DOI: 10.5061/dryad.zgmsbccf5
title: Polyploidization-enhanced effective clonal reproduction endows the successful invasion of Solidago canadensis

description: Clonality and ploidy levels are positively associated with plant
 invasiveness. However, there is still no consensus on whether
 polyploidization can promote the invasion of alien plants by enhancing
 clonality. Our recent long-term community succession study found that the
 more vigorous clone of introduced polyploid Solidago canadensis succeeded
 into mono-dominant community, which seems to be a positive correlationship
 between polyploidization and clonal reproduction. However, how
 polyploidization improves the clonal reproduction of S. canadensis remains
 unknown. Here, we compared clonal growth ability among diploids and
 polyploids of S. canadensis from native and introduced ranges in a common
 garden. Results showed that the rhizomes of S. canadensis originated from
 axillary buds of dense nodes at the basal stem of seedling and then
 produced into clonal ramets. Diploids had denser nodes and more buds,
 developed more rhizomes per unit mass and produced more clonal propagules
 at the early growth stage compared with polyploids. However, the number of
 juvenile and secondary rhizomes, as well as the diameter and length of
 rhizomes in polyploid populations was significant higher than those of
 diploids, and those clonal traits in introduced polyploids were
 significant higher than in native polyploids. Moreover, a phalanx growth
 form was observed in native and introduced diploid populations, which
 allocated about 3% and 5% of the total biomass to rhizomes, respectively,
 resulting in short and weak rhizomes. However, native and introduced
 polyploids allocated about 35% and 40%, respectively, of the total biomass
 to rhizomes, resulting in long and strong rhizomes, which were guerrilla
 growth forms. This study firstly shows that polyploidization enhanced the
 effective clonal reproduction of S. canadensis through pre-adaptation and
 rapid post-adaptation evolution, and consequently contributed to its
 successful invasion.
description: Diploid
 (2n = 2x = 18), tetraploid (2n = 4x = 36), and hexaploid (2n = 6x = 54)
 seeds of S. canadensis were collected from the native region (US and
 Canada) and invasive region (Europe and China). The detailed descriptions
 of material collection and identification had been presented by Cheng et
 al. (2021). These seeds were stored in refrigerator at
 4℃, before we used these seeds, they had been
 stored in the refrigerator for about five ~ eight years in the Weed
 Research Laboratory, Nanjing Agricultural University, China. The ‘NA2x’,
 ‘NA4x’, ‘NA6x’, ‘IN2x’, ‘IN4x’ and ‘IN6x’ were used to represent the
 native diploid, native tetraploid, native hexaploid, introduced diploid,
 introduced tetraploid, and introduced hexaploid, respectively. Each
 geo-cytotype was comprised of 4 populations with 20 individual plants (see
 detailed information of different populations in supplementary material
 Appendix S1: Table S1). <strong>Morphogenesis of rhizome of <em>S.
 canadensis</em></strong> In order to
 determine the origin of rhizome of <em>S. canadensis</em>,
 seeds of<em> S. canadensis </em>of different geo-cytotypes
 (NA2x, NA4x, NA6x, IN2x, IN4x, and IN6x) were sown in pots (11-cm-wide,
 8-cm-deep, 1 L) with a 1:1 mixture of sterilized compost and vermiculite.
 From cotyledon stage to the stage of rhizome formation, 20 seedlings (4
 populations × 5 individuals) from each geo-cytotypes population were
 chosen at each stage that two new leaves outgrew, respectively. Plants
 were pulled out of the pots and washed the soil carefully to ensure that
 the roots and rhizomes are intact and undamaged. The originations of
 rhizomes were observed with a stereomicroscope (SZX7, OLYMPUS, Japan). The
 selected typical seedlings were recorded with a camera.
