INTRODUCTION * Thesis imperative task Seedless citrus fruit was loving at home and abroad market. Seedless or few seed sitrus fruit caused by threefold chromosome mutation, by pollen or ovule sterity or by self-incompatibility included the self-incompatibility caused by self-pollination (Jackson and Futch, 1997; Jackson and Gmitter, 1997) [72] [71]. In the world, almost planted citrus variety was selected from natural mutation and only small ratio was formed from cross breeding program. In world market, many citrus varieties appeared from natural mutation selection, such as Navel orange, Satsuma mandarin…(Vu Cong Hau, 1996) [40] and few citrus pieces was discovered in our nation such as Nam Roi pomelo, seedless Mat orange (Tran Thi Oanh Yen et al., 2005) [35]. Sanh orange and Duong mandarin adjusted to Mekong Delta condition. Plants gave high yield, delicious quality, and got good price so that they were planted widely. However many seed in fruit was the reason that reduced the using and selling of fresh fruit. Fruits researches on Sanh orange and Duong mandarin noted that there had been many seedless or few seed cases (less than 5 seed per fruit). Hence, Can Tho university co-operated with Vinh Long Science and Technology Deparment for finding and studying seedless Sanh orange and Duong mandarin in Mekong delta. That subject had the result that two seedless Duong mandarin were discovered from Duong mandarin population in Tan Phuoc commune, Lai Vung town, Dong Thap province (Nguyen Bao Ve et al., 2007) [13]. Thus, subject “Study of seedless characteristic and morphology characteristics of seedless duong mandarin in the Mekong Delta” was carried out for having basic to develope the new seedless Duong mandarin in Mekong delta 2 * Thesis research target (i) study identification characteristics and relationship of the two seedless Duong mandarin plants to each other and with the normal seedy Duong mandarin plant; (ii) define cause for seedless character of seedless Duong mandarin plants; (iii) evaluate the stability of the seedless characteristic through the croping time, in three cultural regions of Mekong Delta, in three grafting generations and on different rootstocks; (iv) study fruit yield potential and quality of the seedless Duong mandarin in Mekong Delta. * The new of thesis It researched on new materials, the two seedless Duong mandarin plants discovered among seedy Duong mandarin population in Mekong Delta. The “late development” characteristic of ovules resulted for producing completely seedless Duong mandarin fruits. It was the characteristic helped to shape seedless Duong mandarin that was found firstly in Viet Nam. The two seedless Duong mandarin plants could be indentified by the RAPD technique (Random amplified polymorphic DNA) with the aid of DNA markers. * Contribution of thesis + Science signification Providing fairly completed information about two seedless Duong mandarin which were discovered in the Mekong Delta. The morphology characteristics of tree such as branches, leaves, flowers and fruits of the two seedless Duong mandarin plants were homogeneous and not different to the seedy Duong mandarin plant. It was able to identify the two seedless Duong mandarin plants by RAPD technique with the aid of DNA markers. There was the proximity close relationship between the two seedless Duong mandarin plants and they were closed to the normal seedy Duong 3 mandarin plant; The cause of producing completely seedless fruits on these two seedless Duong mandarin plants was the “late development” of ovule; The seedlessness characteristic of the two seedless Duong mandarin plants was stable through croping time, in intercrop condition and due to cross pollination with other species of citrus, at three different cultural regions in Mekong Delta, in three grafting generations and on three kinds of rootstocks (Mat orange, Tau lime and Hanh); These seedless Duong mandarin varieties gave good fruit yield and quality in Mekong Delta. + Reality significant In 5 years, research results helped to