New Insights into Colchicine-mediated Tetraploidy in Actinidia chinensis ‘Donghong’

Abstract

Red-fleshed kiwifruit exhibits favorable nutritional characteristics and is renowned for its sweet flavor. However, the majority of red-fleshed kiwifruit species are diploids, producing diminutive fruit sizes and limited resistance to abiotic stress. To cultivate new cultivars with superior traits, we employed colchicine treatment to induce tetraploidy from the diploid Actinidia chinensis cv. ‘Donghong’. In this study, a 20% induction rate was achieved by immersing the apical meristem in 0.2% colchicine solution for 48 hours. The induced tetraploids demonstrated larger stomatal size, lower stomatal density, higher chlorophyll content, and a lower chlorophyll a/b ratio compared to diploids; however, no significant difference in photosynthetic conversion efficiency was observed. Furthermore, the tetraploid red-fleshed kiwifruit grafted on the same tree exhibited superior fruit shape, quality, and weight compared to their diploid parent ‘Donghong’. The vitamin C content in tetraploid fruits increased by 62.6% compared to that in diploids. Also, the tetraploid ‘Donghong’ showed delayed phenology compared to the diploid from leaf buds to fruit ripening. This research not only generated a novel germplasm of tetraploid kiwifruit for future breeding, but also contributes to understanding the mechanism of polyploidy-induced trait variation in kiwifruit.

Introduction

Polyploidy refers to the presence of more than two sets of homologous chromosomes in an organism (Kuhl et al., 2022). Polyploid species feature many loci that control important traits, and polyploidization can provide substantial resources for agriculture and horticulture (Aono et al., 2022). Polyploidy can be formed naturally or artificially. An artificial polyploid is quickly acquired in the lab or field, whereas natural polyploids evolve, adapt, and speciate very slowly (Clo, 2022). Antimitotic agents like colchicine are used most frequently to accelerate polyploid breeding, as recently reported in garlic (Wen et al., 2022), Magnolia officinalis (Gao et al., 2022), Populus (Wu et al., 2022), Neolamarckia cadamba (Eng et al., 2021), cotton (Maru et al., 2021), orchids (Vilcherrez-Atoche et al., 2022), and so on. Although stronger resistance to stress and faster growth have been widely documented in artificial polyploids, new genetic, cytological, physiological, and molecular variations are still unclear in some new polyploids (Aversano et al., 2012; Chen et al., 2022; Gui et al., 2021; Morgan et al., 2022).

Artificial polyploid technology was first applied to obtain seedless fruits of great economic value, like watermelon (Cao et al., 2022), citrus (Lourkisti et al., 2020), tomato (Deslous et al., 2021), grape (Catalano et al., 2021) and so on. It could also could improve useful cultivar traits like tolerance to stresses, product quality and yield. Triploid breeding to induce 2n pollen in Populus has resulted in an improved growth-rate and timber quality (Zhou et al., 2020). The 2n gametes of orchid Cymbidium Swartz were used to produce triploids, which showed enhanced in vitro regeneration capacity, growth and flowering (Zeng et al., 2020). Artificial triploids and tetraploids of Lippia alba were obtained by colchicine treatment with an altered composition of essential oils (Julião et al., 2020).

Kiwifruit are high-nutrient fruits with health-promoting properties. For instance, there are various morphological and physiological changes among different ploidy races of kiwifruit, that is, diploids, tetraploids, and hexaploids (Huang et al., 2013). Kiwifruit genome duplication could contribute to the formation of the beneficial horticultural, agronomic and economic traits (Wu et al., 2019). Although there are several ploidy races in nature, such as octaploid and decaploid Actinidia arguta var. giraldii, tetraploid A. arguta var. arguta and A. melanandra, polyploid red-fleshed kiwifruit are very rare A. chinensis natural resources (Han et al., 2023; Zhang et al., 2017).

Kiwifruit breeding using traditional hybrids is significantly hindered by a high basic chromosome number, coupled with dioecism and a wide range of ploidy levels. By manipulating ploidy levels in vitro through chromosome doubling, polyploids can be generated to overcome the barriers associated with traditional cross-breeding. In this study, we selected the red-fleshed A. chinensis cv. ‘Donghong’ as the explant and employed colchicine treatment to produce polyploid plants, enabling characterization of their variations in terms of morphology, physiology, adaptability, and stress tolerance. This research will contribute to the development of new red-fleshed kiwifruit germplasms for future breeding endeavors.

