繁星花之花粉發芽、花色遺傳及盆花選育

Abstract

繁星花[Pentas lanceolata (Forsk.) Delf.]耐旱、耐高溫且多年生性，為熱帶及亞熱帶地區重要草花，花色豐富。其育種目標為大花、多花、大花序且多分枝，但現今育出品種變化不大。具有雌雄蕊異長(heterostyly)的兩型花柱(distyly)特性。本研究擬探討短花柱型(thrum form)與長花柱型(pin form)之花粉發芽特性及雜交障礙類型，另亦探討花色及雙色性狀遺傳。建立性狀調查表，並選育具潛力新品系。 取開花當天之繁星花‘Butterfly Deep Pink’與‘Butterfly Red’兩型花柱之花粉置於含15%蔗糖之BK (Brewbaker及Kwack)培養基中，在25℃、黑暗培養1、2、4或6 h，觀察花粉發芽率。結果顯示長花柱型在2 h有最高花粉發芽率；短花柱型則需培養4 h。兩品種之長花柱型花粉發芽率明顯比短花柱型高。 觀察繁星花‘Butterfly Deep Pink’與‘Butterfly Red’開花前一天、開花當天及開花後一天兩型花柱之花冠筒內部縱切結構。結果不論長花柱型或短花柱型，於開花前一天之花藥均飽滿且未開裂，開花當天可見花藥外壁裂開且塌陷，伴隨花粉釋出，開花後一天釋出更多花粉。 分別於開花當天至開花後四天取兩型花柱之繁星花‘Butterfly Deep Pink’與‘Butterfly Red’之花粉，置於含15%蔗糖之BK培養基中，在25℃、黑暗培養2 h。結果兩品種之長花柱型花粉皆於開花當天有最高之發芽率，隨開花日數增加而下降，而短花柱型則於開花後一天才有最高之發芽率，亦隨日數增加而逐漸下降。開花當天與開花後一天，長花柱型‘Butterfly Deep Pink’花粉發芽率顯著高於短花柱型，而‘Butterfly Red’則只有開花當天以長花柱型顯著較高。於開花當天取兩型花柱之繁星花‘Butterfly Deep Pink’與‘Butterfly Red’之花粉，分別置於含0%、5%、10%、15%或20%蔗糖之BK培養基中，在25℃、黑暗培養2 h。結果顯示兩品種之長花柱型花粉皆以含15%蔗糖BK培養基有最高發芽率，而短花柱型‘Butterfly Deep Pink’及‘Butterfly Red’達最高發芽率之蔗糖濃度分別為5%-10%及10%。不論蔗糖濃度，兩品種長花柱型花粉發芽率明顯較短花柱型高。於開花當天分別取繁星花‘Butterfly Deep Pink’與‘Butterfly Red’兩型花柱之花粉，置於含15%蔗糖BK培養基中，分別在0、10、20、30、40及50℃下暗培養2 h。結果顯示兩品種之長花柱型花粉皆以20-40℃有較高發芽率，而短花柱型‘Butterfly Deep Pink’及‘Butterfly Red’達較高發芽率之溫度分別為20-30℃及20-40℃。以乳酸酚棉藍檢驗兩型花柱花粉敗育率之差異，結果顯示長花柱型之花粉稔性明顯比短花柱型高。 以短花柱型及長花柱型的‘Butterfly Deep Pink’、‘Butterfly Red’為材料，自交及雜交後觀察雌蕊內花粉管螢光。結果顯示，繁星花短花柱型雜交障礙類型應屬於孢子體型之受精前障礙；短花柱型自交成功與否依品種而異。自交及雜交所得種子培育至開花，調查後代兩型花柱植株數量比。結果顯示短花柱型(S_)相對於長花柱型(ss)呈顯性。 取‘Butterfly’、‘Graffiti’、‘Kaleidoscope’、‘New Look’、‘Bismarch’及‘Starla’等品種進行自交及雜交，分析後代花冠裂片顏色遺傳。結果顯示繁星花五種花色之顯性順序為紅(Rd_)>桃(Rs_)>紫(V_)>粉紅(P_)>白。當Rd基因為顯性，花色便會形成紅色(Rd_ _ _ _ _ _ _)；當Rd基因為隱性，Rs為顯性，花色會呈桃色(rdrdRs _ _ _ _ _)；當Rd及Rs基因為隱性，V為顯性，花色會呈紫色(rdrdrsrsV_ _ _)；當Rd、Rs和V基因為隱性，P為顯性，花色呈粉紅色(rdrdrsrsvvP_)；而四對基因均呈隱性，則會呈現白色花(rdrdrsrsvvpp)。 另萃取繁星花花冠裂片之花青素並以質譜儀分析，結果顯示紅色的主要花青素(anthocyanin)組成分為矮牽牛素(petunidin)、錦葵素(malvidin)及玫瑰色素(rosinidin)；桃色(深桃與淡桃)均為芍藥素(peonidin)、錦葵素及矮牽牛素；紫色為錦葵素與芍藥素；粉紅色則為飛燕草素(delphinidin)。 將具雙色性狀之選育單株與其他選育單株及商業品種進行自交與雜交。結果顯示雙色性狀受兩獨立遺傳基因控制。以Dc為表現雙色之基因，以Pm為累積雙色性狀色素之基因，當Dc與Pm同時為顯性(Dc_Pm_)，表現雙色性狀，基因型為dcdcPm_時呈純桃色，基因型Dc_pmpm或dcdcpmpm則呈現其他花色。 本研究參照UPOV (International Union for the Protection of New Varieties of Plants)及日本性狀檢定表，從雜交後代中選出純色品系GVKA-1及BRdGRs-13，具眼圈品系BRdKA-7及BRdKA-9，具漸層性狀品系GRsKA-57及KAGV-9。其中，以GVKA-1、BRdGRs-13、BRdKA-7及BRdKA-9分別與對照品種Graffiti Violet、Butterfly Red、Ceylon Hybrida及Kaleidoscope Appleblossom進行性狀比較。結果顯示，GVKA-1分枝性明顯較佳、節位多、葉小、花大且花色更深。BRdGRs-13明顯較低矮，莖節間短且花大。BRdKA-9分枝性明顯較佳、節位多，葉小且花色新穎。BRdKA-7明顯較低矮，莖節間短且花大、花序大，且新穎花色。