determine the stability of seedless characteristic of the two seedless Duong mandarin fast, other characteristics are basically not different to normal seedy Duong mandarin. They are valuable scientific basises for the early development of these precious varieties in widely production. * Main object and range of thesis + Main object of thesis are two seedless Duong mandarin were discovered in the Mekong Delta. + Range of thesis: - Observate on the mother plant and progeny scion grafting plants. - Three different cultural regions (Dong Thap, Can Tho, Vinh Long) in Mekong Delta. - In the period from 2007 to 2011. - Observation of plant morphological characteristics and genetic relationships used RAPD technique, pollen and ovule characteristics, the stability of seedless characteristic (through croping time, in three different cultural regions, in three grafting generations and the three kind of rootstocks (Mat orange, Tau Lime and Hanh), growth, yield potential and fruit quality of seedless Duong mandarin. * Thesis structure This thesis is 131-page thick, consisted four chapters with 35 tables, 34 figures and 3 appendices. There were 152 reference document. 4 CHAPTER 1 LITERATURE OVERVIEW 1.1 DUONG MANDARIN OVERVIEW Nguyen Bao Ve and Le Thanh Phong (2004) [12] suggested that Duong mandarin is belong to the Citrus genus, Eucitrus subgroup, Rutaceae family and Aurantioideae subfamily. In Mekong Delta, Duong mandarin was one of popular mandarin variety had good quality. The total Duong mandarin cultivate region was about 9640 hectares, accounting for 20 - 30% of the citrus fruit regions, focusly cultivate at Lai Vung - Dong Thap, Phung Hiep - Hau Giang, Tra On - Vinh Long, Cai Be - Tien Giang, Cang Long - Tra Vinh and scattered in some provinces (Do Minh Hien, 2008) [2]. 1.2 MOPHOLOGY CHARACTERISTIC OF DUONG MANDARIN In 5 years old, Duong mandarin plants had an average of 4.1 m height (Tran Thuong Tuan et al., 1999) [38]. According to Tran The Tuc et al. (1998) [32] and Duong Hong Dat (2003) [4], plants have thin canopy, acrodome and many secondary branches, canopy diameter is 2.5 m. Tran Thuong Tuan et al. (1999) [38] indicated that Duong mandarin had simple leaf devision, evergreen, its color was green with mark shape. According to Pham Hoang Ho (2003) [22], Duong mandarin leaf have stalk but petiole is absent, leaf blade had many conspicuous oil sacks. Its leaves often have chopped tip (Duong Hong Dat, 2000 [3] Citrus blossom was single or clusters, grew from axillary, be usually bisexual flowers (Tran Thuong Tuan et al., 1994; Nguyen Bao Ve and Le Thanh Phong, 2004; Tran Van Hau, 2009) [37]12] [39]. Full bloom have 1.3 - 1.5 cm petal length, with 5 sepals. Five white thick petals is alternate with the others and with the sepals. According to Pham Hoang Ho (2003) [22], Duong mandarin fruit had from round to slightly flattened shape, with knob at bottom and thin peel. Duong mandarin fruits has a bright yellow and smooth 5 in ripeness stage. Each fruit had average of 10.7 sections, easily separated and peeled by hand. 1.3 CITRUS POLLEN AND OVULE CHARACTERISTIC Pollen have outer wall which consisted of two membrane layers: the outer membrane and the inter membrane. According to Ha Thi Le Anh (2005), [5], inside the wall, there are cytoplasm and two nucleus: the big round somatic nucleus, which will germinate to produce pollen tubes and smaller oval reproductive nucleus, which will became two sperm. Spiegel-Roy and Goldschmidt (1996) [125] considered that the ovule of citrus have contrary attached format that the hole opposite to the axis of the ovary. Matured ovule including embryonic stem, embryo sac with eight cores and two ovule sheath. According to Jackson and Gmitter (1997) [71], the sooner or later mature (maturity, ripeness) depend on variety characteristic. This feature can be affected by inter and outster factors. 