【作者】 刘晓娜；

【导师】 马俊莲；

【作者基本信息】 河北农业大学 ， 农产品加工及贮藏工程， 2004， 硕士

【摘要】 研究了甜柿上西早生品种(Diospyros kaki Thunb．cv．Unishiwase)的组培快繁、生根和植株再生的基本规律，以期为利用转基因技术改良柿果贮运品质奠定基础，同时也为上西早生柿苗木的快速繁殖提供技术指导。试验结果如下：1、上西早生柿组培苗在MS(1／2N)+ZT1.0+IAA0.1mg／L中继代培养14～15代后，转入DKW培养基中能有效提高丛生苗率约60％，转入MS培养基中培养，平均苗高显著增加。在MS+ZT 1.0+IAA 0.1 mg／L培养基中的组培苗高度和增殖系数，与在MS+ZT 0.5+BA 1.0+IAA 0.1 mg／L中培养的组培苗差异不显著，ZT用量的减少显著的降低了组培苗的成本。0.1～0.2 mg／L IAA比IBA或NAA更适于上西早生柿组培苗的增殖生长。 2、经三因素三水平正交实验，组培苗叶片在MS(1／2N)+ZT 4.0+IAA 0.1mg／E培养基中分化率86％，平均不定芽数2.21。MS(1／2N)培养基最好，其次是1／2 MS，MS效果最差；ZT浓度为4.0 mg／L时不定芽的分化率明显高于2.0mg,／L，在ZT1.0mg／L中培养的叶片分化率最低，IAA对叶片分化的效果不明显；组培苗叶片在MS(1／2N)+ZT 1.0+NAA 0.1mg／L中诱导出绿色颗粒状愈伤组织，在MS(1／2N)+ZT 2.0+IAA 0.1mg／L培养基中，经过二次诱导，愈伤组织分化率75％，平均不定芽数2.5；而2，4-D诱导的愈伤组织不能分化不定芽。 经三因素三水平正交试验，组培苗叶片在TDZ 1.0+ZT 1.0+IAA 0.1mg／L的MS(1／2N)培养基中，分化率95％，分化不定芽数1.95。TDZ的浓度对分化率和平均不定芽数的影响最大，1.0 mg／L显著优于2.0，3.0mg／L。将TDZ 1.0 mg／L诱导的组培苗叶片愈伤组织转入含有ZT 2.0mg／L的MS(1／2N)培养基中，分化率为100％，平均不定芽数为9.5。 生根组培苗的根段在MS(1／2N)培养基中分化率显著高于MS，ZT 2.0mg／L优于ZT 1.0+BA 1.0 mg／L，在MS(1／2N)+ZT 2.0+IAA 0.1mg／L培养基中，40～50mm根段分化率100％，平均不定芽3.84；根段长度与与平均不定芽之间线形回归关系显著，回归方程为Y=0.03671+0.0896X；培养基中加入0.1mg／L的IAA，能显著提高根段的分化率和平均不定芽数。 3、采用浸蘸和直接加入激素两种方法诱导20～25mm高的组培苗生根。IBA诱导生根的效果优于IAA，浸蘸250mg／L的IBA溶液，在1／2MS(1／2N)培养基中，生根率66.67%，平均根条数1.7，或在培养基中添加1.0m留L的田A溶液，生根率64.58%，平均根条数L6;添加工BAO.5+认ALOm叭或IBAI.om留L的生根培养基中，在BA+ZT一中培养的组培苗比在ZT中培养的组培苗生根率提高12.4%，平均根条数提高92.2%，生根苗的腋芽萌动率提高了2.5倍。再生苗生根能力高于丛生苗，依次为根段再生苗>叶片再生苗>BA+zT丛生苗>Zr丛生苗，根段再生苗生根率差异显著。随着继代次数的增加根段再生苗的生根能力逐渐降低，5次继代后的生根能力降至继代丛生苗相同。更多还原

