DOI: 10.5061/dryad.stqjq2c0h
title: Effect of sire population on the genetic diversity and fitness of F1 progeny in the endangered Chinese endemic Sinocalycanthus chinensis

description: Sinocalycanthus chinensis Cheng et S. Y. Chang (Calycanthaceae), which has
 a unique systematic status, is listed as a national second-class protected
 plants of China . In this study, the genetic diversity, performance, and
 fitness of F1 progeny from crosses between the Damingshan (DMS) population
 of Sinocalycanthus chinensis and pollen parents from the Daleishan (DLS)
 and Longxushan (LXS) populations were examined. The DLS population has a
 relatively small population size, low genetic diversity, and considerable
 geographical and genetic distances from the DMS population relative to the
 LXS population. Compared with naturally occurring seeds, DLS-sired seeds
 had the highest thousand-seed weight, starch content, fat content,
 germination rate, germination index, and emergence rate, but the lowest
 protein content. Naturally occurring, open-pollinated seeds had the lowest
 thousand-seed weight, starch content, and fat content, but the highest
 protein content. Compared with natural F1 progeny, DMS × DLS seedlings had
 the highest genetic diversity, photosynthetic parameters, and growth
 characteristics, except for leaf mass ratio and stem mass ratio. Under
 strong light, DMS × DLS seedlings exhibited a Fv/Fm value of 0.75, while
 the other two seedling types exhibited Fv/Fm values of 0.65. DLS-sired
 seeds had the most vigorous growth characteristics except for leaf mass
 ratio and stem mass ratio. These results suggest that genetic rescue by
 transplanting seedlings from the DLS population or hand pollination with
 pollen from the DLS population would be effective methods to reduce
 inbreeding depression and obtain strong offspring with high genetic
 diversity and fitness in the DMS population.
description: <em>Pollen
 sources</em> Plants from
 two <em>S. chinensis</em> populations located at two sites
 with different population sizes, levels of genetic diversity, and the
 relative geographical and genetic distances from the DMS seed parent were
 used as the pollen parents. One is located at Longxushan Mountain (LXS) in
 Anhui Province, China; this population is small with relatively high
 genetic diversity and with short geographical and low genetic distances
 from the DMS population (Table 1). This <em>S. chinensis
 </em>population is also within an evergreen broad-leaved forest. The
 main accompanying species are <em>Litsea coreana</em> var.
 <em>sinensis</em>, <em>Symplocos setchuensi
 </em>and <em>Platycarya strobilacea</em>. The other
 pollen parent population is located at Daleishan Mountain (DLS) in Tiantai
 County, Zhejiang Province, China, and it is a medium-sized population
 located a long geographical distance from the DMS population (Table 1).
 This <em>S. chinensis </em>population is located among shrubs
 within a valley. The main accompanying species are <em>Camellia
 cuspidate</em>, <em>Spiraea salicifolia</em>,
 <em>Corylopsis sinensis</em>, <em>Rhododendron
 simsii</em>, <em>Actinidia chinensis,</em> and
 <em>Sargentodoxa cuneata</em>. The DLS <em>S. chinensis
 </em>plants occur in the canopy and are intertwined with<em>
 A. chinensis</em> and <em>S. cuneata</em>.
 <em>Hand pollination
 and seed collection</em> Mature
 <em>S. chinensis </em>plants were selected as pollen donors.
 In May 2009, when the stigmas of <em>S. chinensis</em> flowers
 in DMS had matured, pollens were collected from flowers on 20 mature
 <em>S. chinensis </em>individuals in the LXS and DLS
 populations using Chinese brushes and stored in sterile plastic tubes. The
 pollen samples were maintained at 4°C and quickly transported to
 Damingshan Mountain, where the hand pollination was conducted with the DMS
 population. Pollen was transferred to DMS stigmas within 6 h. 30 mature
 individual plants in the DMS population were hand-pollinated with an
 average of 100 flowers per treatment. Each plant was randomly assigned to
 a crossing treatment and in total of 200 flowers were performed. All
 hand-pollinated stigmas were saturated with pollen. To exclude natural
 pollinators, plants were bagged with fine nylon mesh before flowering.
 Each hand-pollinated flower was also emasculated before the stigma became
 receptive to prevent selfing. Pollen was transferred directly from donor
 flower anthers onto receptive stigmas until the stigmas were saturated.