1.4 POLLINATION AND FERTILIZATION OF CITRUS The pollen is brought to stigma of the same flower or other flowers by wind or insects, it called pollination. The molecules forming the pollen surface interactions with protein and polysaccharide on stigmas. If they are compatible, the pollen will be affected to germinate. Pollen tubes germinate and penetrate into the embryo sac inside the ovule. According to Phan Kim Ngoc and Ho Huynh Thuy Duong (2000) [25], fertilization is the result of the fusion of a sperm and an embryo. Fertilization occurs after pollination. When the pollen tube enters the ovule hole, two sperm cells came out through the ovule holes, two cells escape through a hole in the wall of the pollen tube: a sperm mix with the egg, forming a diploid nucleus; the other penetrate into the large central cell contains two polar nucleus, all three of them combine together to create triploid nucleus, triploid cell develope to be endosperm, first source of nutrients for seedling. Whole of this phenomenon is called the double fertilization (Spiegel- Roy and Goldschmidt, 1996) [125]. 6 1.5 SEEDLESS CHARACTERISTIC OF CITRUS Seedless characteristic is the precious characteristic not only for all kinds of fruit but also Citrus because it was the requirement of the fresh fruit market and even the processing juice industry. Seedless characteristic of citrus has many dominant factors and was influenced by environmental conditions. Forming seedless fruit can or can not be completed, that mean it can have absolutely no seeds or the number of seeds were reduced . Autonomic parthenocarpus is a phenomenon that form seedless fruit without pollination, thus fertilization occurs without external stimulation (Spiegel-Roy and Goldschmidt, 1996)[125]. On citrus, strong sterity combined with Autonomic parthenocarpus to become the necessary condition for production seedless fruit (Ollitrault et al. 2007) [101]. Eventhough sterity is capable, seedless fruit can not be produced because of incapable autonomic parthenocarpus. Ollitrault et al. (2007) [101] considered that the Autonomic parthenocarpus, caused seedlessness without pollination, was a key process in citrus. According Ollitrault et al. (2007) [101], several varieties of male sterility and self-incompatibility can not form seedless fruit because autonomic parthenocarpus was absent. Therefore, the autonomic parthenocarpus ability are indispensable characteristics for the production of seedless fruits and this seem to be popular on citrus. Mooney (1997) [89] found that triploid Citrus have great commercial potential because of the high degree of seedless, but the frequency of triploid plants were very low in the wild. Reed (2003) [113] thought that self-incompatibility is the disfunction of the male and female gametes to form seeds when self-pollinated occurs, the mismatch of the male gametes on the same species stigmas, or prevent the growth of the pollen in the same species styles. The self- incompatibility will create seedless fruit when they were not planted with the other spieces. Self male sterility can result from abnormal development of the stamens leaded to deficiencies in the pollen development process. Tran Thuong Tuan (1992), [36] indicated that unviable male gametes caused of the male sterility. Female sterility is a very important trait 7 which closely related to the seedlesss characteristics and heritability (Yamamoto et al., 1995; Yamamoto et al., 2001) [149] [148]. CHAPTER 2 MATERIAL, CONTENT AND RESEARCH METHOD 2.1 CONTENT AND RESEARCH METHOD Including 4 part, with 8 observations and 5 experiments: 2.1.1 Study of mophology characteristic and genetic relationship Consists of two observations in morphological characteristics and use of RAPD technique to identify and determine the relationship of the two seedless Duong mandarin which had been discovered in the Mekong Delta. 2.1.2 Study of pollen and ovule of seedless Duong mandarin There are 2 pollen observations and 3 ovule observations for finding out the characteristics of pollen and ovule that were related to the seedless characteristics. 