【Abstract】 Diospyros kaki Thunb. cv. Uenishiwase is one of Japanese persimmon varieties. Dormant buds taken from Uenishiwase persimmon trees, as in vitro explants materials, were established successfully. New shoots propagation, shoots rooting and plant regeneration were studied in different basal medium added different hormones. All of the researches in the paper could be useful for modified gene persimmon, through Agrobacterium-mediated transforming, and for fast-propagation of persimmon plants as well. Results as the following:1 Disinfected dormant buds from Uenishiwase-were established in modified MS medium supplemented with ZT 1.0 mg/L and IB A 0.1 mg/L; Micropropagation shoots were subcultured in MS(1/2N) medium added ZT 1.0 +IAA 0.1 mg/L for about 14-15 times, then shoots were transferred to MS medium added ZT 1.0 + IAA 0.1 mg/L or ZT 0.5+BA 1.0+IAA 0.1 mg/L. Shoots grew much better in MS than MS(1/2N) in shoots height and multiple value; and also there was no difference for shoots grown in MS medium adding ZT 1.0mg/L or ZT0.5+BA1.0mg/L. Mere clustering shoots grew in DKW medium than MS medium. The auxin IAA 0.1-0.2 mg/L was superior to IBA or NAA for propagation of Uenishiwase.2 Effects of basal mediums, hormones and their concentrations on shoots regeneration from leaves of Uenishiwase were studied by orthogonal design trial. The results showed that MS (1/2N) was the most optimum for the regeneration and 1/2 MS was better than MS. New shoots differentiation percentage in the medium containing 4.0 mg/L ZT was much higher than 2.0 mg/L and 1.0 mg /L ZT. Differentiated shoots and average shoots per explants were dramatically decreased in the 0.2 mg/L IAA. In the orthogonal trial the best result was achieved in MS (1/2N) medium containing 4.0 mg/LZT and 1.0 mg/L IAA, in which shoot percentage and average shoots per explants were 86% and 2.21 respectively. Green, loose and nodular callus was differentiated from leaves of Uenishiwase in MS(1/2N) medium supplemented with 0.1-0.2 mg/L NAA and 1.0 mg/L ZT; and adventitious shoots were induced on these callus in MS(1/2N) medium supplemented with ZT 2.0 +IAA0.1 mg/L, differentiated percentage was about 75% and average adventitious shoots per calluswas 2.5. Callus of leave induced by 2,4-D could not differentiate shoots.Callus was induced on leaves of Uenishiwase by orthogonal design trial. One hundred percent of leaves differentiated callus in MS(1/2N) medium added TDZ 1.0 + ZT 1.0+ IAA 0.1 mg/L; ninety-five percent callus differentiated adventitious shoots, and average shoots was about 1.95. Callus induced on leaves of Uenishiwase in TDZ 1.0 mg/L was transferred to MS(1/2N) medium added ZT 2.0 + IAA 0.1 mg/L, then number of differentiated shoots were about 9.5, differentiated shoots percentage was 100%.New shoots were differentiated from roots of Uenishiwase shoots much better in MS(1/2N) medium than MS; ZT 2.0 mg/L added in medium was superior to ZT 1.0 + BA 1.0 mg/L. One hundred percent of roots in 40~50mm length could differentiate new shoots, and number of shoots was 3.84. Linear regression relation was distinctive between roots length and no. of new shoots from roots; regression equation was Y=0.03671+0.0896X. And added 0.1 mg/LIAA in MS(1/2N) medium could increase number of new shoots from roots distinctively.3 Roots from Uenishiwase shoots formed by means of dipping auxin solution or adding auxins into 1/2MS(1/2N) medium. IBA was more effective than IAA: after shoots in 20~25mm length were dipped into 250 mg/L IBA solution for 15 min, 66.67 percent rooted, and number of roots was 1.7; 64.58 percent shoots rooted, cultured in medium containing IBA 1.0 mg/L, number of roots was 1.6. Shoots cultured hi MS medium containing BA1.0+ZT0.5mg/L showed higher ability for rooting than containing ZT 1.0mm/L. Regeneration shoots from roots were easier for rooting than regeneration shoots from leaves, subculture shoots; high rooting ability in sequence was regeneration shoots from roots, regeneration shoots from leave, subculture shoots containing BA+ZT, subculture shoots更多还原