 The date was selected to ensure that each flower was encountered at the
 onset of its stigmatic surface receptivity. 30 naturally open pollinated
 DMS plants were selected as the control (Bossuyt, 2007; Holme, James
 & Hoffmann, 2008). <em>Fitness measurement
 </em> Fitness of the
 F1 progeny was defined as the relative reproductive success of a genotype
 as measured by survival, fecundity and other life history parameters
 (Molina-Montenegro et al., 2013) and was indicated by seed number per
 fruit, seed weight, seed size germination days, total germination rate,
 seedling emergence rate (Baker, Richards & Tremayne, 1994) and
 seedling biomass (Du, Yang, Guan & Li, 2016). In October
 2009, <em>S. chinensis </em>fruits were collected and
 air-dried, and seeds were collected from the dried fruits. Thousand-seed
 weights were measured using an electronic balance with an accuracy of
 0.0001 g. The starch, lipid, and protein contents of seeds were measured
 using anthrone–sulfuric acid colorimetric, Soxhlet extractor, and
 UV-spectrophotometric methods, respectively (Song, Cheng, Jiang, Long
 & Huang, 2008). In March,
 2010, DMS × DLS (pollen) seeds (hereafter, DLSH), DMS × LXS (pollen) seeds
 (hereafter, LXSH), and control seeds were germinated in an illuminated
 incubator (Jiangnan Instrument Inc., Ningbo, China) with 30/15°C and 12
 h/12 h light/dark cycles at 80% humidity. Seeds were immersed in
 H<sub>2</sub>SO<sub>4</sub> for 3 min, rinsed with
 sterilized water, and surface sterilized with 70% ethanol. Seeds were
 immersed in sterilized water and incubated at 28ºC for 2 days. Fifty seeds
 were placed into 3-mm deep silicon sand. Seeds were covered with moist
 filter paper to prevent them from drying out. Three replicates were used
 with a total of 150 seeds for each treatment. Seeds were
 considered to be germinated when their radicle length exceeded 2 mm
 (Hussain, Aljaloud, Alshammafy, Karimulla & Al-Aswad, 1997), and
 germination was recorded daily for 100 days. Germination-related indices
 were calculated as follows, according to previously described methods
 (Cai, 2008): (i) germination days were recorded as the number of days
 after sowing when the seeds begin germinating; (ii) total germination rate
 (%) = total number of germinated seeds on the 100<sup>th</sup>
 day / total number of seeds used for germination experiment × 100
 %. In March 2010,
 <em>S. chinensis</em> seeds of F1 DLSH, F1 LXSH, and control
 progeny were immersed in
 H<sub>2</sub>SO<sub>4</sub> for 3 min, rinsed with
 sterilized water, and surface sterilized with 70% ethanol. Seeds were
 immersed in sterilized water and incubated at 28ºC for 2 days. Seeds were
 planted 2-cm deep into soil-filled pots. Three seeds were planted per pot,
 and a total of 50 pots were used for each treatment. Sufficient tap water
 was added every day. One hundred days after planting, the seedling
 emergence rate was defined as the percentage of healthy seedlings that
 emerged, with a hypocotyl appearing on or above the soil surface (Demir
 & Mavi, 2004; Bolek, 2010). After the
 performance measurement described below, plants were harvested and divided
 into leaves, stems, and roots. Plant material was oven-dried at 105°C for
 1 h and then at 80°C until a constant weight was reached. The leaf, stem,
 and root biomasses were weighed with a balance to an accuracy of 0.1 mg,
 and the total biomass of seedlings was subsequently calculated.