Star cluster [Pentas lanceolata (Forsk.) Delf.] is drought- and heat-tolerant, thus an important herbaceous perennial ornamental suitable for tropical and sub-tropical areas. The breeding objectives are larger florets, floribunda, larger inflorescences and good branching. However, major cultivars did not provide much variation. This study aimed to investigate pollen germination requirement and hybrid barriers between distylic plants, and the inheritance of floral colors and bicolor traits. This thesis also includes and selects Pentas cultivars lines with market potentials. Pollen from distylic florets of ‘Butterfly Deep Pink’ and ‘Butterfly Red’ at anthesis were cultured in BK medium containing 15% sucrose and incubated in dark at 25°C for 1, 2, 4, or 6 h. Results showed that highest pollen germination percentage was recorded after culture for 2 and 4 h for pin and thrum forms, respectively. Pollen from pin flowers had higher germination percentage than thrum flowers in both cultivars. Distylic florets of ‘Butterfly Deep Pink’ and ‘Butterfly Red’ were observed longitudinally at one day before anthesis, at anthesis, and one day after anthesis. Regardless of floret type, the anthers were full and intact at one day before anthesis. Dehiscence of anthers and pollen release could be observed at anthesis, and more pollen released at one day after anthesis. Pollen from distylic florets of ‘Butterfly Deep Pink’ and ‘Butterfly Red’ was collected from anthesis to four days after flowering, and cultured in BK medium containing 15% sucrose, and incubated in dark for 2 h at 25°C. Pollen from pin and thrum florets had the highest germination percentage at anthesis and one day after anthesis, respectively, and decreased gradually afterwards in both cultivars. Germination percentage was higher for pollen from pin florets than thrum florets at anthesis and one day after anthesis in ‘Butterfly Deep Pink’. While significant difference was observed only at anthesis in ‘Butterfly Red’. Pollen from distylic florets of ‘Butterfly Deep Pink’ and ‘Butterfly Red’ was collected at anthesis and cultured in BK medium containing 0%, 5%, 10%, 15%, or 20% sucrose, then incubated in dark for 2 h. Results showed that pin pollen had the highest germination percentage with 15% sucrose. The optimum sucrose concentration for thrum pollen was 5%-10% for ‘Butterfly Deep Pink’ and 10% for ‘Butterfly Red’. Pollen germination percentage was higher in pin than thrum florets regardless of sucrose concentration in both cultivars. Pollen from distylic florets of ‘Butterfly Deep Pink’ and ‘Butterfly Red’ was taken at anthesis and cultured in BK medium containing 15% sucrose, then incubated at 0, 10, 20, 30, 40, or 50°C in dark for 2 h. Results showed that the optimum temperature for pin pollen ranged between 20 and 40°C in both cultivars. The optimum temperature for thrum pollen was 20-30°C for ‘Butterfly Deep Pink’and 20-40°C for ‘Butterfly Red’. Pollen abortion percentage was examined by