Materials and Methods

Plant materials

The kiwifruit cultivar ‘Donghong’ was selected by Wuhan Botanica Garden, CAS, with a plant variety right number of National S-SV-AC-031-2012. The fruit is long, cylindrical, glabrous, while the fruit flesh is golden yellow and bright red around the core. Female aseptic seedlings of A. chinensis ‘Donghong’ were cultured in vitro and preserved in a tissue culture nursery. The induced female tetraploid vines were grafted to wild diploid rootstock and fertilized with male diploid A. chinensis ‘Donghong’ in the kiwifruit orchard of Wuhan Botanical Garden.

Induction of polyploid kiwifruit

The 1–2 mm apical meristems of sterile materials were taken and immersed in different concentrations of colchicine solution (Sinopharm Chemical Reagent Co., Ltd., Shanghai, China) before shaking in dark culture (80 rpm). The colchicine concentrations (w/v) were 0.2%, 0.3%, and 0.4%, respectively. The soaking times under different concentrations were 24 h, 30 h, and 48 h, respectively. At the same time, other apical meristems were soaked in sterile water without colchicine for 24 h, 30 h, and 48 h as a control. After the shaking culture, the apical meristems were washed 4–5 times with sterile water. Adventitious buds were then induced on a regeneration culture media. Each treated material was cultured in 50 bottles, with three apical meristems for each bottle. After regenerating adventitious buds, they were transferred to shoot strengthening and root induction media.

Identification of the chromosome ploidy of kiwifruit ‘Donghong’

About 0.5 cm × 0.5 cm of young leaves from ‘Donghong’ induction plants were placed in a clean plastic culture dish. About 300 μL of cell lysate with 2.46 mg·mL⁻¹ MgSO₄·7H₂O, 3.7 mg·mL⁻¹ KCl, 1.2 mg·mL⁻¹ hepes (pH = 8.0), 0.005 mg·mL⁻¹ dithiothreitol and 20 μL Triton X-100 stock (10%, w/v) (Sinopharm Chemical Reagent Co., Ltd.) was added, and the leaves were quickly chopped with a sharp blade. Then, 200 μL of DAPI stock (5.0 mg·mL⁻¹, propidium iodide) (Amresco, PA, USA) was added and mixed. The sample was filtered into a test tube through a 30 μm microporous filter membrane (CellTrics^TM, Partec, Münster, Germany) and loaded into a flow cytometer (PA-2, Partec). The distribution curve of sample DNA content was generated automatically based on the diploidy kiwifruit A. eriantha as a reference (It is preserved in the national kiwifruit germplasm resources garden of Wuhan Botanical Garden). The ploidies of samples were determined and analyzed by referring to Li (Li et al., 2010).

Observation of the cells, stomata and trichomes on the leaf surface of tetraploid and diploid kiwifruit

Two tetraploid plants D-88 and D-107 were grafted to unique A. deliciosa rootstocks for cultivation, and the grafted plants settled fruit in the third year. The young buds, leaves, flowers and fruits of tetraploid kiwifruits were recorded and compared with the diploid kiwifruit ‘Donghong’ (4-2 and 4-4). The photos were taken with a digital camera (Canon Inc., Tokyo, Japan).

The leaves of the tetraploid plants and the control plants were cut into 0.5 cm × 0.5 cm pieces, placed in a centrifugal tube containing 2.5% (v/v) glutaraldehyde (Sinopharm Chemical Reagent Co., Ltd.) for 4 hours, dehydrated according to the requirements of scanning electron microscopy, and dried with a critical point dryer (Leica EM CPD300; Leica, Germany). Then, the samples were loaded on a platform coated with gold powder. Cells and stomata distribution on the leaf surfaces were observed and photographed under a scanning electron microscope (Quanta 250 FEG; PHILIPS/FEI, OR, USA).

Measurement of the chlorophyll content and chlorophyll fluorescence of the tetraploid and diploid kiwifruit

The fourth fully expanded leaves from the vine tip were taken to measure the chlorophyll content according to the spectrophotometer method (Li and Wang, 2019). Subsamples were collected to detect the photosynthesis capability of the tetraploid and diploid ‘Donghong’ kiwifruit with a Monitoring-PAM Multi-Channel Chlorophyll Fluorometer (Walz, Effeltrich, Germany). The operation protocol and parameter designment referred to the chlorophyll fluorometer method (Jiang et al., 2017). The initial fluorescence yield (F_o), the maximum fluorescence yield (F_m) and the maximum quantum yield of PSII (F_v/F_m) were measured and calculated to detect the primary light energy conversion efficiency in leaves of the tetraploid and diploid ‘Donghong’ kiwifruit.