2.1.3 Evaluate the stability od seedless characteristic Includes an observation and five experiments to evaluate the stability of seedless characteristic through croping time, on 3 cultivation regions in the Mekong Delta, in 3 grafting generation and on different rootstocks. 2.1.4 Evaluate yield potential and fruit quality of seedless Duong mandarin Evaluate the growth ability, yield potential and fruit quality in scion grafting plants with Mat orange rootstock in Lai Vung district, Dong Thap province, where is one of the typical mandarin cultivate region in Mekong Delta. 2.2 REGIONS, MATERIAL AND RESEARCH PERIOD The study was carried out in Dong Thap, Can Tho and Vinh Long. Main material are seedless Duong mandarin No. 1 (code 1), seedless Duong mandarin No. 2 (code 80) and normal seedy Duong mandarin as controls (code 63) (Nguyen Bao Ve et al., 2007) [13]. The research period is from 2007 to 2011. 8 CHAPTER 3 RESULTS AND DISCUSSIONS 3.1 MOPHOLOGY CHARACTERISTIC AND GENETIC RELATIONSHIP OF SEEDLESS DUONG MANDARIN The results showed that canopy got elliptical shaped, vertical posture, branch density were sparse, scion direction and branching devision much, sparse spine density, spine is straight and short, smooth tree-trunk and angle of branching were moderate. In general, the mophology characteristics of the two seedless Duong mandarin were same to normal seedy Duong mandarin. Leaf morphological characteristics between the two seedless Duong mandarin is same to each other and not different with seedy Duong mandarin. The flowers characteristics of two seedless Duong mandarin are the same and similar to seedy mandarin. Time from the onset of bud to the blooming of the two seedless Duong mandarin is about 11 - 12 days, from blooming to stigma fall is about 2 - 3 days, the same to normal seedy Duong, mandarin (control). These shows that development period and capable blooming period for receiving pollen in two seedless Duong mandarin is normal, and not different to seedy Duong mandarin (control). The flower development of seedless Duong mandarin No. 1 is same with seedless Duong mandarin No. 2 and be same to seedy Duong mandarin(control) Fruits of two seedless Duong mandarin and fruits of seedy Duong mandarin have the same shape. They are big circular and rather flat. Bottom of fruit have knob, top of fruit got slightly concave. Peel transfer to smooth yellow Green when ripe. Basically, the morphological characteristics between the two seedless Duong mandarin are same, and not different to seedy Duong mandarin (completely seedless characteristic). In short, except the completely seedless characteristic, morphological characteristics such as stems, leaves, flowers and fruits are the same between the two seedless Duong mandarin and seedy Duong mandarin (control). 9 Figure 3.4 Seedless Duong mandarin fruit Shape Due to analyzing results, all 7 used primer steady brought useful magnification products with the aid of RAPD technique. Using RAPD technique with seven primers used, a total of 46 tape recorded. Including nine tapes appear polymorphisms in the primer SO15 (1), SN20 (4 tape), SN06 (2 tape) and A13 (2). SO15 and A13 primers can distinguish two seedless Duong mandarin with seedy Duong mandarin. By using primers A13, seedy Duong mandarin had the amplification of DNA at two positions 650 bp and 750 bp (noted a and b on figure 3.5), while the two seedlesss Duong mandarin do not give amplification products in this location. Meanwhile, the prey SN06 and SN20 can distinguish two seedless Duong mandarin to each other and with the seedy Duong mandarin (to distinguish the three plants together). Seedless No. 1 Seedless No. 2 Seedy (control) Up side Downside Cross side 10 Figure 3.5 Seedless Duong mandarin PCR record with A13 primer M: standard meter 1 kb; 1: seedless Duong mandarin No.1; 2: seedless Duong mandarin No.2; 3: seedy Duong mandarin (control). Figure 3.8 shows two seedless Duong mandarin and seedy Duong mandarin as control fusion to be a main branch. It included seedy Duong mandarin, seedless Duong mandarin No. 1 and seedless Duong mandarin No. 2 with the high coefficient (0.87). Two seedless Duong mandarin constitute a