Adventitious shoot regeneration from in vitro leaves of adult Camellia reticulata

Experiments were conducted with Camellia reticulata cv. Captain Rawes to develop an adventitious regeneration system. Leaves were taken from axillary shoot proliferation cultures in WPM medium that had been established from mature trees. They were sectioned, and then plated in Petri dishes on media containing various combinations of cyto- kinins and auxins; the best response was induced by 2 mg I“1 BA + 1 mg I-1 IBA. The shoots obtained were multiplied by axillary branching on WPM + 2 mg I-1 BA + 2 mg I-1 Z + 2 mg I-12iP + 0.01 mg I-1 IBA. There was no significant difference in multiplication coefficient (the product of the proportion of explants forming axillary shoots and the mean number of new segments per explant) between shoots of adventitious and axillary origin, but there was between the various types of explant used in the multiplication stage: shoot tips (ST1) and nodal segments (ns) of harvested shoots longer than 14- 15 mm, and whole harvested shoots 7-10 mm long (ST2). The best results were achieved with ns and ST2 explants. Shoots of adventitious origin rooted very poorly in comparison with those of axillary origin under the same culture conditions.     

The effects of explant and cytokinin type on regeneration of Prunus microcarpa

We investigated in vitro regeneration ability of Prunus microcarpa subsp. tortusa using various explants (root, cotyledon and hypocotyl pieces) and cytokinins [benzyladenine (BA), meta-Topolin (mT) and thidiazuron (TDZ)]. Sectioned cotyledon, root and hypocotyl pieces of in vitro grown seedlings were cultured on Nas and Read Medium (NRM) containing BA (7.5, 10, 12.5, 15 or 17.5 μM), mT (2.5, 5.0, 7.5, 10 or 12.5 μM) or TDZ (2.5, 5.0, 7.5, 10 or 12.5 μM). As a measurement of morphogenetic reaction, the ratios of regenerating explants and the numbers of primary adventitious shoots per regenerating explant were analyzed. Cotyledon explants exhibited higher regeneration ratios than hypocotyl explants, and the root explants were inappropriate for regeneration. Both BA and mT were effective on shoot regeneration but higher regenerating explant ratios were obtained when BA was used. In comparison with BA and mT, the effect of TDZ on enhancing explant regeneration ability was insignificant. Mean number of adventitious shoot per regenerating explant was between 1 and 4, and regenerating explant ratios were between 0% and 77%. The practical appliacations of the results are discussed.     

‘艾西丝’南瓜子叶的离体培养

用‘艾西丝’南瓜子叶作外植体离体培养,成功地建立了一套子叶高频率诱导再生芽的程序。在ＭＳ＋ＢＡ ４．０ｍｇ／Ｌ＋ＩＡＡ０．４ｍｇ／Ｌ的培养基上,以４ｄ苗龄子叶基部切块为外植体,其不定芽的诱导率最高,达５０％,在ＭＳ＋ＢＡ０．２５－０．５ｍｇ／Ｌ的培养基上继代,增殖培养效果较好,在无激素的１／２ＭＳ培养基上最易生根。     

欧洲甜樱桃幼胚子叶离体培养再生植株研究

 以欧洲甜樱桃（Prunus avium L.）幼胚子叶为试材,研究了子叶不同发育时期、BA和NAA配比、品种基因型和培养条件等对幼胚子叶离体再生不定芽的影响。结果表明,以MS为基本培养基,附加BA 2.0 mg·L^-1＋NAA 0.2 mg·L^-1,用PF（子叶的长度/胚的长度×100）﹦50～80发育阶段的子叶,那翁幼胚子叶再生不定芽效果最好,再生率最高可达79.2﹪。暗培养2周后再转入光照下培养,对欧洲甜樱桃子叶再生不定芽具有一定的促进作用。欧洲甜樱桃其他品种的子叶再生率,雷尼尔为72.2﹪,先锋为61.1﹪,拉宾斯为33.3﹪。子叶再生不定芽全部发生于子叶的正面近胚芽端切口处,表现出明显的极性效应。     

Direct somatic embryogenesis and adventitious shoot formation from immature axillary buds of Melastoma affine

Summary

Direct somatic embryogenesis and adventitious shoot formation were induced from immature axillary bud explants of Melastoma affine cultured on induction medium containing both naphthaleneacetic acid (NAA) and cytokinins. Among the cytokinins tested, thidiazuron (TDZ) played a greater role in the induction of somatic embryogenesis than 6-benzylaminopurine (BA) and kinetin (KT). Histological studies of paraffin sections showed that the same tissue from immature axillary buds produced different types of explants that developed floral primordia and vegetative bud primordia during the earlier and later flowering stages, respectively. These could result in different developmental pathways. Efficient mass propagation and plant regeneration systems were established for Melastoma affine.     

