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园艺学报 ›› 2014, Vol. 41 ›› Issue (10): 2139-2146.

• 研究报告 • 上一篇    下一篇

红姜花体细胞胚胎发生及植株再生的研究

涂红艳,肖 望*,邓崇会   

  1. 广东第二师范学院生物系,广州 510303
  • 收稿日期:2014-04-01 出版日期:2014-10-25 发布日期:2014-10-25
  • 基金资助:

    广东省自然科学基金博士启动项目(10451030301004286);2013年广东省高等院校学科与专业建设专项资金项目(2013KJCX0137);广州市科技计划项目(2014J4100151)

Somatic Embryogenesis and Plant Regeneration of Hedychium coccineum

TU Hong-yan,XIAO Wang*,and DENG Chong-hui   

  1. Department of Biology,Guangdong University of Education,Guangzhou 510303,China
  • Received:2014-04-01 Online:2014-10-25 Published:2014-10-25

摘要: 以红姜花(Hedychium coccineum)的花丝和花药为外植体,通过体细胞胚胎发生途径建立了植株再生体系。结果表明:外植体在MS + 4 mg ? L-1 2,4-D + 4 mg ? L-1 NAA + 1 mg ? L-1 6-BA + 30 g ? L-1 蔗糖 + 7 g ? L-1 琼脂的培养基上经过120 d诱导出愈伤组织,愈伤组织在MS + 1 mg ? L-1 2,4-D + 0.25 mg ? L-1 NAA + 0.25 mg ? L-1 6-BA + 30 g ? L-1 蔗糖 + 7 g ? L-1琼脂的培养基上经过继代筛选,获得浅黄色松散易碎的胚性愈伤组织。胚性愈伤组织在MS无机盐 + B5维生素+ 100 mg ? L-1谷氨酰胺 + 230 mg ? L-1 脯氨酸 + 100 mg ? L-1麦芽提取物 + 0.02 mg ? L-1 NAA + 0.02 mg ? L-1 TDZ + 0.5 ~ 1.0 mg ? L-1 2,4-D + 45 g ? L-1蔗糖的培养基中通过3个月的悬浮培养,可得到均质稳定的胚性细胞悬浮系(ECS)。胚性悬浮细胞在SH无机盐 + B5维生素 + 100 mg ? L-1谷氨酰胺 + 230 mg ? L-1脯氨酸 + 100 mg ? L-1麦芽提取物 + 0.25 mg ? L-1 NAA + 0 ~ 0.20 mg ? L-1 TDZ + 45 g ? L-1蔗糖 + 7 g ? L-1琼脂的体胚诱导培养基中培养10 d,可见到白色半透明体胚发生,20 d后体胚发育成熟。当TDZ浓度为0.15 mg ? L-1时,体胚诱导率高达4 500个 ? mL-1 PCV ECS(PCV:细胞密实体积)。在SH无机盐 + B5维生素 + 0.20 mg ? L-1 IAA + 0.25 ~ 1.0 mg ? L-1 6-BA + 30 g ? L-1蔗糖 + 7 g ? L-1琼脂的体胚萌发培养基上,体胚萌发率高达100%。将萌发的体胚转移到1/2MS + 1 g ? L-1活性炭成苗培养基中,进一步发育成正常的再生植株,植株室外栽培成活率达90%。

关键词: 红姜花, 胚性细胞悬浮系, 体细胞胚胎发生, 植株再生

Abstract: A reproducible protocol for somatic embryogenesis was established for the ornamental ginger Hedychium coccineum using filaments and anthers. After culture for 120 days,callus was induced on Murashige and Skoog(MS)medium supplemented with 4 mg ? L-1 2,4-D,4 mg ? L-1 NAA,1 mg ? L-1 6-BA,30 g ? L-1 sucrose and 7 g ? L-1 agar,proliferated on MS medium containing 1 mg ? L-1 2,4-D,0.25 mg ? L-1 NAA,0.25 mg ? L-1 6-BA,30 g ? L-1 sucrose and 7 g ? L-1 agar. Light yellow and friable embryogenic callus was obtained. Embryogenic calli were suspended in liquid medium containing MS basal salts,B5 vitamins,100 mg ? L-1 glutamine,230 mg ? L-1 proline,100 mg ? L-1 malt extract,0.02mg ? L-1 NAA,0.02 mg ? L-1 TDZ,0.5–1.0 mg ? L-1 2,4-D and 45 g ? L-1sucrose. After 3 months culture,a homogeneous and stable embryogenic cell suspension(ECS),composed of small cell aggregates,was established. Planting of stable ECS on somatic embryo induction media containing Schenk and Hildebrandt(SH)basal salts,B5 vitamins,100 mg ? L-1 glutamine,230 mg ? L-1 proline,100 mg ? L-1 malt extract,0.25 mg ? L-1 NAA,0–0.20 mg ? L-1 TDZ,45 g ? L-1 sucrose and 7 g ? L-1 agar. White and translucent globular embryos were induced after 10 days culture,and mature somatic embryos were obtained after 20 days culture. The highest induction frequency of 4 500 somatic embryos ? mL-1 PCV ECS(PCV:packed cell volume)was derived from medium with 0.15 mg ? L-1 TDZ. A germination percentage of 100% were observed in germination media,which consisted of SH basal salts,B5 vitamins,0.20 mg ? L-1 IAA and 0.25–1.0 mg ? L-1 6-BA. Regenerated plantlets with normal shoot and root were developed after one month on half strength MS medium with 1 g ? L-1 active charcoal. Well rooted plantlets were successfully acclimatized with a survival rate of 90% and grew vigorously.

Key words: Hedychium coccineum, embryogenic cell suspensions, somatic embryogenesis, plant regeneration

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