Genetic Engineering Approach to Enhance Adventitious Root Formation of Hardwood Cuttings

Abstract

Vegetative propagation by cuttings is critical to the horticultural industry, particularly the ornamental and fruit sections. This article briefly reviews the traditional approaches to improve root formation, specifically with hardwood cuttings. The focus of this review has been placed on the nontraditional strategies, including infection with Agrobacterium rhizogenes and genetic engineering with rol genes, or ro/B gene alone from A. rhizogenes and iaaM gene from auxin biosynthesis genes (iaaM, iaaH) from A. tumefaciens. Recent results with these new strategies have shown great potential in enhancing adventitious root formation in difficult-to-root woody species. The future perspective in understanding adventitious rooting mechanisms and new strategies for improving rooting are also discussed.     

全光雾喷法在刺玫果嫩枝扦插上的应用

全光雾喷法扦插育苗是近几年应用较多的快速育苗技术,它具有生根快、繁殖率高的特点,是快速繁殖优良品种苗木的一条重要途径。笔者对刺玫果分别用普通扦插和全光雾喷法进行夏季嫩枝扦插试验,结果表明用浓度为2×10-5g／mL的IBA浸泡16小时处理的插穗使用全光雾插法生根率可高达96％。     

不同激素种类及浓度对红厚壳嫩枝扦插育苗的影响

以红厚壳（Calophyllum inophyllum）1年生嫩枝为扦插材料,探讨不同浓度梯度的萘乙酸（NAA）、吲哚丁酸（IBA）及生根粉1号（ABT）对插穗抽叶特性和生根特性的影响,并运用隶属函数值法对各处理组合的扦插效果进行综合评价。结果表明：与对照处理相比,不同激素种类及浓度处理显著影响嫩枝插穗的抽叶率、新叶数量、新叶面积等抽叶特性指标,以及生根率、偏根率、不定根数量、不定根体积、侧根数量和侧根长度等生根特性指标,但对宿叶率影响不显著。不同激素处理下嫩枝插穗的抽叶率和生根率分别为21.7%~66.7%和43.3%~98.3%,而对照处理的抽叶率和生根率均为0。基于隶属函数值法综合分析,激素处理对扦插效果的促进作用依次为NAA>ABT>IBA。NAA处理插穗的抽叶率和生根率均值为62.1%和95.4%,分别比ABT处理增加47.5%和30.2%,比IBA处理增加62.0%和57.9%。综合分析,红厚壳嫩枝扦插的最佳处理组合为1000 mg/L NAA,其抽叶率为61.7%,生根率为95.0%。     

澳洲坚果扦插繁殖技术在生产中的应用

在南亚所苗圃对6个澳洲坚果品种和在西双版纳苗圃对9个澳洲坚果品种插穗，分别以WGD-1和IBA、ABT—1号、IAA等生长调节剂处理，进行扦插繁殖比较试验。结果表明：WGD—1、IBA、ABT-1号和IAA的合理使用均有显著促进澳洲坚果插穗生根的作用。WGD-18000mg/L处理的插穗成苗率显著高于8000mg/L IBA及ABT-1号、IAA处理：IBA 8000mg/L处理成苗率显著高于IAA；但IBA与ABT—1号之间，ABT—1号与IAA之间，差异不显著。经2个苗圃3a以上应用WGD—1配方生产扦插苗，成苗率达82．34％；品种344在版纳苗圃成苗率显著高于660、741，而344、246、O．C、788、800、H2之间差异不显著，但788、294扦插苗根系少，扦插难度较大：在常规条件下，我国植区的澳洲坚果扦插繁育期最宜在冬季，以长20～25cm皮色灰白木栓化枝作插穗为佳，带叶扦插，插后40～45 d所有插穗均有愈伤组织形成，生根高峰期在插后115～150 d之间，但品种间有差异。     

