Stability of watermelon phenotype characters (Citrullus lanatus (Thunb.) Matsum. & Nakai) from crossing ♀ ‘Putri Delima’ with ♂ ‘Maduri’

Watermelon (Citrullus lanatus (Thunb.) Matsum. & Nakai) is a horticultural plant that belongs to the Cucurbitaceae family with high public demand, however, local markets sometimes have limited supply. The existence of watermelon varieties that are not pest-resistant causes its production to be erratic. Therefore, plant breeding efforts are required to produce superior varieties through the stability test of plant characters. For watermelon to be certified as a new variety, it needs to possess a stable and adaptive character to various conditions. The F1 watermelon from crossing ♀ ‘Putri Delima’ with ♂ ‘Maduri’ produces inole-shaped fruit, red flesh, and a sweet taste. Therefore, this study aims to determine the stability of the phenotypic character of watermelon F2 from crossing ♀ ‘Maduri’ with cultivar ♂ ‘Putri Delima’ and was conducted in Jamusan, Bokoharjo, Prambanan, Sleman, D.I.Yogyakarta fields from August to December. There are five samples of ripe watermelons that were selected randomly while their phenotypic characters were observed qualitatively and quantitatively. Each F2 watermelon character was compared to F1, and the quantitative analysis was conducted using one-factor ANOVA with a confidence level of 5%. The results of quantitative character analysis between F2 and F1 showed a P (P-value)> 0.05. Meanwhile, the results of qualitative observations of F2 watermelon showed different flesh and skin color, while the harvest time from F1 was caused by the segregation of heterozygous crosses. Therefore, it is necessary to select superior phenotypic characters as desired for the next breeding.


INTRODUCTION
Indonesia is an agricultural country with rapid development in growing horticultural plants, such as watermelon (Citrullus lanatus (Thunb.) Matsum. & Nakai) which belongs to the Cucurbitaceae family with high public demand and is classified as a seasonal creeping plant with a tool holder like gyre (Hannah & Krishnakumari, 2015;Pan et al., 2020). The surface of this plant has sharp fine hairs that cover the stems and leaves (Wei et al., 2020). Guo et al. (2013), stated that watermelon is a type of fruit that is very popular among people. Furthermore, the plant has high water content and also rich in minerals such as Ca, Mg, P, niacin, riboflavin, thiamin, and carotenoid (Grassi et al., 2013;Ilahy et al., 2019) and as stated by Syukur et al. (2012), an average watermelon has a solid content of 8-10% and 20-25% sucrose.
Due to its fresh, sweet taste, and popularity with the public, the demand for watermelons in Java is high which leads to an increase in market demand every year. In 2019, Kementerian Pertanian stated that watermelon had an average production yield of 13.86 tons/ha, while in 2014, it was 18.27 tons/ha. This increase in market demand becomes an obstacle for farmers due to the absence of local seeds that are stable and superior enough to balance needs. Meanwhile, one of the efforts of producing superior and stable seeds is the use of plant breeding techniques through quality development on hybrid varieties of watermelon seeds (Ceccarelli, 2015;Cardi, 2016;Dou et al., 2018). With the development of hybrid watermelon seeds' quality, it is expected that new certified cultivars with high stability and adaptability created for large-scale cultivation.
Some of the superior watermelons that have been tested for stability are cultivars 'Putri Delima' and Maduri. Meanwhile, the 'Putri Delima' watermelon cultivar has an oblate or oval (inole) fruit shape, dark green skin color, and red flesh with an average fruit weight of 3.5-6 kg, and Brix or a sweetness level of 13. This cultivar is harvested in the range 60-75 day after seeding/sowing (DAS). The Maduri watermelon is a superior cultivar with the code WM 2479 (Kementerian Pertanian, 2016). Furthermore, Maduri has an oval fruit shape (inole), dark green skin color with blackish stripes, and yellow flesh. Meanwhile, this cultivar is harvested in the range 56-60 DAS, has a long shelf life, and tolerant of fusarium wilt with a leaf crackle.
This study aims to test the stability of the phenotypic character of F 2 watermelon from crossing watermelon cultivar ♀ 'Putri Delima' with ♂ 'Maduri' and comparing with the F1 results of a previous study by Putri & Daryono (2017) which is stable and superior.