Fruit characterization of the tetraploid and diploid kiwifruit

The fruits of tetraploid and diploid ‘Donghong’ were collected and stored in a lab until they soft ripened at room temperature for about 30–40 days. The weight of single fruit, fresh fruit, and dry fruit of the tetraploid and the diploid kiwifruit ‘Donghong’ were measured with an electronic balance (Sartorius, Gottingen, Germany). Next, 2–3 mm thick transverse sections from the fruit’s equator were taken and dried at 60°C for 24 hours. The dry matter content was calculated from the ratio of dry weight to fresh weight of fruits. The ratios of each fruit’s longitudinal diameter to transverse diameter were measured by vernier calipers (0–15 mm; SHAHE, Zhejiang, China). The content of soluble solids was determined by a digital refractometer (PAL.1; ATAGO, Tokyo, Japan). The firmness of the fruits was measured with a 7.9 mm digital desktop diameter probe to assess fruit hardness (GY-4; HANDPI, Beijing, China). The L-ascorbic acid (AsA, vitamin C) content was measured according to the HPLC method (Liu et al., 2023). All fruits from all plants were collected and recorded.

Statistical analysis

The experimental data were expressed as mean ± SE (n = number of explants or plants or fruits) and analyzed using EXCEL and SPSS 19.0 software for the average value, standard error and Analysis of Variance (ANOVA). Duncan’s Multiple Range test and t-test were used to analyze pairwise comparisons with a significance level of 0.05.

Results

Colchicine inhibited kiwifruit regeneration at high concentrations and for a long duration

The regenerative capacity of the apical meristem of ‘Donghong’ significantly declined with increasing colchicine concentration, as shown in Table 1. Adventitious shoots from explants soaked in sterile water without colchicine exhibited a direct regeneration rate of 90.8%; however, after immersion in 0.4% colchicine, the direct regeneration rate decreased to 48.6%, and most treated material succumbed to direct browning. Furthermore, at the same concentration of colchicine, the regeneration rate of adventitious shoots from explants also markedly diminished with prolonged treatment duration. These findings indicate that increasing concentrations and durations of colchicine treatment are toxic and impede differentiation of adventitious buds.

Table 1

Effects of colchicine treatment on shoot apices of Actinidia chinensis ‘Donghong’ kiwifruit.

Flow cytometry analysis of the tetraploid kiwifruit ‘Donghong’

In order to characterize the ploidy of the colchicine-treated kiwifruit ‘Donghong’, young leaves were collected to measure their chromosome ploidy based on the diploidy of A. eriantha. The peak value of the diploid A. eriantha showed the presence of 2C DNA content, which was designed as a standard reference sample (Fig. 1A). The peak values of untreated and colchicine-treated kiwifruit ‘Donghong’ material were recorded in Figures 1B, 1C and 1D. The 4C DNA contents of kiwifruit ‘Donghong’ were shown in Figure 1C with twice the peak values compared to the standard reference sample and colchicine-untreated sample, so it is regarded as tetraploid kiwifruit ‘Donghong’. However, DNA contents of some samples were somewhere between diploid and tetraploid kiwifruit as shown in Figure 1D, and these were regarded as chimera individuals.

Fig. 1

Histograms of flow cytometry for kiwifruit. A: diploidy Actinidia eriantha (control); B: diploid Actinidia chinensis ‘Donghong’ (control); C: tetraploid Actinidia chinensis ‘Donghong’; D: chimera Actinidia chinensis ‘Donghong’.

Optimal conditions for inducing polyploidy in kiwifruit with colchicine

The results of the ploidy test conducted through flow cytometry indicated that the mutagenesis rate of shoot apices differentiation and polyploid induction in ‘Donghong’ is influenced by different concentrations of colchicine and treatment durations. Specifically, exposure to gradually increasing concentrations of 0.2%, 0.3%, and 0.4% colchicine yielded various mutagenesis rates for ‘Donghong’, ranging from 4.3% to 20.0%, 5.1% to 6.4%, and 9.0% to 10.3%, respectively (Table 1). Likewise, under identical treatment durations, higher concentrations of colchicine resulted in an elevated mutagenesis rate for obtaining polyploids through shoot apices differentiation in ‘Donghong’.

