重瓣日日春之花芽形態、花形遺傳及育種

Abstract

日日春[Catharanthus roseus (L.) G. Don]喜高溫強光，花期長，是近年來全球發展快速的花卉，新品種持續育成推出。本研究比較重瓣變異日日春TYV1品系及其單瓣親本‘Pacifica Polka Dot’不同生長期的花苞，進行解剖及埋蠟切片觀察，以瞭解新增生內花瓣的起源與形態特徵。並將其栽培於不同溫度下，探討溫度對生育、開花、細胞膜熱穩定性和花粉發芽之影響。TYV1品系雌雄蕊仍具有功能，與其他單瓣品系進行正反交、自交及回交，瞭解日日春重瓣花型的遺傳模式。最後，利用上述所得形態及遺傳資訊，建立重瓣日日春新品種之選育流程。 日日春TYV1外花瓣5片輪生成高腳碟狀(salverform)，新增生內花瓣5片輪生成漏斗形(funnel-shaped)，內花瓣起源於花冠筒頂端與花瓣連接處。將TYV1及‘Pacifica Polka Dot’栽培於日夜溫30/25 ℃下，可觀察到TYV1及‘Pacifica Polka Dot’花開1-2天時柱頭上部和花藥下端重疊距離分別為0.56 ± 0.01及0.82 ± 0.02 mm。TYV1子房內胚珠數明顯較‘Pacifica Polka Dot’少。依花蕾長度將發育分為12個階段，取其中前6個階段進行解剖及切片觀察，其中第1階段(0.5-0.9 mm)時，可觀察到5枚細長萼片包圍5枚離生花瓣，其內又有5個雄蕊，而正中心皮尚未完全分化。第2階段(1.0-1.9 mm)時，5枚花瓣已相互接觸並將雄蕊及心皮包裹在其中，解剖後顯示TYV1花冠筒與花瓣交界處已可觀察到內輪花瓣原體，而‘Pacifica Polka Dot’則無此構造。第3階段(2.0-2.9 mm)外觀上萼片與外花瓣約等長，內花瓣明顯伸長且邊緣可見絨毛，切片顯示內外花瓣共用一層細胞。自第5階段(4.0-4.9 mm)開始花冠筒開始快速伸長，至第10階段(21.0-25.9 mm)才減緩。第11階段時花瓣開始展開，第12階段時花瓣完全平展。TYV1自交實生苗6對葉片時期，其葉片表面皺褶，而‘Pacifica Polka Dot’則平滑。TYV1和‘Pacifica Polka Dot’之葉柄長度分別為1.3及0.4 cm。 將日日春 TYV1及‘Pacifica Polka Dot’栽培於日夜溫20/15、25/20及30/25℃下，並調查生育開花表現及細胞膜熱穩定性。結果顯示20/15℃不利TYV1及‘Pacifica Polka Dot’生育，株高矮、花徑小、分枝數少、到花日數長，花下節位數少。以25/20℃為日日春TYV1及‘Pacifica Polka Dot’之生育適溫。日日春TYV1及‘Pacifica Polka Dot’葉片相對傷害值與水浴溫度呈S形曲線關係，以‘Pacifica Polka Dot’較TYV1有較明顯熱馴化反應，但以TYV1較耐熱。取栽培於30/25℃之花粉培養於含20%蔗糖之B&K培養基並置於10-45℃環境，結果顯示TYV1花粉發芽適溫25-30℃及‘Pacifica Polka Dot’花粉發芽適溫為20-40℃，TYV1發芽適溫範圍較‘Pacifica Polka Dot’窄。 以營養繁殖之TYV1、‘Little Pinkie’及‘Titan Burgundy’為材料，進行自交、重瓣與單瓣品系雜交(F1)、F1自交(F2)及F1與種子親回交(BCF1)等組合。結果顯示TYV1自交後代皆為重瓣型，而‘Little Pinkie’及‘Titan Burgundy’自交後代全為單瓣型。四組F1後代全為單瓣型，四個F1組合中有三組之F2後代單瓣及重瓣型植株數符合3：1 (χ2 = 0.78-1.53, P = 0.22-0.37)，二組BCF1後代單瓣及重瓣植株數符合1：1 (χ2 = 0.25-0.69, P = 0.41-0.62)，顯示日日春之重瓣花性狀由一對隱性對偶基因控制，同質或異質顯性時表現單瓣花。重瓣日日春TYV1及單瓣‘Pacifica Polka Dot’之DNA長度與二倍體的‘Little Pinkie’無顯著差異，三者分別約有1468.0、1508.1及1457.2百萬鹼基對。 以TYV1為種子親與單瓣品系TYV3及TYV4為花粉親經雜交授粉，得單瓣F1後裔，選其中15株花徑大、生長強勢單株自交得F2單重瓣分離世代族群，再經初選及兩次品系比較試驗，選出性狀優良的五個重瓣日日春品系，分別命名為‘桃園1號-玫瑰女孩’、‘桃園2號-桃花女’、‘桃園3號-紅蝴蝶’、‘桃園4號-夏雪’及‘桃園5號-紅娘’。重瓣日日春新品種側枝數及花朵數多、株高矮，適合盆花栽培，花形均具兩輪10片花瓣，花瓣排列緊密呈漏斗狀，平均花徑3.3 ± 0.3 cm。本研究所育出之重瓣日日春品種通過中華民國農業委員會植物品種審議委員會審查通過，為目前全世界首度有品種保護的重瓣日日春，權力期間至民國119年。