secondary branch and they have the same coefficient (0.92). Thus, the two seedless Duong mandarin had the close genetic relationship to each other and they have the close genetic relationship to seedy Duong mandarin. Figure 3.8 Branch diagram of seedless Duong mandarin 1: seedless Duong mandarin No. 1; 2: seedless Duong mandarin No. 2; 3: seedy Duong mandarin (control)). In summary, the morphological characteristics of two seedless Duong mandarin are basically same about trunk, branches, leaves, 0, 87 0,92 M 1 2 3 a b 1.000 500 650 850 11 flowers and fruits and not different seedy Duong mandarin (excluding completely seedless characteristic). It was able to identify two seedless Duong mandarin by RAPD technique with DNA markers aid. Beside, the genetic relationships analysing result showing two seedless Duong mandarin have close relationships to each other and close to the commercial seedy Duong mandarin in the region. 3.2 POLLEN AND OVULE CHARACTERISTIC OF TWO SEEDLESS DUONG MANDARIN 3.2.1 Pollen characteristic Observations results showed that the pollen germination and pollen tube length of the seedless Duong mandarin No. 1 and seedless Duong mandarin No. 2 seed were not significant different to seedy Duong mandarin (control). Force pollination results also showed that using natural pollen of seedless Duong mandarin No. 1 and No. 2 in seedy Duong mandarin, the number of full seeds/ fruit, flat seeds/ fruit were no significant differences to each other and to seedy Duong mandarin (control). It showed that pollen of seedless Duong mandarin were unsterile. Thus, seedless characteristics of the two seedless Duong mandarin is not caused by pollen. 3.2.2 Ovule characteristic 3.2.2.1 Ovule Developement * Before full bloom stage In seedy Duong mandarin, ovule appeared early and their quantity was stable on 5 flower stage (from 10.4 to 10.8 ovule). In contrary, there are completely no ovule in seedless Duong mandarin No. 1 and No.2 till full bloom stage. Late development characteristic in full bloom stage may help to form seedless fruit in seedless Duong mandarin No.1 and No.2 Table 3.18 and Table 3.19 show that in small flowers (flower diameter is 3.5 mm), the ovule size of seedy Duong mandarin was quite large (203 µm length and 130 µm width) and rapidly developed in 5 mm. from this moment till the full bloom stage, the ovule size of seedy Duong mandarin virtually unchanged (396 - 404 µm length and 12 246 - 258 µm width). At this time, the ovule had the pear shape (Figure 3.14). The stability of the size of the ovule in the period before and on flowering time allows to infer that ovule was mature (ripe) and may be ready for fertilization. Ovule growth characteristics on flowering time is a common feature of crops, because it is the best time for pollination, fertilization. This feature has helped commercial seedy Duong mandarin to have normal seeds. Thus, in the cross-sectional area of the ovary surgery of seedless Duong mandarin, mature ovule is pear-shaped and about the size of 404 ± 21 µm x 258 ± 23 µm. Table 3.18 Ovule width (µm) at different flower size of seedless Duong mandarin Flower size (diameter, mm) 3,5 4,0 4,5 5,0 Bloo m Mandarin tree X Sd X Sd X Sd X Sd X Sd Seedless No.1 0 - 0 - 0 - 0 - 0 - Seedless No.2 0 - 0 - 0 - 0 - 0 - Seedy (control) 203 20 279 28 331 48 396 35 404 21 X : average; Sd: standard deviation; Table 3.19 Ovule width (µm) at different flower size of seedless Duong mandarin Flower size (diameter, mm) 3,5 4,0 4,5 5,0 Bloom Mandarin tree X Sd X Sd X Sd X Sd X Sd Seedless No.1 0 - 0 - 0 - 0 - 0 - Seedless No.2 0 - 0 - 0 - 0 - 0 - Seedy (control) 130 17 202 32 222 27 246 17 258 23 X : average value; Sd: standard deviation; 13 Figure 3.14 Ovary Cross slices before blooming of seedless Duong mandarin; zoom 100 time. Flower with 4 mm diameter Flower with 4.5 mm diameter Flower with 5 mm diameter Blooming Seedless No. 1 Seedless No. 2 Seedy (control) Flower with 3.5 mm diameter 100 µm 14 * After blooming stage On seedy Duong mandarin, number of ovule remained stable at 3 flower size