 (OC)	Missouri Botanical Garden, cultivated in Climatron	Cultivated	PureLink	Unpublished
Primulaceae	Primula auriculata Lam.	Hannah E. Marx 2014-006 (UI)	Provence-Alpes-Côte d’Azur, France: Hautes-Alpes, Villar-d’Arêne: Jardin Botanique Alpin du Lautaret: invaded plot	Field	PureLink	Unpublished
Primulaceae	Primula farinosa L.	Hannah E. Marx 2014-001 (UI)	Provence-Alpes-Côte d’Azur, France: Hautes-Alpes, Villar-d’Arêne: Jardin Botanique Alpin du Lautaret: native plot	Field	PureLink	Unpublished
Primulaceae	Primula grandis Trautv.	Hannah E. Marx 2014-003 (UI)	Provence-Alpes-Côte d’Azur, France: Hautes-Alpes, Villar-d’Arêne: Jardin Botanique Alpin du Lautaret: invaded plot	Field	PureLink	Unpublished
Ranunculaceae	Caltha fistulosa Schipcz.	Hannah E. Marx 2014-009 (UI)	Provence-Alpes-Côte d’Azur, France: Hautes-Alpes, Villar-d’Arêne: Jardin Botanique Alpin du Lautaret: invaded plot	Field	PureLink	Unpublished
Ranunculaceae	Caltha palustris L.	Hannah E. Marx 2014-002 (UI)	Provence-Alpes-Côte d’Azur, France: Hautes-Alpes, Villar-d’Arêne: Jardin Botanique Alpin du Lautaret: native plot	Field	PureLink	Unpublished
Ranunculaceae	Ranunculus aconitifolius L.	Hannah E. Marx 2014-022 (UI)	Provence-Alpes-Côte d’Azur, France: Hautes-Alpes, Villar-d’Arêne: Jardin Botanique Alpin du Lautaret: native plot	Field	PureLink	Unpublished
Ranunculaceae	Ranunculus acris L.	Hannah E. Marx 2014-017 (UI)	Provence-Alpes-Côte d’Azur, France: Hautes-Alpes, Villar-d’Arêne: Jardin Botanique Alpin du Lautaret: native plot	Field	PureLink	Unpublished
Ranunculaceae	Ranunculus caucasicus M. Bieb.	Hannah E. Marx 2014-005 (UI)	Provence-Alpes-Côte d’Azur, France: Hautes-Alpes, Villar-d’Arêne: Jardin Botanique Alpin du Lautaret: invaded plot	Field	PureLink	Unpublished
Ranunculaceae	Trollius europaeus L.	Hannah E. Marx 2014-018 (UI)	Provence-Alpes-Côte d’Azur, France: Hautes-Alpes, Villar-d’Arêne: Jardin Botanique Alpin du Lautaret: native plot	Field	PureLink	Unpublished
Rosaceae	Alchemilla mollis (Buser) Rothm.	Hannah E. Marx 2014-007 (UI)	Provence-Alpes-Côte d’Azur, France: Hautes-Alpes, Villar-d’Arêne: Jardin Botanique Alpin du Lautaret: invaded plot	Field	PureLink	Unpublished
Rosaceae	Alchemilla xanthochlora Rothm.	Hannah E. Marx 2014-008 (UI)	Provence-Alpes-Côte d’Azur, France: Hautes-Alpes, Villar-d’Arêne: Jardin Botanique Alpin du Lautaret: native plot	Field	PureLink	Unpublished
Sarcobataceae	Sarcobatus vermiculatus (Hook.) Torr.	Michael J. Moore 1773 (OC)	New Mexico, USA: Torrance, along US 60 ca 5 mi. E of jct. w/ NM 42, near S end of Laguna del Perro	Field	Bio-Rad	Unpublished
Scrophulariaceae	Leucophyllum frutescens (Berl.) I. M. Johnst.	Michael J. Moore 1810 (OC)	Texas, USA: Travis, on the campus of the University of Texas, behind the Littlefield House	Cultivated	Bio-Rad	Unpublished
Stegnospermataceae	Stegnosperma halimifolium Benth.	Desert Botanical Garden accession 1973-0120-01-1 W	Cultivated at Desert Botanical Garden	Cultivated	Bio-Rad	Unpublished
Talinaceae	Talinum paniculatum (Jacq.) Gaertn.	Michael J. Moore 1789 (OC)	Cultivated; purchased from Oasis Nursery in Stillwater, OK	Cultivated	Bio-Rad	Unpublished

Herbaria codes are from Index Herbariorum (http://sweetgum.nybg.org/science/ih/).

For samples without herbaria listed, we have provided the accession number from the living collection (e.g., Cambridge University Botanic Garden, Sukkulenten-Sammlung Zürich), seed bank (e.g., Millennium Seed Bank), or seed supplier.