| 摘要: | 本研究旨在篩選適合的培養基條件，以建立山胡椒(Litsea cubeba(Lour.) Persoon)的試管內植株再生途徑。以山胡椒之頂芽及側芽為培植體，分別探討不同消毒劑條件對山胡椒培植體消毒的效率差異，並且優化培養基成分對於山胡椒芽體發育、誘導形成叢生芽體之影響。
初代培養以NaOCl、HgCl2及PPMTM作為消毒劑處理山胡椒培植體，以1% HgCl2消毒1分鐘的消毒效果最佳，汙染率在所有處理中最低(51.2%)，以WPM基本培養基添加BAP (0, 1, 5, 10 mg L-1)或TDZ (0, 1, 2 mg L-1)組合2,4-D (0, 1, 5 mg L-1)培養培植體，以1 mg L-1 BAP處理組的萌蘗率最佳，可達42.9%。
以WPM、1/2 WPM、MS及1/2 MS四種基本鹽類培養基培養山胡椒培植體，觀察得到MS基本培養基對於培養培植體有15.1%的發芽率(P<0.01) ，優於其餘處理組。培植體初代培養4週後，將無菌芽體繼代培養於四種基本鹽類培養基，再培養6週，以1/2WPM培養基中的芽體成長狀況最為優異，不論是株高(1.8 cm)、叢生芽數量(5個)或是葉片數量(19片)，都是四種培養基中最佳；以WPM基本培養基添加BAP(0, 1, 2, 4 mg L-1)或TDZ(0, 1, 2, 4 mg L-1)組合0.5 mg L-1 2,4-D培養山胡椒無菌芽體4週，以2 mg L-1 TDZ組合0.5 mg L-1 2,4-D對於芽體誘導叢生芽數(4.5)的效果最佳。
培養過程中山胡椒芽體會出現不同程度的褐化情形，添加維生素C處理的改善效果有限。
This study aimed to establish the in vitro plant regeneration pathway of Litsea cubeba (Lour.) Persoon. The stem-nodes and the shoot tips were used as explants, and we evaluated the different disinfectant conditions on the disinfection efficiency of explants, and optimized the medium composition for the bud sprouting and multiple shoots induction.
In the primary in vitro culture, NaOCl, HgCl2 and PPMTM were used as disinfectants for the disinfection of the explants of Litsea cubeba. The results showed that the optimal disinfection condition is 1% HgCl2 1 min with the lowest contamination rate of 51.2%. Bud explants were cultured in WPM medium supplemented with TDZ (0, 1, 2 mg L-1) or BAP (0, 1, 5, 10 mg L-1) combined with 2,4-D (0, 1, 5, 10 mg L-1). The composition of 1 mg L-1 BAP showed the highest bud sprouting rate of 42.9%.
The explants were cultured in four different basal medium WPM, 1/2WPM, MS and 1/2 MS basic media, and MS medium showed the highest bud sprouting rate of 15.1% (P<0.01). In vitro shoots after primary culture of four weeks were subcultured to four basic medium for six weeks, and 1/2 WPM basal medium is optimal for shoot development and multiple shoots formation. In the combination experiment of plant growth regulators of BAP (0, 1, 2, 4 mg L-1) or TDZ (0, 1, 2, 4 mg L-1) and 0.5 mg L-1 2,4-D, the condition of 2 mg L-1 BAP and 0.5 mg L-1 2, 4-D showed the maximum multiple shoots of 4.5.
However, the shoots of Litsea cubeba showed different levels of browning during the culture. Though the application of vitamin C has only limited effect on improving the browning. |