after bloom (11.0 - 11.3 ovule). Ovule of two seedless Duong mandarin appeared similarly and later than seedy Duong mandarin ovules. Till the petal fall stage, it recored that ovule appeared, but the number of ovule of seedless Duong mandarin is less than the number of ovule seedy Duong mandarin (2.4-4.5 ovule). Figure 3.15 Cross slice of ovary after blooming of seedless Duong mandarin; zoom 100 time. Seedless Duong mandarin ovule have unchange width (in the range of 266 - 270 µm), the length of ovule developed followed Petal fall Stigma fall Fruit At 2.5mm diameter 100 µm Seedless No. 1 Seedless No. 2 Seedy (control) 15 flower side (petal fall: 411 µm, stigma fall: 443 µm, fruit sixe at 2.5 mm: 470 µm). The change of this size may be due to the process of developing seeds after the fertilized. Meanwhile, in two seedless Duong mandarin there are few ovules appear, but the size is smaller, petal fall had the range of 77-81μm length and in the range of 63 - 66 µm width, in stigma fall, the length is in the range of 127 - 131 µm and at fruit size 2.5 mm had length in 191 - 199 µm. The results showed that, in seedless Duong mandarin No. 1 and No. 2, till 2.5 mm diameter fruit, ovule is still developing, has not yet reached mature size. So maybe, ovule of two seedless Duong mandarin still immature at this time. In summary, the ovule of two seedless Duong mandarin had the same "late development" characteristics to the ovule of seedy Duong mandarin. May be the ovule characteristics "late development" is cause for seedless fruit is seedless Duong mandarin. 3.2.2.2 Pollent tube appearance in ovary * Self pollination condition In self-pollination condition, in all parts of the female structure (stigma, above styles, below styles and ovary) at 3 days after pollination of the three plants, the observation reported the presence of pollen tubes (Figure 3.16). The observation results also demonstrated for ovule characteristics of seedy Duong mandarin had grown in the blooming stage because of the fertilization occurence (pollen tube came through the hole in the ovule and came to embryo sac) . Meanwhile, in seedless Duong mandarin No. 1 and No. 2, pollination still occurs, pollen tubes reach to the ovary, but ovule is immature at this time so fertilization can not occur so that it can not create seeds. It also confirmed the late development characteristics of two seedless Duong mandarin(Figure 3.16). 16 Figure 3.16 Slice of female tructure of flower on 3 days after sefl pollination of seedless Duong mandarin; zoom 100 times; white narrow direct the pollen tube. * Open pollination condition The two seedless Duong mandarin and seedy Duong mandarin on 3 days afer pollination, with 3 types of pollination (self pollination, pollinate with seedy Duong mandarin pollen and 100 µm Stigma Above styles Below styles Ovary Seedless No. 1 Seedless No. 2 Seedy (control) 17 pollinate with Mat orange pollen), most parts of the female structure (stigma, above styles, below styles and ovary) were recorded that there were the presence of pollen tubes (93.3 to 100%), difference was not significant by χ2 testing. 3.2.2.3 Ovule fertilization bability Seedless Duong mandarin No. 1 and No. 2 had fruits with absolutely no-pollinated seeds in 3 types of pollination. Meanwhile, the seedy Duong mandarin with 3 types of pollination have the formation of seeds, number of full seeds/fruit was in range of 9.07 to 11.53 and the number of immature seeds/ fruit was 1.93 to 2.03 3.2.3 Discussion about seedless Cause Flowering stage is the best time for pollination, fertilization and forming seed. At this time, in the seedy Duong mandarin, through morphological anatomy cross-sectional ovary, most ovule is mature, perhaps due to this characteristic made famous commercial Duong mandarin in the Mekong Delta has many seeds. Meanwhile, ovule of two seedless Duong mandarin were still immature. By the petal fall stage (about 2 - 3 days after blooming), the ovule of two seedless Duong mandarim appeared, but they got small size and far from mature