 <em>Seedling
 photosynthetic physiological performance measurement</em>
 In May
 2010, healthy F1 DLSH, FI LXSH, and control seedlings were transplanted
 into pots, with each pot containing one seedling. In August 2010, we
 conducted in-situ photosynthetic trait measurements on a sunny day using a
 mature middle leaflet at the same position across plants, using a GFS-3000
 Portable Gas Exchange Fluorescence System (Heinz Walz GmbH, Effeltrich,
 Germany). The photosynthetically active radiation (PAR) was maintained at
 800 μmol m<sup>-2</sup> s<sup>-1</sup> using a
 red-blue LED light source, and the temperature was maintained at 25°C with
 a relative humidity of 70% inside the leaf measurement chamber. The
 CO<sub>2</sub> concentration within the chamber was maintained
 at 400 µmol mol<sup>-1</sup>. We recorded the net
 photosynthetic rate (<em>P</em>n), stomatal conductance
 (<em>G</em>s), transpiration rate (<em>T</em>r),
 and intercellular CO<sub>2</sub> concentration
 (<em>C</em>i) between 08:30 and 11:30. Three leaves per plant
 were chosen, and six consecutive measurements were performed (Li, Liao,
 Guan, Wang & Zhang, 2012). To construct
 light response curves, we obtained all photosynthesis measurements between
 09:30 and 11:00 (Beijing time) on a mature leaf from each plant with a
 leaf temperature of 25°C, a CO<sub>2</sub> concentration of
 400 ppm, and a relative humidity of 70%. We used a red-blue LED light
 source attached to the system to produce steady photosynthetically active
 radiation (PAR). Prior to the measurements, we allowed the mature leaf to
 acclimate under a PAR of 2000 μmol m<sup>-2</sup>
 s<sup>-1</sup> for 30 min to avoid photo-inhibition. As soon
 as the value stabilized, we exposed the leaves to a series of PAR values
 for 20 min or so in the following order: 2000, 1500, 1200, 1000, 800, 600,
 400, 200, 100, 50, 20 and 0 μmol m<sup>-2</sup>
 s<sup>-1</sup>. The temporal interval between each
 concentration was 3 min. We fitted the entire photosynthetic light
 response curve in Origin 8.0 (OriginLab, Northampton, MA, USA) as a binary
 linear equation by calculating the maximum of the net photosynthetic rate
 (<em>P</em>n) as <em>P</em>max. We utilized the
 following definitions. The light intensity at the maximum
 <em>P</em>n value (<em>P</em>max) was defined as
 the light saturation point (<em>LSP</em>). The light intensity
 at a zero <em>P</em>n value was defined as the light
 compensation point (<em>LCP</em>). The <em>P</em>n
 at a maximum PAR of zero was defined as the dark respiration point
 (<em>R</em>d). Chlorophyll
 fluorescence parameters under strong light (1200 μmol photons
 m<sup>-2</sup> s<sup>-1</sup>) were measured
 between 12:00 and 14:00 with a portable chlorophyll fluorometer (OS30P,
 Opti-Science Inc., Hudson, NH, USA). Measurements were performed on the
 third undamaged adult leaf from the top of each plant after 30 min of dark
 adaptation using light exclusion clips. For each measurement, three leaves
 per plant from three randomly selected plants per measurement were
 sampled. The data of three measurements of three leaves were averaged and
 used as the mean for each plant. The variable-to-maximum fluorescence
 ratio (<em>F</em>v/<em>F</em>m), which has been
 used to express the maximum PSII photochemical efficiency, was
 calculated. <em>Seedling morphological
 performance measurements</em> In September
 2010, plant height, leaf length, and leaf width were measured using a
 ruler to an accuracy of 0.1 cm, and the leaf length-to-width ratio was
 then calculated. The number of leaves was also recorded. The basal
 diameter was measured using Vernier callipers to an accuracy of 0.02 cm.
 Furthermore, whole pots were immersed in water, and then entire root
 complexes were excavated and carefully washed with running water to remove
 fine soil particles. Intact root systems were spread in a Perspex tray
 (A3-size) to minimize overlap, scanned (resolution 300 dpi, Epson 1680,
 Seiko Epson Corporation, Japan), and analysed using the WinRhizo software
 package (Version 3.10, Regent Instruments Inc., Quebec City, Quebec,
 Canada) to obtain the root volume (RV), total root length (RL), root
 surface area (RSA) and the number of root tips (RT).
description: Original data for the figures and light response curve were
 provided. 