Periwinkle [Catharanthus roseus (L.) G. Don] is a heat and sun-loving plants with prolonged flowering duration. Cultivars have been developed rapidly in recent years. This study aimed to compare flower bud development of TYV1, a double-flowered periwinkle mutant, and its parent, single-flowered ‘Pacifica Polka Dot’ by dissection and wax sectioning to determine the origin of doubled petals. Plants of TYV1 and ‘Pacifica Polka Dot’ were also grown under different temperatures to establish cultural, cell membrane thermostability, and pollen germination information. With functioning pistils and stamens, TYV1 was cross-pollinated with single-flowered cultivars to create reciprocal F1 hybrid, F2, and back-crossed F1 generation to determine the inheritance of double-floweredness in periwinkle. Moreover, based on morphological and genetical information, the procedure of breeding and selection for double-flowered periwinkle was established. TYV1 has five salverform outer corolla lobes and five funnel-shaped inner corolla lobes. Sectioning results showed inner corolla lobes of TYV1 originated from adaxial meristem originally formed into orifice in ‘Pacifica Polka Dot’. Plants of TYV1 and ‘Pacifica Polka Dot’ were grown at day/night temperature of 30/25oC, flower buds at different developmental stages were taken for anatomical observations. The overlap between the top end of the pistil and bottom ends of anthers in TYV1 and ‘Pacifica Polka Dot’ flowers at 1-2 d after anthesis was 0.56±0.01 and 0.82±0.02 mm, respectively. TYV1 had fewer ovules per ovary than ‘Pacifica Polka Dot’. Based on flower bud longitudinal length, 12 stages were proposed for flower bud development of periwinkle and the first six stages were taken for further observations. At stage 1 (0.5-0.9 mm long), undifferentiated carpel was surrounded by five stamens, then five polypetals, and five thin sepals outermost. At stage 2 (1.0-1.9 mm long), polypetals had fused into sympetal and stamens and pistil were enclosed within. Dissecting stage 2 flowers showed that TYV1 had initiated inner corolla lobe at the transition of corolla tube and lobe, but not for ‘Pacifica Polka Dot’. At stage 3 when petals and sepals have same lengths, inner corolla lobes started to elongate and had marginal hairs. Sectioning results showed both outer and inner corolla lobes had the same originality. Corolla tube rapidly elongated from stage 5 (4.0-4.9 mm long) until stage 10 (21.0-25.9 mm long). Flower anthesis at stage 11 and corolla lobe fully expanded at stage 12. Young seedlings from self-pollinated TYV1 had crinkled leaf before the sixth pair of leaves emerged, as compare with smooth leaf surface in ‘Pacifica Polka Dot’. Petiole length of TYV1 and ‘Pacifica Polka Dot’ was 1.3 and 0.4 cm, respectively. Plants of TYV1 and ‘Pacifica Polka Dot’ were grown at day/night temperatures of 20/15, 25/20, and 30/25oC, for the evaluation of the responses of growth, flowering, and cell membrane thermostability. Results showed that plants at 20/15oC had poor growth, both TYV1 and ‘Pacifica Polka Dot’ exhibited reduced plant height, flower diameter, and branching, and more days to flowering, but fewer node number below the flower. Optimum temperature for growth and flowering of TYV1 and ‘Pacifica Polka Dot’ was 25/20oC. The relationship between the RI value, as measured with water bath temperature from 25 to 75℃ for 30 min, occurring in leaf tissue discs and the water bath temperature was sigmoidal in both TYV1 and ‘Pacifica Polka Dot’. ‘Pacifica Polka Dot’ showed more heat acclimatization response than TYV1, but TYV1 could tolerate higher temperatures than ‘Pacifica Polka Dot’. Pollen from plants grown at 30/25oC was tested for incubation temperatures of 10-45oC, and results showed that the optimum pollen germination temperature was 25-30℃ for TYV1 and 20-40oC for ‘Pacifica Polka Dot’. Vegetative propagated double-flowered TYV1, single-flowered ‘Little Pinkie’, and ‘Titan Burgundy’ were used for self-pollination, cross between single- flowered and double-flowered lines (F1), F2, and back-crossed F1 of seed parents. Self-pollinated TYV1 produced all double-flowered progeny compared to self-pollinated single-flowered lines which produced all single-flowered progeny. F1 plants between TYV1 and ‘Little Pinkie’ or ‘Titan Burgundy’ were all single. Three among four F2 population segregated into 3 single : 1 double ratio (χ2 = 0.78-1.53, P = 0.22-0.37). Two BCF1 population segregated into 1 single : 1 double ratio (χ2 = 0.25-0.69, P = 0.41-0.62). Results indicated a double-flowered form was controlled by a recessive allele. A single dominant gene expressed in the homozygous or heterozygous state resulted in the single-flowered phenotype. DNA length of double-flowered TYV1 and single-flowered ‘Titan Burgundy’ did not differ with diploid ‘Little Pinkie’, and estimated to have 1468.0, 1508.1, and 1457.2 Mbp, respectively. Double-flowered TYV1 was used as female parent and crossed with single-flowered TYV3 and TYV4. Fifteen single-flowered F1 seedlings were selected based on bigger flower diameter and vigorous growth habit. Five double-flowered lines with desirable traits were selected after the first selection and two line comparison tests, and were named as ‘Taoyuan No.1 Rose Girl ’, ‘Taoyuan No.2-Peach Lady’, ‘Taoyuan No.3-Red Butterfly’, ‘Taoyuan No.4-Summer Snow’, and ‘Taoyuan No.5-Red Lady’. The new double-flowered cultivars were suitable for potted flower production characterized with more branching, flower number per plant, and shorter plant height. All the selectrd cultivars had two whirls of compacted petals in funnel-shaped, and had flower diameter of 3.3 ± 0.3 cm. The selected new double-flowered cultivars had been certified of plant breeders’ right (PBR) by Council of Agriculture, Taiwan, and were the world’s first PBR protected double-flowered periwinkle cultivars.