顶梢和功能叶对茶树全光照弥雾扦插快繁的影响研究

为进一步缩短茶树无性苗繁育周期,提高无性苗质量,本试验以3年生台茶12号（又名金萱）茶树嫩枝为扦插材料,采用全光照弥雾快繁育苗技术,研究了插穗顶梢保留与摘除、不保留成熟功能叶、保留1~3片成熟功能叶对茶树扦插苗根系和地上部生长的影响,并于扦插前对茶树嫩枝不同部位芽叶的相关生理生化指标进行了测定分析。结果表明,随叶片成熟度增加,可溶性糖和叶绿素含量呈递增趋势。净光合速率也随叶片成熟度的增加呈递增趋势,但以第2片成熟功能叶最高。可溶性蛋白含量随叶片成熟度的增加呈下降趋势,以顶部一芽一叶含量最高。带成熟功能叶插穗成活率和生根率均为100.00%,而不带成熟功能叶的一芽一叶插穗和一芽二叶插穗成活率和生根率均仅为5.00%和23.33%。根系生长以带3片功能叶留梢插穗最好,生根早且根量大,10βd内就有大量白色根点从茎段基部冒出,60%~70%茶苗形成二次根仅需37βd,且地上部新形成的成熟功能叶数也最多,但是新梢高度增长值小于保留顶梢带1片成熟功能叶插穗,基部茎粗增长值小于带2片成熟功能叶插穗。成熟功能叶片数的增加有利于根系生长发育,提高成活率和生根率;保留顶梢插穗的根系和地上部的生长优于去梢插穗。     

茶树插穗发根的解剖学和生物化学研究

通过对浙农12等七个无性系茶树品种扦插发根过程的显微切片观察和主要生化指标的测定,结果表明:茶树红棕色枝和绿色枝的不定根原基分别起源于木栓形成层和中柱鞘内侧的韧皮薄壁细胞。根据插穗基部解剖学和形态学特征的变化,茶树插穗不定根的形成过程可分为5个时期,即:根原基未分化期、根原基分化期、根原体分化前期、根原体分化后期和不定根形成期。用80ppm IBA 喷射母树两周后,个别品种的插穗在离开母树时已分化出不定根原体。插穗母叶内的淀粉和非蛋白氮高,母茎内的碳、氮比高,以及母叶内的蛋白氮含量低的品种,具有较强的发根能力。     

Influence of proprietary rooting compounds and basal internode length on the rooting of potato stem cuttings

Summary Potato stem cuttings, cv. Désitrée, were treated with a powder formulation of 4-indol-3-ylbutyric acid (IBA), a gel formulation of 1-naphthylacetic acid (NAA), or a powder formulation of NAA. Both the IBA powder and the NAA gel stimulated significantly more roots per rooted cutting than were produced by the untreated controls but the NAA gel also significantly reduced the number of root branches per rooted cutting (Table 1). Untreated cuttings of cvs. Désirée and Recond were more likely to form only roots when the basal internode was long and more likely to form only tubers when the basal internode was short (Table 2). We suggest that the number of stem cuttings forming roots would be increased if, where necessary, the cuttings were trimmed to leave a long basal internode.     

辣木育苗试验

采用容器育苗,种子需经特殊催芽处理后播种,幼苗期严防苗木猝倒病。辣木属难生根树种,扦插繁殖生根率低,使用低浓度（100和200mg·L^-1）萘乙酸或吲哚丁酸处理,可提高插穗生根率。     

不同激素种类、浓度及浸泡时间对金蒲桃扦插生根的影响

以金蒲桃半木质化枝条作为插穗,以黄心土+砂（1∶1）为基质,通过正交试验针对不同激素种类、浓度以及浸泡时间等因素研究金蒲桃扦插生根的效果,从而获得最适宜金蒲桃扦插生根的处理,为其扦插繁殖提供理论基础和技术支持。结果表明:金蒲桃以皮层组织生根为主,较好的处理插穗成活率超过90%,激素浓度是影响金蒲桃扦插生根最主要的因素,其次是激素种类,浸泡时间的影响最小。极差分析结果发现,扦插生根率最适合的因素组合A₂B₂C₃（500 mg/L IBA浸泡2 min）或A₂B₃C₁（200 mg/L IBA浸泡30 min）,扦插生根数量最适合的因素组合为A₁B₁C₁（1000 mg/L NAA浸泡30 min）,扦插生根平均根长最适合的因素组合为A₃B₃C₂（200 mg/L“根太阳”生根剂浸泡10 min）,最适合扦插生根最长根长的因素组合为A₂B₁C₂（1000 mg/L IBA浸泡10 min）,最适合扦插生根效果指数的因素组合为A₁B₁C₁（1000 mg/L NAA浸泡30 min）。主成分分析联合生根效果指数指标极差分析结果表明,A₁B₁C₁（1000 mg/L NAA浸泡30 min）和A₁B₂C₂（500 mg/L NAA浸泡10 min）是适宜金蒲桃扦插较好的组合。总之,金蒲桃是一种扦插相对比较容易的植物,该研究结果可为生产中金蒲桃的扦插繁殖提供参考。     