兰州百合花瓣高频再生体系研究

以兰州百合为试材,研究总结出了利用花瓣建立外植体的高频再生体系:用花瓣建立外植体时,灭菌时间控制在15min效果较佳,污染率、失活率均低;适宜诱导花瓣愈伤组织的培养基为MS BA0.5mg/L 2,4-D 0.5mg/L,诱导率达到90%;适宜芽分化培养基为MS BA1.5mg/L NAA0.1mg/L,分化率为75%;适宜继代和增殖的培养基为MS BA1.5mg/L IBA0.5mg/L,增殖倍数可达5.8;生根培养基以GS NAA0.25mg/L最佳,生根率高达100%,试管苗生长势强,且根系发达;用草炭作移栽基质,成活率高,为89.6%.     

Regeneration of plants from Fraxinus pennsylvanica hypocotyls and cotyledons

An adventitious shoot regeneration and rooting protocol was developed for green ash (Fraxinus pennsylvanica) seedling explants. The best regeneration medium for freshly isolated hypocotyls and cotyledons was Murashige and Skoog (MS) supplemented with 13.3 μM 6-benzylaminopurine (BA) plus 4.5 μM thidiazuron (TDZ), and 22.2 μM BA plus 4.5 μM TDZ, respectively. Seventy-six percent of hypocotyl segments and 24% of cotyledon segments produced adventitious shoots, with a mean number of adventitious shoots per explant of 2.7 ± 0.5 and 2.3 ± 1.3, respectively. The effect of in vitro-germinated seedling age on adventitious shoot regeneration from hypocotyl and cotyledon explants was also studied. Results showed that hypocotyl and cotyledon explants from freshly isolated embryos exhibited a higher organogenesis potential than 4–15-day-old explants. Adventitious shoots from hypocotyls and cotyledons were established as proliferating shoot cultures following transfer to MS basal medium with Gamborg B5 vitamins supplemented with 10 μM BA plus 10 μM TDZ. A high rooting percentage (73–90%) was achieved when in vitro shoots were rooted on woody plant medium (WPM) containing 4.9 μM indole-3-butyric acid (IBA) and IAA (0, 2.9, 5.7, or 8.6 μM) with a combination of 10-day dark culture period followed by a 16-h photoperiod. The highest rooting (90%) of adventitious shoots or the number of roots per shoot (3.0 ± 1.0) was obtained on WPM with 4.9 μM IBA plus 5.7 μM IAA. Rooted plants were successfully acclimatized to the greenhouse and 100% survived after overwintering in cold storage. This regeneration system using hypocotyls and cotyledons provides a foundation for Agrobacterium-mediated genetic transformation of F. pennsylvanica for resistance to the emerald ash borer.     

Factors affecting in vitro adventitious shoot formation on internode explants of Citrus aurantium L. cv. Brazilian

The in vitro formation of newly formed adventitious buds and shoots from internodal branch segments was studied on 12-month-old plants of Citrus aurantium L. cv. Brazilian. The effects of 6-Benzyladenine (BA) and α-Naphthalene acetic acid (NAA) treatments were evaluated on adventitious bud and shoot regeneration. High rates of bud initiation and shoot development were obtained both with BA supplemented medium, in the range from 1 mg L⁻¹ to 3 mg L⁻¹, and with 0.1 mg L⁻¹ NAA supplemented medium. NAA concentrations above 1 mg L⁻¹ significantly reduced bud initiation and shoot elongation. The results obtained using different in vitro culture vessels such as Petri dishes, tubes and glass culture jars were compared. The highest adventitious bud induction was observed in Petri dishes for internodes cultured in 2 mg L⁻¹ BA supplemented medium, with 95% responsive explants forming 9.0 ± 2.4 adventitious buds. The adventitious buds observed in Petri dishes reached a maximum height of 1 mm, with no further development, while some of the adventitious shoots cultured in tubes and glass culture jars grew over 1 cm in height. A shoot regeneration gradient of the internodes collected along the branch axis was noticed, with basal ones exhibiting higher regeneration frequency.     

Comparison of shoot induction ability of different explants in herbaceous peony (Paeonia lactiflora Pall.)

Shoot induction ability of explants of herbaceous peony was investigated in semisolid MS medium containing BA, TDZ and GA₃. Callus was readily induced from stem without node and petiole explants within 2 days of culture but failed to generate shoots. Adventitious shoots were successfully produced from meristematic regions only: bud eyes on nodal stem sections, and junctions of petioles and petiolules. No shoots were induced from internode sections, petiole without junctions, or leaf sections. Nodal sections were the most efficient explants. There were up to 20 shoots in one explant generated within 20 days of culture. TDZ was more effective than BA to induce shoots. The 100% shoot induction rate was obtained in medium containing 0.1–3 mg L⁻¹ of TDZ. However, higher concentrations of TDZ inhibited shoot elongation and only large leaf clusters were produced. Combinations of BA and TDZ failed to increase shoot induction rates but caused shoots shorter. The 2–60-min pretreatment of explants with 20 mg L⁻¹ TDZ solution was very effective to induce adventitious shoots directly, but both shoot number and shoot length tended to decrease as treatment time increased. GA₃ was beneficial for shoot and stem elongation.     

