白姜花胚性愈伤组织的诱导与植株再生体系的建立

将白姜花(Hedychium coronarium)未成熟花丝接种在MS+4 mg·L~(-1) 2,4-D+4 mg·L~(-1) NAA+1 mg·L~(-1) 6-BA的培养基上,经过180 d的培养,诱导出3种愈伤组织:Ⅰ,浅黄色松散易碎;Ⅱ,白色疏松半透明水渍状;Ⅲ,黄色致密颗粒状。对愈伤组织继代培养基中的2,4-D、NAA和6-BA浓度进行优化,结果表明培养基中含有1 mg·L~(-1) 2,4-D、0.25 mg·L~(-1) NAA、0.25 mg·L~(-1) 6-BA时可获得胚性状态良好的愈伤组织。胚性愈伤组织在MS无机盐+0.25 mg·L~(-1) NAA+0.5 mg·L~(-1) TDZ+维生素B5+100 mg·L~(-1)谷氨酰胺+230 mg·L~(-1)脯氨酸+100 mg·L~(-1)麦芽提取物+45 g·L~(-1)蔗糖的体胚诱导培养基中培养20d后,转移到MS基本培养基上培养30 d,1 g胚性愈伤组织可获得50~60个体胚。成熟体胚在MS+0.2mg·L~(-1) IAA+0.5 mg·L~(-1) 6-BA的萌发培养基上培养20 d时,体胚萌发率为85%。萌发的体胚转移到1/2MS+1 g·L~(-1)活性炭的成苗培养基中可发育成正常植株,植株室外栽培成活率达90%。对100株再生植株的染色体数进行分析,发现3株三倍体(2n=3x=51)、6株四倍体(2n=4x=68)和2株六倍体(2n=6x=102)。

Embryogenic Callus Induction and Plant Regeneration from Hedychium coronarium via Somatic Embryogenesis

A reproducible protocol for somatic embryogenesis was established for the ornamental ginger Hedychium coronarium. Immature filaments were cultured 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. After culture for 180 days，three types of callus were obtained，type Ⅰcallus were light yellow and friable with tightly packed cell contained dense cytoplasm that appeared to be embryogenic；type Ⅱ callus were white with long，highly vacuolated cells that appeared to be non-embryogenic；type Ⅲ callus were yellow with large，less packed cells contained dense cytoplasm that appeared to between embryogenic and non-embryogenic. Medium containing 1 mg · L^-1 2,4-D，0.25 mg · L^-1 NAA，0.25 mg · L^-1 6-BA was suitable for embryogneic callus proliferation. Embryogenic calli were cultured on somatic embryo induction medium containing MS basal salts，vitamin B₅，100 mg · L^-1 glutamine，230 mg · L^-1 proline，100 mg · L^-1 malt extract，0.25 mg · L^-1 NAA，0.5 mg · L^-1 TDZ，45 g · L^-1 sucrose and 7 g · L^-1 agar for 20 days，then transferred on MS medium free hormone for 30 days. About 50–60 somatic embryos were induced from per gram callus. Germination percentage of 85% were observed in germination medium，which consisted of MS basal salts，0.2 mg · L^-1 NAA and 0.5 mg · L^-1 6-BA. Regenerated plantlets with normal shoot and root were developed after 30 d 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. The chromosome number of 100 regenerated plants showed a wide range of variation，89 of them were diploid（2n = 2x = 34），3 were triploid（2n = 3x = 51），6 were tetraploid（2n = 4x = 68）and 2 were hexaploid（2n = 6x = 102）.