size. So, whether pollination at this time (very difficult to occurs due to petal fall, dried stigma, color changed and no longer to receive pollen anymore), then fertilization is difficult to implement due to ovule is not mature. The "late development" characteristic was not in sync with other progression of the flower is the cause of seedlessness of the two seedless Duong mandarin. Characteristics that ovule will be mature for few days later than the time of blooming was recorded on Washington Navel oranges and Satsuma tangerine (Jackson and Gmitter, 1997) [71], is the world's seedless citrus. Although possible pollination process but fertilization can not occurs because there is no mature ovule, so that it form the seedless fruit. In summary, two seedless Duong mandarin pollen were unsterile, They are capable to germinate, fertilize and form seeds as seedy Duong mandarin, hence two seedless Duong mandarin pollen were not related to seedlessness. Seedlessness characteristic of two seedless Duong mandarin is not related to self-incompatibility 18 because pollen tubes was still present in the ovary itself and totlaly seedless when cross-pollinated with other citrus (Duong mandarin seeds, Sanh orange). "Late development" characteristic caused seedlessness of seedless Duong mandarin No. 1 and No. 2 . 3.3 STABILITY OF SEEDLESSNESS CHARACTERISTIC 3.3.1 Stability of seedlessness through croping time The results showed that all fruits in the 5-year observation from 7 years old until 11 years old, the trees have absolutely seedless fruits (no mature seeds and no immature seeds) in intercrop conditions with other citrus (Duong mandarin, Tieu mandarins, Tau lime, Mat orange, Sanh orange). Completely seedlessness characteristic of the two seedless Duong mandarinare stable through croping time in intercropped condition with other citrus varieties. 3.3.2 Stability of seedlessness in different cultivation regions Results indicated that there are no presence of ovule in most ovary at the flowering time in seedless Duong mandarin No. 1 and No. 2 in all 3 experiments regions (Dong Thap, Can Tho, Vinh Long). At the time of 3 days after blooming (petal fall), in all 3 experiments regions, the ovule in seedless Duong mandarin No. 1 and No. 2 ovules developed more than blooming stage, but there are few ovule with small size. All harvest fruits were completely seedless . Meanwhile, the seedy Duong mandarin in three point Lai Vung - Dong Thap, Can Tho city and Tra On Vinh Long sequential of full seed/ fruit is 8.56, 10.26 and 12.22; and immature seeds / fruit was 2.33, 2.08 and 2.36. In short, absolutely seedlessness characteristics of two seedless Duong mandarin may have genetic ability and maintain stable in clonal generations (on Mat orange rootstock) in 3 cultivation regions (Dong Thap, Can Tho and Vinh Long) of Mekong Delta. 3.3.3 Stability of seedlessness in grafting generations Ovary observation results showed that do not have the presence of the ovule at the blooming stage. At three days after blooming stage there are no ovule with the mature ovule morphology and size (ripening). Two seedless Duong mandarin through three generations 1 a 1 b 19 on Mat rootstock gave absolutely no seeds in the fruit of all treatments. In short, absolutely seedlessness characteristics of seedless Duong mandarin No. 1 and No. 2 have the genetic ability and maintain stability in the three grafting generations on Mat orange rootstock. 3.3.4 Stability of seedlessness on different rootstock At the blooming stage and 3 days after blooming stage, threr are no ovule with mature ovule morphology and size in all treatments. Full seeds in all treatments was 0.00. Immature seeds also have similar results (0.00). This is approriate with the recorded that ovule at the blooming stage and 3 days after blooming stage. The late development characteristics of ovule remain stable at seedless Duong mandarin on different rootstock. Thus, seedlessness characteristics of seedless Duong mandarin remain stable in the next generation by asexual multiplication on different rootstock (Mat orange, Tau lime and Hanh) (Figure 3.26). 