1. Quantifying genetic diversity in
Lepidospartum burgessii using microsatellites
Chloe Siegel1, Rachel Cheung2, and Evelyn Williams3
1 – University of Illinois – Urbana-Champaign, 2 – Carleton College, Northfield, MN, 3 – Chicago Botanic Garden, IL
Introduction
Materials & Methods
Population Sampling: 320 Lepidospartum burgessii samples, 65 Lepidospartum latisquamum samples
from 25 different patches
•L. burgessii from 5 patches (74 plants) in Texas and 15 patches (246 plants) in New Mexico
•L. latisquamum from 5 patches in California
DNA Extraction: Previously extracted DNA using Qiagen DNeasy Plant Mini Kits
PCR: Loci amplified by screening previously developed primers for Senecio
•29 primers were screened, 4 were used previously, 3 additional primers amplified here
•Loci amplified using two-step M13 PCR protocol
Beckman and Scoring: Ran samples with 400bp or 600bp ladder on a Beckman Coulter Ceq 8000
analysis system
•Scored loci using a peak ratio analysis and hand scoring
Results
From scoring our three additional primers on the Beckman Coulter, we have
seen a vast majority of homozygotes with little variation among alleles (Fig. 3).
Primer S23 is largely monomorphic for a single allele across populations, though
heterozygotes do occur. Primer S20 is similarly low in heterozygotes. Primer V45
is invariable for L. burgessii but likely helps separate it from L. latisquamum.
While our study has not yet been completed, data will be compared using
diversity statistics, genetic structure patterns, and genotype analyses. Based on
results that have been scored, we anticipate that similar results will arise from
the three primers we tested (Fig. 2). From this previous work, 16 out of 39
putative clonal samples were previously confirmed to be clones, which will
similarly be studied from our results (2). Our results suggest fewer heterozygotes
circulating through the L. burgessii gene pool, likely a result of inbreeding among
the individuals. We expect to see more heterozygotes among the L. latisquamum
population due to its higher population size and thus likely more diverse gene
pool.
Discussion
Previous work with fewer loci showed that L. burgessii and L. latisquamum
populations are not comparatively genetically depauperate. Our results will help confirm
this result and provide more power to differentiate populations. Asexual reproduction
likely makes up a great deal of the growth the species, but variation comparable to L.
latisquamum shows that sexual reproduction may still be occurring. In future studies it
would be useful to investigate the specific reason as to why sexual reproduction is so
stagnant now.
It has been observed that transplanted L. burgessii plants do well when tended
regularly tended (Fig. 4a). Chicago Botanic Garden scientists have already tried to
vegetatively propagate new shrubs to increase population numbers and create a
common garden for cross-pollination experiments. Vegetation propagation failed, though
there are current efforts in tissue culture. Future cross-pollination experiments will be in
the field (Fig. 4b).
We speculate that the two populations could have previously been a single grouping
given the relatively short distance between them. If the two populations could be
reintroduced to one another, sexual reproduction might be more successful if there is a
sufficient amount of genetic variation between the two populations.
Because the species was never actively monitored as one population, we don’t know
exactly what environmental conditions could contribute to population’s decline.
Environmental degradation, changing stages of succession, or the presence of insects or
fungi (Fig. 4c) could be causes for population and diversity decline. Our work will provide
a better understanding of the two populations’ genetics and inform additional courses of
action to conserve this species in its given environments.
Acknowledgements
• Mike Howard and Lepidospartum collection teams
References
1. Ladyman & Gegick. (2000) The Status of Lepidospartum burgessii (Burgess Broomshrub or
Gypsum Broomscale). In Machiniski & Holter Southwestern rare and endangered plants:
Proceedings of the Third Conference. U.S. Dept. of Agriculture, Fort Collins, CO.
2. Williams, E.W., et al. in prep. Narrowly endemic does not mean depauperate: population
genetics of the gypsophile Lepidospartum burgessii (Asteraceae).
Lepidospartum burgessii (Asteraceae) is a desert plant native to the Western United States with two
shrinking populations remaining on public lands. The species was previously studied in the 1990s, but
has recently attracted the attention of researchers for its perplexing lack of seed production. The only
populations are located in Southern New Mexico (Otero) (Fig. 1a) and Northern Texas (Hudspeth) (Fig.
1b), approximately 10 kilometers apart. Lepidospartum burgessii grows only in gypsum soils, some on
mobile dunes, and others on flatter gypsum deposits. Its limited distribution is of particular concern
because it is producing little to no new seeds and no new recruits. This may be due to the species’ low
genetic diversity causing the plants to reject pollen from genetically identical individuals. Lepidospartum
burgessii is able to reproduce clonally (1), which proves problematic when the species tries to reproduce
sexually- the individuals are too similar to be compatible for reproduction. Other hypotheses consist of
insects eating the seeds before the plants are able to reproduce.
In order to find out more about inbreeding and population structure of L. burgessii, we used
microsatellites. Previous work analyzed the differences between L. burgessii and related species (Fig. 2,
ref. 2). We used one of the other two species in the genus (both more common than L. burgessii), L.
latisquamum, for the purpose of comparison. Previous work found some genetic differences between
species and limited differences between populations. We looked at the microsatellites of both species to
determine their genetic structures on a fine scale. To improve our conclusions, we added three more
microsatellite loci to the previous analysis.
Figure 2. Principal Coordinates Analysis ordination of previously collected data
using Nei genetic distances shows variation among both species and
individuals within L. burgessii populations.
Figure 4 (clockwise) a. Example of thriving
transplanted individual. b. Hand pollinating flowers to
test cross-pollination effects. c. Lace bug (Corythuca
marmorata), which infects L. burgessii and potentially
spreads fungi.
Figure 1 (from left) a. Lepidospartum burgessii in Otero
County, NM b. Lepidospartum burgessii in Hudspeth
County, TX
Figure 3. Scoring on Beckman Coulter shows dominance of homozygotes for
loci S20.