MATERIALS AND METHODS
This study was conducted in Jamusan (Greenhouse), Bokoharjo, Prambanan, Sleman, D.I.Yogyakarta, and the Laboratory of Genetics and Breeding, Faculty of Biology, Universitas Gadjah Mada.
Cultivation. Watermelon cultivation consists of three stages, namely, land preparation, seed germination, and maintenance. Land preparation was carried out by cultivating through taxation and the establishment of good irrigation channels. Meanwhile, the seed germination stage began with soaking for one day, followed by sorting and planting in warm conditions with a temperature of ± 28°C which is equipped with a 10-watt lamp. The 10 days old germinated seeds were transferred to the field for permanent planting while the third stage was carried out by routine fertilizers and irrigation. Moreover, watermelons that have male and female flowers were pollinated manually and lateral branches were trimmed by removing seven to eight leaves and separating them into two leaves. This process was carried out on subsequent branches until the 20 th or 25 th internode is reached. After obtaining the ovaries from the crosses, the watermelons were nurtured until ripe and ready to be harvested. Meanwhile, the watermelon that is ready for harvest has a dark yellow spot and a loud sound when tapped.
Phenotype characterization. Generally, the reference for sampling and testing the difference, uniformity, and stability of watermelons is according to UPOV (2012). In this study, five samples were tested for stability and the phenotypic character which include qualitative and quantitative data. The qualitative data were measured by observing the fruit and seed shape, skin and flesh color, together with pulp texture according to Kementerian Pertanian (2016). Moreover, the fruit color observations refer to the RHS mini chart 2015. Furthermore, the quantitative characters observed include fruit size, thickness, skin and flesh weight, sweetness level, storage capacity, number of seeds, and weight of 100 seeds.
Data Analysis. The quantitative data obtained were tested using one-way ANOVA with a confidence level of 5% through a comparison of the results between F2 and F1 watermelon.