3.3.5 Seedlessness characteristic stability disscussion Results of ovule presence recorded from the experiments showed that late development characteristics of seedless Duong mandarin No. 1 and No. 2 ovules were maintained when they had grown in three cultivation regions in the Mekong Delta, on three grafting generations and on 3 kinds of rootstock (Mat orange, Tau lime and Hanh). Thus, the Late development characteristic of ovules depends largely on the variety property. According to Jackson and Gmitter (1997) [71] maturity characteristics of the ovule occurs sooner or later will depends mainly on seeds, Foster grapefruit ovule are mature on few days before the blooming stage, Pineapple Orange ovule are mature right on blooming stage and Washington Navel orange and Satsuma mandarin ovule are mature few days after blooming stage (these are 2 famous seedless citrus in the world). Due to late development characteristics of ovule may have genetic ability, that helped the seedlessness characteristics stability of two seedless Duong mandarin discovered in Mekong Delta in high 20 degree. The absolutely seedlessness characteristic remained stable through croping time, in intercropped with other citrus; in three cultivation regions of Mekong Delta; on 3 grafting generations, and on 3 different rootstocks (Mat orange, Tau lime and Hanh). This result allows to propagate rapidly seedless Duong mandarin No. 1 and No.2 by grafting on 3 types of rootstock (Mat orange, Tau lime and Hanh), planted in regions that had comfortable condition for growing citrus in Mekong Delta and especially can be intercropped with other citrus that absolutely seedlessness characteristic remained stable. However, accordiing to Jackson and Gmitter (1997) [71] sooner or later maturity characteristics of the ovule can be affected by a number of agents inside and outside. So, there may be some factors that make the development of seedless Duong mandarin No. 1 and 2 ovule mature earlier and the time that stigmas receive pollen will be longer (the period from blooming to petal fall for about 2 - 3 days in seedless Duong mandarin), then this time can occur fertilization and formation of seeds in seedless Duong mandarin. This issue should be studied further. 3.4 YIELD POTENTIAL AND FRUIT QUALITY OF SEEDLESS DUONG MANDARIN 3.4.1 Growing of tree Diameter of rootstock, scion diameter, the ratio of rootstock diameter and scion diameter, plant height, canopy width through croping time showed that there are no difference between three treatments. Therefore, the growth of seedless Duong mandarin No. 1 No. 2 seed is not equivalent to each other and do not different with seedy Duong mandarin which were grown popular in the Mekong Delta. 3.4.2 Yield Percentage of fruit set can be used to evaluate the yield potential of seedless Duong mandarin No. 1 and No. 2. Table 3.32 shows the percentage of fruit set of three treatments with the general decreasing trend in the period from 30 - 90 days after flowering, but [...]... and due to cross pollination with other species of citrus, at three different cultural regions in Mekong Delta, in three grafting generations and on three kinds of rootstocks (Mat orange, Tau lime and Hanh) - These seedless Duong mandarin varieties gave good fruit yield and quality in Mekong Delta 4.2 SUGGESTION - Studying continuously about yield and fruit quality of seedless Duong mandarin at three . focusly cultivate at Lai Vung - Dong Thap, Phung Hiep - Hau Giang, Tra On - Vinh Long, Cai Be - Tien Giang, Cang Long - Tra Vinh and scattered in some provinces (Do Minh Hien, 2008) [2]. 1.2. orange, Tau lime and Hanh). This result allows to propagate rapidly seedless Duong mandarin No. 1 and No.2 by grafting on 3 types of rootstock (Mat orange, Tau lime and Hanh), planted in regions. and progeny scion grafting plants. - Three different cultural regions (Dong Thap, Can Tho, Vinh Long) in Mekong Delta. - In the period from 2007 to 2011. - Observation of plant morphological