description: This README file was generated on 2024-02-20 by Junmin Li. # Title of
 Dataset Effect of sire population on the genetic diversity and fitness of
 F1 progeny in the endangered Chinese endemic Sinocalycanthus chinensis ##
 Date of data collection Fruit data were collected in 2009. Seed data and
 light response data were collected in 2010. ### Links to publications that
 cite or use the data Li JM, Qi CH, Gu JJ, Jin ZX. Effect of sire
 population on the genetic diversity and fitness of F1 progeny in the
 endangered Chinese endemic Sinocalycanthus chinensis. Ecology and
 Evolution, 2020, 10: 4091-4103. #### File interview Data for figures is an
 Excel document with three sheets. Light response curves data is an Excel
 document. #### Description of the data 1. Data for Fig. 2 and Fig. 3(a)
 sheet in **Data for figures.xls** Number of variables: 6 Number of
 characteristics: 3 Variable List: Seeds number per fruit (#): The number
 of seed in every fruit Thousand seeds weight (g): the dry weight of 1000
 seeds Germination days: the number of days after sowing when the seeds
 begin germinating Total germination rate (%): total number of germinated
 seeds on the 100th day/total number of seeds used for the germination
 experiment*100%. Seedlings emergence rate (%): the percentage of healthy
 seedlings that emerged, with a hypocotyl appearing on or above the soil
 surface Plant biomass (g): The dry weight of whole plant Characteristics
 list: Nautral: open pollinated DMS plants LXSH: F1 progenies of
 Sinocalycanthus chinensis DMS population hybridized with pollen collected
 from LXS population DLSH: F1 progenies of Sinocalycanthus chinensis DMS
 population hybridized with pollen collected from DLS population Note:
 Different lower case letters indicate significant differences between
 different pollen sources at a p &lt; 0.05 level. 3. Data for Fig. 4, Fig.
 5 and Fig. 3(b) sheet in **Data for figures.xls** Number of variables: 6
 Number of characteristics: 3 Variable List: Plant height (cm): The height
 of plant Basal diameter (cm): The diameter of plant at the location with 1
 cm above soil Number of leaves (#): The number of all leaves Root volume
 (cm3): The volume of all roots Root surface area (cm2): The surface area
 of all roots Total root length (cm): The length of all roots Number of
 root tips (#): The number of all root tips Characteristics list: Nautral:
 open pollinated DMS plants LXSH: F1 progenies of Sinocalycanthus chinensis
 DMS population hybridized with pollen collected from LXS population DLSH:
 F1 progenies of Sinocalycanthus chinensis DMS population hybridized with
 pollen collected from DLS population Note: Different lower case letters
 indicate significant differences between different pollen sources at a p
 &lt; 0.05 level. 5. Data for Fig. 6, Fig. 7 and Fig. 3(c) sheet in **Data
 for figures.xls** Number of variables: 6 Number of characteristics: 3
 Variable List: Net photosynthetic rate (umol m-2 s-1): Dry matter
 accumulation per unit leaf area per unit time Intercellular concentrations
 of CO2 (umol mol-1): The concentration of nitrogen dioxide between
 biological cells Stomatal conductance (mol m-2 s-1): The degree of
 stomatal opening Transpiration rate (mmol m-2 s-1): The rate of
 decomposition of organic matter per unit area of leaves per unit time
 Pmax(umol CO2 m-2s-1): The maximum of the net photosynthetic rate Light
 saturation point (umol m-2 s-1):The light intensity at the maximum net
 photosynthetic rate value Light compensation point (umol m-2 s-1): The
 light intensity at a zero net photosynthetic rate value Apparent quantum
 efficiency (mmol m-2 s-1): The slope of the light response curve in the
 low light stage Dark respiration rate (mmol CO2 m-2 s-1): The net
 photosynthetic rate at a maximum PAR of zero Fv/Fm under strong light: The
 variable-to-maximum fluorescence ratio under strong light (1200 umol
 photons m-2 s-1) Characteristics list: Nautral: open pollinated DMS plants
 LXSH: F1 progenies of Sinocalycanthus chinensis DMS population hybridized
 with pollen collected from LXS population DLSH: F1 progenies of
 Sinocalycanthus chinensis DMS population hybridized with pollen collected
 from DLS population Note: Different lower case letters indicate
 significant differences between different pollen sources at a p &lt; 0.05
 level. 7. **light response curve.xls** Number of light intensities: 12
 Number of treatments: 3 Light intensities: 0-1800 umol m-2 s-1 Treatments
 list: Nautral: open pollinated DMS plants LXSH: F1 progenies of
 Sinocalycanthus chinensis DMS population hybridized with pollen collected
 from LXS population DLSH: F1 progenies of Sinocalycanthus chinensis DMS
 population hybridized with pollen collected from DLS population