異源四倍體LO及LA百合微體繁殖、組培苗促成栽培 及其生育評估

Abstract

本論文利用研究室所育成異源四倍體LO及LA (4n=48, 8M+40A)百合為參試材料，探討微體繁殖其影響再生及體外結球因子，組培苗促成栽培技術，評估其生長開花表現，以及複二元體化染色體配對行為及其可稔性。異源四倍體LO百合增殖培養基以0.1 mg L-1 NAA組合0.1 mg L-1 BA不定芽再生效率較高，每一迷你鱗片培植體可誘導再生5.0±0.9不定芽，而其小鱗莖發生則以鱗片葉型為主。結球培養基以MS+ 200 mg L-1 酵母抽取物+ 80 mg L-1 腺嘌呤硫酸+ 45 g L-1 蔗糖培養8週後，對形成總變型葉數效果較佳，內輪鱗片葉可得3.6±1.4枚，而外輪鱗片葉為1.6±0.9枚。利用短暫浸漬系統(temporary immersion system, TIS)培養異源四倍體LO百合小鱗片之不定芽發生。結果顯示將小鱗片以MS固體培養基添加0.1 mg L-1 NAA+0.1 mg L-1 BA培養9週後，浸漬頻率為每4小時打氣10分鐘，浸漬6分鐘，其可提高不定芽再生數為7.0±1.2個，變型葉則為7.6±2.1枚，鱗片葉佔5.3±1.4片。異源四倍體LA百合則以單獨添加0.1 mg L-1 NAA對小鱗片再生不定芽效果較好，其小鱗莖發育接種8週後，總變型葉可形成8.6±1.9枚，但以外輪鱗片居多為4.6±1.9。MS培養基添加活性碳有利小鱗莖形成較多比例鱗片葉數。另外以1/2MS+ 2 g L-1活性碳對小鱗莖根系建立有促進作用，或添加2 mg L-1 IAA對根發生有較好之效果。 組培苗促成栽培及生育評估，異源四倍體LO百合簇生苗期表現中強生長勢，涼溫有利鱗片葉形成，生長於25/20℃鱗片葉發生速率為0.4片/週。相對地異源四倍體LA選系苗期則偏向鱗片型生長，經冷溫5℃恆溫處理有助導向鱗片葉形成，但後續結球會呈鱗片與鱗片葉型交錯發生。異源四倍體LO百合簇生苗含8~10枚鱗片葉，經冬季自然低溫處理4週再移至生長溫度條件，對照二倍體有100%抽莖，而異源四倍體LO百合僅有20%。以15/13℃處理10週後，抽莖率可達76.9%，其對冷溫單位需求仍有待探討。異源四倍體LO百合簇生苗低溫處理後至抽莖、露蕾及開花所需之生育日數為95.3±11.1天，呈現中生性。從組培苗種植到開花總生育日數為251~273天，可將歸之為早熟性。觀賞性狀表現花被平伸寬瓣而對稱，質地厚而優美。異源四倍體LO百合減數分裂於前期I完全24 II有12.7%，平均配對則有21.2±3.8雙價體。花粉母細胞減數分裂產物之小孢子數平均為4.8個。顯示，染色體延遲分離及裂開分離，此等分裂異常性易招致染色體組不整數，及超額小型孢子形成。花粉活力以Alexander’s solution檢定為3.08%，稍比二倍體佳。異源四倍體LO百合年生球周徑約14~20.8cm，隨生長溫度而異，顯示其可發展一作兩收之潛力。採收的年生球打破休眠後促成栽培，其從種植到開花生育日數為102.4±9.7天，類似中早生性。

In this research, the rapid propagation and bulbing pattern of allotetraploid LO and LA (4n=48, 8M+40A) lilies was developed in vitro. The rosette growth and phase changes of vitroplant planting were conducted under various controlled conditions. The performance in morphological and ornamental characters was evaluated, and the amphidiploids chromosome pairing and fertility were also studied. The efficently adventitious bud multiplication 5.0±0.9/mini-scale explants of allotetraploid LO lily was achieved on MS medium supplemented with 0.1 mg L-1 NAA and 0.1mg L-1 BA. The modified leaf pattern of vitroplant bulbing was dominated in scale leaf type on MS medium supplemented with 200 mg L-1 yeast extract, 80 mg L-1 adenine sulfate and 45g L-1 sucrose, than total modified leaves have 3.6±1.4 of inner scale leaves and outer scale have only 1.6±0.9. The temporary immersion system (TIS) produced 7.0±1.2 buds/mini-scale explants, and tend to scale of allotetraploid LO given 10 min/4hr immersion time and the liquid medium containing 0.1 mg L-1 NAA and 0.1 mg L-1 BA. The maximal shoot multiplication rate of allotetraploid LA lily was5.9±0.6 buds/mini-scale explant on MS medium supplement with 0.1 mg L-1 NAA alone. Howeven, the subsequent bulbing was characterized with larger portion of outer scales and a few inner scale leaves. The scale leaf bulbing growth and development was enhanced on MS medium supplemented with 2 g L-1 activated charcoal. The rooting of allotetraploid LA lily bulblet was improved on 1/2 MS medium or added with 2.0 mg L-1 IAA. The vitroplants of allotetraploid LO and LA lilies were growing at various phytotron temperatures. The rosette stage of allotetraploid LO lily mainly display scale leaf type bulbing. The unfolding rate of scale leaf was 0.4/week at 25/20℃ condition considered as middle high vigor. The allotetraploid LA lily was dominant in scale type bulbing, and could be enforced by precooling treatment. Subsequently, accompany with alternation of scale and scale leaf type bulbing.The green plants with 8-10 scale leaves of allotetraploid LO lily were exposed to nature winter cooling for 4 weeks obtained 20% shoot emergence, 15/13℃ precooling for 10 weeks obtained 76.92% shoot emergence. Contrast to the diploid LO lily that given 6 weeks precooling is saturated for shoot emergence. The total timing from shoot emergence to flowering was calculated 95.3±11.1 days, characterized as middle earliness. The total timing from planting to flowering was 251~273 days, showed as precocity. The graceful flowers appear symmetric, flatten and wild inner perianthus, and thick texture superior to diploid. The meiotic behavior of pollen mother cell was examined as 12.67% of 24 bivalents on prophase I, the average pairing was 21.2±3.8 bivalents. The average of meiotic product was 4.8. The result of chromosome delay disjunction and splitting division may conduct to aneuploid and extra microspore formation. The Alexander’s staining of pollen viability was 3.08%. The yearling bulb circumference of allotetraploid LO lily was 14~20.8 cm, which revealed highly potential for two harvest of cut flower and bulb production in one cycle. The harvested yearling was also forcible to bloom by 102.4±9.7 days after planting, resemble to middle earliness.