Notably, under the high concentration of 0.4% colchicine, the likelihood of explant differentiation to obtain chimeras significantly diminished, and a chimera rate of 0 was observed. However, it is important to highlight that the high concentration of colchicine often led to increased mortality rates. For instance, a mortality rate as high as 20.8% was noted under the 0.2% colchicine treatment, reaching 48.9% under the highest concentration tested (Table 1). Consequently, when considering equal treatment durations, overall induction rates were lower for higher concentrations compared to lower concentrations of colchicine. In light of these findings, we recommend adopting a treatment duration of 48 hours with a colchicine concentration of 0.2% as an optimized approach to achieve both shoot apices differentiation and polyploid induction.

The leaf characteristics of diploid and tetraploid ‘Donghong’

The high grafting technique was employed to compare the young buds, leaves, flowers, young fruits, and mature fruits of tetraploid kiwifruits with those of the diploid ‘Donghong’, as depicted in Figure 2. Based on the phenology of diploid and tetraploid ‘Donghong’ from 2020 to 2022, we found that the diploid developed earlier at all growth stages compared with the tetraploid ‘Donghong’ over several days.

Fig. 2

Comparison of young leaf buds, young leaves, flowers, young fruits, and mature fruits in diploid and tetraploid kiwifruit ‘Donghong’ during developmental stages. A, F: young leaf buds; B, G: young leaves; C, H: flowers; D, I: young fruits; E, J: mature fruits. A–E: diploid ‘Donghong’ on the left, and F–J: tetraploid ‘Donghong’ on the right. Scale bar = 1 cm. The exact date of the photograph is indicated next to it.

Scanning electron microscopy was utilized to observe the leaf tissue structure of both tetraploid and diploid kiwifruits. As illustrated in Figure 3, the leaf surface is adorned with bifurcated unicellular trichomes lacking glandular properties. In contrast to diploid lines, tetraploid mutants possess sparser 3–5 bifurcated trichomes, while diploid lines exhibit denser 5–7 bifurcated trichomes.

Fig. 3

The leaf surface morphology of diploid and tetraploid kiwifruit ‘Donghong’ as observed by scanning electron microscopy. A, B: tetraploid kiwifruit ‘Donghong’; C, D: diploid kiwifruit ‘Donghong’.

The stomata are predominantly distributed on the abaxial surface of kiwifruit leaves, exhibiting an almost round or near-round shape in tetraploid varieties compared to the oval shape observed in diploid kiwifruit. Measurements of stomatal apparatus length and width revealed that tetraploid stomatal cells possessed a long axis measuring 35.06 ± 1.34 μm, a length-to-width ratio of 1.01 ± 0.04, and a stomatal density of 183.3 ± 43.03 per mm². In diploid lines, stomatal cells exhibited a long axis measuring 23.86 ± 1.74 μm, a length-to-width ratio of 1.52 ± 0.08, and a density of 291.7 ± 31.91 per mm² (Fig. 4). Significant differences were observed between the two ploidy levels in kiwifruits (P < 0.05).

Fig. 4

Comparison of stomatal size, density, aspect ratio between tetraploid and diploid plants. A: Length of stomatal; B: Width of stomatal; C: Ratio of length/width; D: stomatal density. SD bars indicate standard error. The asterisk stands for significant difference (P < 0.05). n = 3 biological replicates.

The photosynthesis of diploid and tetraploid ‘Donghong’

As shown in Figure 5, tetraploid kiwifruits have higher chlorophyll a, chlorophyll b, chlorophyll a+b and carotenoid levels than those of diploid controls. The total chlorophyll concentration of tetraploids increased by 24.2% to 44.2% compared to the control. However, the chlorophyll a, chlorophyll b, and chlorophyll a+b contents of the tetraploid kiwifruit were almost the same as that of the control diploid cultivated under a sun-shade condition with a 75% shading rate. Consequently, the induced tetraploid kiwifruit has a clear advantage in open fields, but not in shady cultivation conditions, due to its superior photosynthesis.

Fig. 5

Chlorophyll contents in tetraploid and diploid kiwifruit leaves in an open field or under 75% shading cultivation conditions. A: Content of chlorophyll a; B: Content of chlorophyll b; C: chlorophyll a/b; D: Content of carotenoid. SD bars indicate standard error. The asterisk stands for significant difference (P < 0.05). n = 3 biological replicates.