RESULTS AND DISCUSSION
Watermelon has a distinctive phenotypic character which is the external appearance or other characteristics of every organism that are observed. This character is determined by the interaction results between proteins in the cell. Meanwhile, observations of phenotypic characters are classified into two types based on the measurement and variety continuity, namely the qualitative and quantitative phenotype character (Daryono et al., 2019). In this study, F2 was from hybrid crosses (heterozygous) with unstable phenotypic characters and there was segregation. Therefore, there is a need to select the desired phenotypic characters.
Qualitative characters. These are visible characters that are not measured in certain measurement units. Also, qualitative characters have discontinuous diversity and are controlled by each single gene (Govindaraj et al., 2015;Cao et al., 2016;Peterson & Müller, 2016). From the results, as shown in Table 1, the fruit of the F2 watermelon from crossing ♀ 'Putri Delima' with ♂ 'Maduri' has the same shape as the F1 breeders, namely oblate/innole, while the fruit skin color and the flesh color are different. The F1 fruit skins, namely (RHS 2015137A, RHS 2015N 189A, RHS 2015 showed more diverse colors than in F2 (RHS 2015 139A; RHS 2015 145A). Moreover, fruit skin is one of the important qualitative phenotypic characters in watermelon breeding because it determines consumer interest. The green skin color of the watermelon is the wild type and the changes in skin color that occur are caused by evolution, artificial selections, and gene mutations (Yang et al., 2015;Li et al., 2019;Sun et al., 2020). These gene mutations occur on chromosome four with a single gene dominant pattern of inheritance (Guo et al., 2013;Dou et al., 2018). The watermelon pulp produced by F1 shows a different appearance compared to F2 while the color of F2 has similar properties with the parental 'Maduri', which is yellow to dark (RHS 2015 5A; RHS 2015 21 A). This is due to the existence of two loci (C locus and i-C locus) which play a role in inhibiting and controlling the color of watermelon flesh (Bang et al., 2010;Zhao et al., 2013;Liu et al., 2016). In addition, several exogenous factors that affect the phenotypic characters and fruit quality include light and temperature conditions, pathogen attacks, as well as pre-and postharvest manipulations (Ilahy et al., 2019).
Moreover, the color of the seeds produced by F1 watermelon is blackish (RHS 2015 200A) while the color of the F2 is black with yellow spots (RHS 201510C, RHS 2015. The color of the seeds in F2 is similar to the yellow watermelon cultivar 'Maduri' while F1 is similar to the red watermelon broodstock of the 'Putri Delima' cultivar. Meanwhile, the difference in seed color is due to the instability of phenotypic characters or certain genes that regulate the watermelons pigment. The F1 watermelon from crossing ♀ 'Putri Delima' with ♂ 'Maduri' has a shorter harvest age than F2, due to the difference in planting time and season. When the rainy season is high, rainfall is higher and the phosphorus content of the soil decreases as it is eroded by the rainfall. This inhibits plant growth, fruit, and seed maturity (White & Veneklaas, 2012;Bai et al., 2013;Emongor et al., 2017). Meanwhile, F2 watermelon which is the result of crossing ♀ 'Putri Delima' with ♂ 'Maduri', was planted during the rainy season to have a low intensity of irradiation was low that reduces photosynthate accumulation. Unlike F1 which is planted in the dry season, F2 shows a shorter harvest life due to more radiation. Meanwhile, more radiation causes the fruit to ripen quickly due to the higher accumulation of photosynthate. In addition to adequate radiation, watermelons grow optimally when planted with a temperature range of 22°C-30°C, humidity less than 80%, an average rainfall of 40 to 50 mm/month, and influenced by the soil structure as well as microbial composition , Wang et al., 2019.
The differences in the appearance of the flesh and seed color between the F1 watermelon from crossing ♀ 'Putri Delima' with ♂ 'Maduri' with F2 watermelon shown in Fig. 1. Quantitative characters. Phenotypic characters are quantitative that is measured clearly and have continuous diversity which forms a spectrum, and when the population is large enough it forms a normal distribution curve (Daryono et al., 2012). This quantitative character is controlled by several genes (polygene) that have little influence and complement each other for clear observation of quantitative changes. Based on One way ANOVA test, there was no significant difference (P <0.05) in the quantitative phenotypic characters between F2 and F1 watermelon. It is shown in Table 2 where the F2 watermelon from crossing ♀ 'Putri Delima' with ♂ 'Maduri' has a fruit weight of 2.82 kg, horizontal and vertical circumference of 43.60 cm and 68.40 cm respectively while the upper and bottom diameters were 9.60 cm and 10.40 cm. Furthermore, the skin thickness of 1.64 cm, thick flesh of 11.40 cm, horizontal and vertical diameters of 14.68 cm and 26.70 cm, Brix of 9-13, fruit skin and flesh weights was 1.6 kg and 1.22 kg while the number of seeds per fruit was 234, and 4.01 gr weight of 100 seeds. Therefore, the overall quantitative character of F2 watermelon from crossing 'Putri Delima' with ♂ 'Maduri' was higher than F1. In this study, the fruit weight is greater than the results of Putri & Daryono (2017). Meanwhile, heavier fruit weight is associated with an increase in fruit diameter, pulp and seeds weight, as well as fruit skin. The sweetness level of Brix in F2 watermelon is of a higher range than F1 due to different harvesting times. In F2 watermelon, it is estimated that the harvest is on time to produce good fruit and an optimal level of sweetness compared to F1. The level of sweetness was influenced by the accumulation of sucrose, depends on genotype expression and environmental factors (Verma et al., 2017;Kyriacou et al., 2018). Moreover, the genotypes express the taste of sweet flesh due to the high accumulation of sucrose from metabolic processes while the environmental factors influence genotype during the accumulation period. Furthermore, low temperatures in the rainy season cause sugar accumulation in form of starch, while high  Generally, the phenotypic characters between F2 watermelons from crossing ♀ 'Putri Delima' with ♂ 'Maduri' with F1 watermelon by Putri & Daryono (2017) show different results a previous study stated that the difference in location, time, and weather conditions affect the resulting fruit. Furthermore, Daryono et al. (2012), stated that the differences in phenotypic characters produced between the two studies were caused by the interaction between the genotype and the environment. Meanwhile, polygene interacts with various environmental factors which affect plant growth.
Influencing environmental factors are water, nutrients, temperature, light intensity, soil moisture, and others Ilahy et al., 2019;Wang et al., 2019). Meanwhile, Suprapto & Jaya (2000) stated that the ideal environmental conditions for growing watermelons include an altitude of 100-300 m above sea level, basic topography soil which sandy or loam with a loose texture, and crumbs that contain several organic substances with a pH of 5.9-7.2, full radiation and an open space with a temperature range of 22°C to 30°C, humidity less than 80%, and an average rainfall between 40-50 mm/month.

CONCLUSION
The qualitative character of the F2 watermelon phenotype from crossing ♀ 'Putri Delima' with ♂ 'Maduri' shows segregation and non-uniformity (unstable). Therefore, it is necessary to select the desired phenotypic characters for breeding according to the consumer's point of views such as fruit shape, skin and flesh color, pulp texture, sweetness level, harvesting age, and disease resistance.