欧李离体叶片再生体系的建立

【目的】以欧李试管苗叶片为外植体进行不定芽再生研究,以期为核果类果树的品种改良、基因遗传转化和功能验证等生物技术育种奠定一定的基础,【方法】使用不同的基本培养基和激素组合促进其不定芽再生并建立再生体系。【结果】结果表明,欧李幼芽增殖对基本培养基类型较敏感,改良MS培养基优于MS、F14和WPM。在MS(改良)+NAA 0.1 mg.L-1+BA 0.2 mg.L-1培养基上,增殖系数为5.6,增殖效果较好。离体叶片不定芽再生的最适植物生长调节剂组合为MS(改良)+TDZ 4.0 mg.L-1+IBA 0.2 mg.L-1,再生率达到63.1%,平均每外植体再生芽数为4.9。暗培养10 d可以提高不定芽再生率。再生不定芽1/2 MS(改良)+IBA3.0 mg.L-1培养基上生根率最高,生根率达93.3%。【结论】在改良MS培养基上TDZ与IBA组合诱导不定芽再生的效果最好。     

Micropropagation of fastigiate bird cherry (Prunus padus L.) and adventitious shoot formation from leaves

A shoot tip of a mature clone of fastigiate bird cherry (Prunus padus L.) was successfully established in vitro. Culture of shoot tip explants on a Murashige and Skoog (MS) based medium with phloroglucinol (PG) resulted in micropropagation, but the clonal line gradually became hyperhydric on this medium. This problem was overcome using PG- free medium based on either MS or Driver and Kuniyuki Walnut medium (DKW). Heavier cultures with more shoots were obtained on DKW medium with fructose or glucose rather than sucrose or sorbitol. Leaf explants placed on DKW basal medium with benzyladenine (BA) produced adventitious shoots. Addition of 1-naphthaleneacetic acid to media with BA increased regeneration. More leaves produced shoots on medium with sucrose or sorbitol rather than glucose or fructose. Adventitious shoots were excised and micropropagated. Shoots were rooted by insertion into DKW medium with indol-3yl- butyric acid, followed by transfer to hormone-free DKW. PG increased the proportion of shoots that produced adventitious roots.     

野生水芹组培快繁技术研究

以野生水芹茎尖、腋芽为外植体,研究了影响愈伤组织诱导和分化的激素浓度配比.结果表明,最佳外植体是茎尖;最佳愈伤组织和不定芽诱导培养基为MS 6-BA 4 mg/L,最佳继代增殖培养基为MS 6-BA 2 mg/L,最佳生根培养基为MS IBA 1 mg/L.并提出试管苗炼苗与移栽的最佳试验条件.     

山杜英离体培养植株再生的研究

 研究了山杜英[Elaeocarpus sylvestris(Lour．)Poir．]带芽茎段的离体培养及植株再生。筛选出最佳培养基：(1)启动培养：Ms+BA 1.0～2.0 mg·L^-1 (单位下同)+NAA 0.05+蔗糖3%；(2)愈伤组织发生型丛生苗增殖培养：MS+BA 1.0+NAA 0.5+蔗糖3%；腋芽发生型丛生苗增殖培养：MS+BA2.0+IBA 0.1+蔗糖3%；(3)有效苗诱导培养：MS+BA 0.5+IBA 0.1+GA 1.0+蔗糖3%；(4)生根培养：1/2 MS+IBA 3.0+蔗糖2%。用透气膜封FI比用聚乙烯菌膜生根率明显提高，生根明显提前。     

高品质甜叶菊品系“1096”的组培快繁

采用改良DNS等方法筛选出的生长快、糖苷含量高的甜叶菊品系＂1096＂为材料,研究了不同外植体、不同激素种类和配比对愈伤组织诱导、不定芽分化和生根的影响。结果表明：以茎尖为外植体愈伤组织和不定芽诱导率均最高;其次是带腋芽茎段;再次为叶片。无腋芽茎段和无菌根均能诱导出愈伤组织,但不能诱导成芽。茎尖不定芽诱导的最佳培养基为MS＋6-BA 1.0mg/L;叶片不定芽诱导的最佳培养基为MS＋6-BA 1.0mg/L＋IAA 1.0mg/L;带腋芽茎段不定芽诱导的最佳培养基为MS＋6-BA 1.5mg/L。不定芽继代增殖培养采用MS＋6-BA 0.5mg/L＋NAA0.05mg/L;在1/4MS＋IBA 0.1mg/L＋NAA 0.1mg/L＋1g/L活性炭的培养基上诱导生根,生根率可达100%,生根苗移栽后成活率达95%。     