The F_o, F_m, F_v and F_v/F_m values of photosynthesis efficiency in kiwifruit are shown in Figure 6. Tetraploid and diploid kiwifruit were grown under different conservation environments; an open field or under 75% shading. Photosynthetic efficiency produced a bimodal curve with a peak at 8 am and a peak at 6 pm and a plateau at noon. There was almost no difference between tetraploid and diploid kiwifruit under any cultivated conditions.

Fig. 6

Comparison of the maximum quantum yield (F_v/F_m) in diploid and tetraploid kiwifruit ‘Donghong’. Data with different letters are significantly different (P < 0.05). SD bars indicate standard error. The asterisk stands for significant difference (P < 0.05). n = 3 biological replicates.

The fruit characteristics of diploid and tetraploid ‘Donghong’

As shown in Figure 7, tetraploid fruits were significantly rounder than diploid fruits. The single fruit weight of tetraploid D88 was significantly heavier by 36.3% than that of diploid 4-4 (Fig. 8A). The ratio of the longitudinal to transverse diameter in the tetraploid was nearer to 1 than that of the diploid as shown in Figures 8B, C and D. The soluble solids content in tetraploid fruits was 7.02°Bx, and was significantly higher than that of 5.09°Bx in diploid fruits (Fig. 8E). The firmness of tetraploid fruits was 5.1 kg·cm⁻² on average, and was comparatively softer than diploid fruits that had 7.12 kg·cm⁻² firmness (Fig. 8F). The dry weight of tetraploid D107 fruits was lighter that in other fruits, but the other one of tetraploid D88 specimens had the opposite characteristics; there were not enough statistical samples to compare differences in dry matter between diploid and tetraploid ‘Donghong’ (Fig. 8G). Surprisingly, the L-ascorbic acid (AsA, vitamin C) content in tetraploid ‘Donghong’ significantly increase by 62.6% compared with that in diploid fruits, reaching 163.41 mg/100 g FW on average (Fig. 8H).

Fig. 7

The fruit morphology of tetraploid and diploid kiwifruit ‘Donghong’. A: longitudinal and cross sections of diploid kiwifruit; B: longitudinal and cross sections of tetraploid kiwifruit. Scale bar = 1 cm.

Fig. 8

Comparison of the fruit characteristics between tetraploid and diploid kiwifruit ‘Donghong’. A: Single fruit weight; B: Longitudinal diameter of the fruit; C: Transverse diameter of the fruit; D: Ratio of longitudinal to transverse diameter; E: Soluble solids content of the fruit; F: Fruit firmness; G: Dry matter of the fruit; H: Content of vitamin C in the fruit. SD bars indicate standard error. The asterisk stands for significant difference (P < 0.05). n ≥ 3 biological replicates.

Discussion

The antimitotic agent colchicine is often used to induce polyploidy artificially. Seeds are immersed in, or cultured on, media supplemented with colchicine at various concentrations. Nevertheless, colchicine inhibits seed germination and seedling growth and thus it is time-consuming and inefficient for plants with low seed germination rates (Lv et al., 2021; Maru et al., 2021). Therefore, more research has been conducted on tissue immersion methods, such as seedling immersion, root immersion, and explant immersion (Chaikam et al., 2020; Wen et al., 2022). High-frequency direct-shoot organogenesis is fundamental to the tissue immersion method. It could significantly shorten the development time compared to seed immersion. In this study, the regeneration rate from shoot apices reached more than 90% in kiwifruit ‘Donghong’ without colchicine treatment (Table 1). With increases in colchicine concentration and treatment period, the regeneration rate gradually decreased (Table 1), showing that the colchicine retarded the regeneration of shoot apices in kiwifruit. Polyploidy is induced more rapidly with increasing colchicine concentrations and treatment duration (Table 1). At the 0.2% colchicine concentration and immersion for 48 h, the regeneration rate was 69.8% and the induction rate was 20%, which will be used as the optimum colchicine treatment in further research.