lactophenol-aniline blue. Results showed that pin pollen was more fertile than thrum pollen. Distylic plants of ‘Butterfly Deep Pink’ and ‘Butterfly Red’ were self- and cross-pollinated and in vivo pollen tube growth was observed using aniline-blue fluorescent staining. Results showed some thrum florets had sporophytic incompactibility and pre-fertilized hybrid barrier. Seedlings of above crosses were raised and observed for pin and thrum plants when flowering. Results showed that thrum type (S_) is dominant to pin type (ss). Self- and cross-pollination between cultivars Butterfly, Graffiti, Kaleidoscope, New Look, Bismarch, and Starla were made to determine the inheritance of corolla lobe color. Results showed that the dominance of corolla lobe color is red (Rd_) > rose (Rs_) > purple (V_) > pink (P_) > white. When the Rd gene was dominant, the corolla lobe color would be red (Rd_ _ _ _ _ _ _ ); when the Rd gene was recessive, Rs was dominant, the corolla lobe color would be rose (rdrdRs _ _ _ _ _ ); when Rd and Rs gene were recessive, V was dominant, the corolla lobe color would be purple (rdrdrsrsV_ _ _). When Rd, Rs, and V genes were recessive, P was dominant, the corolla lobe color would be pink (rdrdrsrsvvP_); when the four genes were all recessive, white corolla lobe (rdrdrsrsvvpp). Anthocyanin extraction and mass spectrometry analysis of the corolla lobes were made. Major anthocyanin compositions of red corolla lobes are petunidin, malvidin, and rosinidin. Peonidin, malvain, and petunia were found in rose corolla lobes (deep and pale). Purple corolla lobes composed malvidin and peonidin majorly. Pink corolla lobes contained delphinidin. The selected progenies with bicolor traits were self- and cross-pollinated. Results showed that bicolor trait was governed by two independent loci, Dc and Pm. Dc was proposed to be responsible for the expression of bicolor, and Pm is responsible for the accumulation of pigments. Bicolor trait present when Dc and Pm were proposed to be both dominant (Dc_Pm_), but solid rose when the genotype was dcdcPm_, and other colors when the genotype was Dc_pmpm or dcdcpmpm. DUS test guideline was developed for Pentas. Pure color lines GVKA-1 and BRdGRs-13, eye trait lines BRdKA-7 and BRdKA-9, and dilute trait lines GRsKA-57 and KAGV-9 were selected from the hybrid progeny and compared with the comparative cultivars. Results showed that GVKA-1 has better branching, multiple nodes, smaller leaves, larger florets, and deeper floral color. BRdGRs-13 is shorter, with shorter internodes, and larger florets. BRdKA-9 has more branching, multiple nodes, smaller leaves, and new floret color trait. BRdKA-7 had shorter internodes, larger florets and inflorescences, and new flower color.