红肉苹果组织培养及转基因体系的建立与优化

于早春取红肉苹果植株上当年生茎段,建立其组织培养体系,并对其组织培养体系进行优化.结果表明:在添加0.2 mg/L IBA+-1.0 mg/L TDZ的MS培养基上红肉苹果叶盘不定芽诱导率最高,再生率为100％,平均每个叶盘再生9.57个不定芽;1 mg/L IBA+I mg/L BAP组合有利于植株扩繁,其扩繁率为8.27;添加0.6mg/L IBA的培养基上的生根率最高,为93.3％.转基因测试结果表明,农杆菌菌株EHA105适于红肉苹果的基因转化;农杆菌浸染后进行3d的共培养有利于转化效率的提高.     

桃离体组织分化再生植株的研究

  以奉化‘玉露’桃叶片、幼茎、下胚轴、子房、胚珠、胚乳及花后45～50 d 和75 d 的未成熟胚、完全成熟的种胚为外植体, 进行离体培养再分化试验。结果表明只有花后45～50 d 和75 d 的未成熟胚产生白色致密节球状的愈伤组织, 并可诱导出不定芽, 不定芽分化率分别为73. 8 %和15. 5 %; 62BA 诱导愈伤组织不定芽的效果优于KT。不定梢转移生根成苗获得了完整的再生植株。探讨了不同激素和碳源等对不定芽诱导的影响。     

黑莓试管苗叶片植株的再生

以黑莓带芽茎段试管苗叶片为外植体,诱导形成不定芽并进一步形成再生植株。在MS+TDZ2.0mg/L的培养基中叶片不定芽最高分化频率达93.7%(6.52不定芽/外植体),在不定芽伸长培养基BA0.5mg/L上,分化完全的不定芽能够长大伸长,当其高度达到4cm时,切下转接于IBA为0.3mg/L培养基上诱导生根,生根率100%。     

朝鲜白头翁的组织培养与快繁技术

以朝鲜白头翁的叶片、叶柄和花梗为材料,研究了细胞分裂素zeatin、kinetin、6-BA 和生长素IAA 组合对诱导不定芽的影响;以不定芽为材料研究了植物生长调节剂对不定芽的增殖和生根的影响;研究了不同基质配比对组培苗移栽成活的影响。结果表明: 1）叶柄和花梗接种在MS + 0.5 mg · L^-1 zeatin +0.05 mg · L^-1 IAA 培养基培养6 周后不定芽再生率为100%。2）不定芽切成小块或单株带类似愈伤的组织块转到MS + 0.5 mg · L^-1 zeatin + 0.05 mg · L^-1 IAA 或MS + 1.0 mg · L^-1 6-BA + 0.05 mg · L^-1 IAA 的培养基时不定芽的增殖与生长良好。 3）在增殖培养基中长至3 cm 以上的小植株转入到1/2 MS + 1.5 mg · L^-1NAA的生根培养基中培养42 d 后,生根率高达93%,平均根长12.3 cm,每株平均根数11 条,且可缩短生根时间。4）组培苗移栽到东北山土和珍珠岩比例为2︰1 的混合基质时,成活率高达66.7%。     

金花梨子叶不定梢再生研究

以金花梨花后65 d的成熟胚子叶为外植体,研究了不同培养基、BA浓度、暗培养条件及AgNO3浓度对子叶不定梢再生的影响。结果表明:梨成熟胚子叶在适宜培养条件下能发生大量不定梢,培养15 d为初发生期,25~30d为高峰期,40 d后结束发生。暗培养预处理有利于提高子叶再生频率、再生不定梢数和不定梢质量。在NN69+BA3.0mg/L+NAA 0.2mg／L培养基上,暗培养3周后转光下培养获得了85.2％不定梢再生率,平均再生不定梢4.4个。培养基中加入0~4mg／LAgNO3对梨子叶不定梢再生有促进作用,但浓度大于4mg/L时抑制子叶不定梢再生。     

香石竹快速繁殖技术研究

以中花散枝香石竹"红芭芭拉"的茎段为外植体,探讨不同培养基对香石竹的诱芽萌动、继代增殖和生根培养的影响。结果表明:外植体切取部位的诱导芽萌动率从高至低排序为:顶芽带腋芽的茎段不带腋芽的茎段;当培养基为MS+6-BA 1.0mg/L时,对香石竹带腋芽的茎段芽萌动最好;在培养基为1/2MS+6-BA 1.0mg/L+NAA 0.2mg/L时,继代增殖长芽的效果最好;在培养基为1/2MS+6-BA 0.2mg/L+NAA 0.5mg/L时,诱导不定根效果最好。