Polyploids exhibit useful morphological, physiological, and ecological traits (Sattler et al., 2016), which provide superior germplasms for agricultural domestication and breeding. In fact, polyploidy drives evolution in the plant kingdom (Van de Peer et al., 2017, 2021). Petioles from five diploid A. chinensis Planch. genotypes were selected as explants to obtain a tetraploid polyploid using colchicine treatment (Wu et al., 2011), and the main improvement was the increased fruit size, without other fruit quality assessments (Wu et al., 2012). Here, tetraploid kiwifruit performed better than diploid kiwifruit in terms of stomatal size, density, chlorophyll contents, and fruit weight and quality (Figs. 4, 5, and 8). From the SEM results, we found that the stomatal cells were bigger, rounder and fewer in the tetraploid kiwifruit compared to those in diploid fruit (Fig. 4), which is consistent with the fact that 2C-values and stomatal lengths were positively correlated in Vanilla planifolia (Bory et al., 2008), Rhododendron fortune (Mo et al., 2020) and Cannabis sativa L. (Parsons et al., 2019). The larger stomata size and reduced stomatal density possibly improved stomatal conductance, increasing the stress resistance and water-use efficiency as in higher plants (Guo et al., 2019; Monda et al., 2016). Trichomes are the first line of plant defense (Han et al., 2022). The branched unicellular non-glandular trichomes in kiwifruit play an important role in resisting various abiotic and biotic stresses. The trichomes on the leaf epidermal cells were less branched and sparse in the tetraploid than that in the diploid kiwifruit. The interlink between the branch numbers and the stress resistance should be intensively observed and discussed in the future. Going forward, kiwifruit polyploids may promote fruit yield and quality.

Higher chlorophyll content generally leads to darker green leaves, increased photosynthetic capacity and higher yields in tetraploids compared to diploids (Mo et al., 2020; Šmarda et al., 2018). Kiwifruit has a high ratio of chlorophyll a/b, so it can absorb long-wavelength light. However, the tetraploid kiwifruit had a lower percentage of chlorophyll a/b in the high grafting condition, possibly because of a lower rootstock effect. The tetraploid kiwifruit had a slightly higher ratio of chlorophyll a/b in the open field but was not significant, combined with undistinguished chlorophyll fluorescence and a maximum quantum efficiency of PSII photochemistry between tetraploid and diploid kiwifruit plants (Baker, 2008), suggesting that tetraploid kiwifruit may have no advantages over diploid kiwifruit in terms of photosynthetic efficiency. The most important factor influencing the photosynthetic efficiency of tetraploid kiwifruit in this study was the growth conditions in the open field.

The tetraploid ‘Donghong’ showed delayed phenology compared to the diploid from leaf buds to the fruit ripening based on observation for consecutive three years, excluding the fruit development stage on the tree. This stage lasted several months and the quality of unripe fruits was not compared. Previous studies have reported similar observations in Cardamine flexuosa, where the flowering time of the allopolyploid wild plant C. flexuosa was delayed compared with its diploid parents C. hirsute (Akiyama et al., 2020). However, not all polyploids exhibit the same characteristics. In Arabidopsis, for example, the tetraploid Col-0 had a delayed flowering time, whereas the tetraploid Ler did not (Cheng et al., 2022). Therefore, the influence of ploidy on phenology depends on the species and genetic background.

Kiwifruit is called the king of vitamin C fruits. The vitamin C content is usually regarded as an important indicator for evaluating the quality of kiwifruit. The tetraploid kiwifruit ‘Donghong’ has more vitamin C than diploid kiwifruit, and this can greatly improve the nutritional value of kiwifruit ‘Donghong’ (Fig. 8H). Previous research has indicated that the duplication of regulators played a crucial role in vitamin C biosynthesis and accumulation, particularly in A. latifolia, which has the highest vitamin C content among 54 kiwifruit species (Han et al., 2023). Therefore, the tetraploid kiwifruit ‘Donghong’ contains some duplicated genomic information, which may contribute to its higher vitamin C content compared to diploid ‘Donghong’. Vitamin C can also reduce oxidative damage and strengthen the stress tolerance of kiwifruit (Liu et al., 2023). The yellow-fleshed tetraploid cultivars of A. chinensis have been selected for their resilience against Pseudomonas syringae pv. actinidiae (Psa) biovar 3 disease (Tahir et al., 2020). In the future, it will be useful to intensively study the fitness of red-fleshed tetraploid kiwifruit ‘Donghong’ with higher vitamin C content and resistance to different kiwifruit diseases.

Conclusion

In order to develop more kiwifruit elite varieties, we produced a tetraploid kiwifruit ‘Donghong’ by artificial colchicine treatment. In general, the tetraploid kiwifruit showed a better performance in terms of plant growth and fruiting. This will lay the foundation for conserving and developing A. chinensis resources and provide a clear direction for future attempts to extensively breed new kiwifruit cultivars.

Acknowledgements

We thank Wenjun Huang (Associate professor, Wuhan Botanical Garden) for critical reading of this manuscript.

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