 <strong>Difference of basal nodes and the number of
 axillary buds of different geo-cytotypes of <em>S.
 canadensis</em></strong> After
 determining that the rhizome of<em> S. canadensis</em>
 originated from axillary buds, we counted the number of axillary buds at
 the basal stem of <em>S. canadensis </em>to study the
 potential of rhizome production in different geo-cytotype populations. The
 basal stem here refers to the stem starting between nodes 1 and 10 (node 1
 = cotyledonary node). <strong>Comparison of rhizomes and clonal ramets of
 different geo-cytotypes of<em> S. canadensis</em> in pot
 common garden experiment and field
 test</strong> Seeds of six
 geo-cytotypes (4 populations with 20 individuals were selected from each
 geo-cytotype) were sowed in small germination pots respectively, which
 were filled with the same soil substrate as described above and grown for
 one month. Then, uniform plants in each population were selected and
 transplanted into 1 L pots (10-cm-wide, 9-cm-deep) with the same soil
 substrate. Each of the pots contained one individual and was placed in a
 greenhouse. Pots were randomly arranged on tables and changed their
 positions weekly. They were watered every two ~ four days and manually
 removed the other weeds regularly. Three months later, they were
 transplanted into 6.6 L pots (24-cm-wide, 21-cm-deep) with a 1:2 mixture
 of sterilized compost and soil and each of the plots contained 1 seedling.
 The plots were placed in the outdoor of Pailou Teaching and Research
 Station of Nanjing Agricultural University (32°02′N, 118°37′E), Nanjing,
 China. This region is characterized as being a subtropical monsoon
 climate, with an average annual rainfall of 1090mm and the mean annual
 temperature is 15°C (lowest in Feb at 2.7°C and highest in July at
 28.1°C). They were watered, weeded, and sprayed with pesticides regularly.
 After these plans grew for six months in the outdoor (October 2018), the
 number of clonal ramets and rhizomes developed from axillary buds at the
 basal stem of each genet was counted by removing the topsoil around the
 basal stem of genet to expose stem above the cotyledon node without
 damaging the roots, and the height (cm) and diameter (mm) of each ortet
 were measured with a straight-edge and a vernier caliper, respectively.
 After these measurements, the ramets were cut off in winter except the
 basal stem (5~6 cm) and belowground parts and some soils were added to
 these pots. In the next two years, we only watered, weeded, sprayed with
 pesticides regularly and cut off the aboveground part in winter. Three
 years later, in September 2021, the number, diameter (mm), height/length
 (cm), and biomass (g) of ramets and rhizomes of different geo-cytotypes
 were measured by digging out these plants from pots. Because the rhizomes
 are produced not only by the axillary buds on the basal stem of mother
 ramet and daughter ramet but also by the axillary buds on the rhizome, we
 defined rhizome that produced from mother ramet as ‘old rhizomes’, rhizome
 that produced from daughter ramet as ‘juvenile ramet’ and rhizome that
 produced from old rhizome as ‘secondary rhizome’ in this
 study. To determine what is the difference of clonal
 reproduction among the six geo-cytotypes in a field environment, we used
 the field test of Cheng et al. (2020) to analyze the clonality of
 different cytotypes of <em>S. canadensis</em>. This study was
 also conducted at the Pailou Teaching and Research Station of Nanjing
 Agricultural University from July 2011 to November 2015. The field
 experiment was established without artificial watering and fertilization
 during the whole period of experiment. Plots were randomly designed with
 four replicates for the six geo-cytotypes. <em>S.
 canadensis</em> were planted in a plot (3 × 2 m) including 12 plants
 and named the plot with the corresponding geo-cytotype (see supplementary
 material Appendix S1: Figure S1). Local weeds were also transplanted in
 every plot in order to create a scene where<em> S.
 canadensis</em> compete with local weeds (detailed information had
 been shown in paper (Cheng et al. 2020)). In November 2011 to November
 2015, before the <em>S. canadensis</em> seeds matured, the
 aboveground of <em>S. canadensis</em> and four local weeds
 were cut off every year
 during the winter, maintaining the growth of belowground root and rhizome.
 The spacer length between mother ramet and daughter ramet (cm) of each
 geo-cytotype with 48 mother ramets (4 plots × 12 individuals), in the
 first year were measured. From 2012 to 2015, the total number of ramets in
 each plot was counted in the spring and the autumn respectively. At the
 same time, the average clonal ramets produced by each ortet, the total
 number of ramets in each plot, plant height (cm) and fresh biomass (g) of
 aboveground of different geo-cytotypes in the autumn of the first year
 were measured.