Optimizing the propagation of potato (Solanum tuberosum L.) by stem cuttings

Stem cuttings from potato plants are frequently used in multiplication programs for disease-free cultivars. The survival of these leafy cuttings is often very poor, and thus various techniques were investigated which were designed to increase rooting and cutting survival. Stock plants of cultivars “Caribe,” “Jemseg,” “Keswick,” “Russet Burbank,” “Sable,” “Sebago” and “Shepody” were grown in the greenhouse under long days. Promoting growth of the axillary shoots was successfully achieved by shading the shoot apex with aluminum foil. Trimming the outer leaves of the cuttings was deleterious to both survival and rooting. A rooting hormone improved root initiation and development, and the use of shade cloth or intermittent mist increased survival by reducing wilting. A rooting substrate of vermiculite: perlite (1:1) provided sufficient aeration, support and drainage for the cuttings. Antitranspirants sprayed on the stock plants prior to the excision of cuttings and also on the excised cuttings did not improve rooting or survival. The addition of activated charcoal (1% w:w) increased rooting of the cuttings and improved vigor of potted plants obtained from the cuttings.     

Large scale in vitro propagation of Stevia rebaudiana (bert) for commercial application: Pharmaceutically important and antidiabetic medicinal herb

Stevia rebaudiana is a valuable medicinal plant species and it is being used for the treatment of diabetes. Currently, there is a high demand for raw material of this medicinal herb due to ever increasing diabetes disorder among the population. In order to meet the increased demand an efficient in vitro propagation of S. rebaudiana was established. Nodal explants collected from the field were cultured on MS basal medium fortified with different concentrations of BAP (0.5-3.0 mg/l) and KIN (0.5-3.0 mg/l) individually for shoot bud induction. In vitro derived nodal buds were cultured on MS medium supplemented with different concentrations (0.5-3.0 mg/l) of BAP and KIN for multiple shoot bud regeneration. In the second experiment, in vitro derived buds were placed on MS medium supplemented with different concentrations of BAP (0.5-3.0 mg/l) in combination with 0.5 mg/l IAA or IBA or NAA for shoot bud multiplication. The highest frequency (94.50%) of multiple shoot regeneration with maximum number of shoots (15.69 shoots/explant) was noticed on MS medium supplemented with 1.0 mg/l BAP. For large scale plant production, in vitro derived nodal bud explants were cultured on MS medium fortified with 1.0 mg/l BAP, in which about 123 shoots/explant were obtained after three subcultures on the same media composition. Elongated shoots (>2 cm) dissected out from the in vitro proliferated shoot clumps were cultured on half-strength MS medium containing different concentrations of NAA (0.1-0.5 mg/l) and/or MS medium fortified with various concentrations (0.5-2.0 mg/l) of auxins (NAA, IAA and IBA) for root induction. Highest frequency of rooting (96%) was noticed on half-strength MS medium augmented with 0.4 mg/l NAA. The rooted plantlets were successfully transferred into plastic cups containing sand and soil in the ratio of 1:2 and subsequently established in the greenhouse. The present in vitro propagation protocol would facilitate an alternative method for rapid and large-scale production of this important antidiabetic medicinal plant.     

H．11648麻珠芽组织培养技术研究

用H．11648麻珠芽为材料进行组织培养，结果表明：在MS＋6-BA5mg／L＋NAA0．3mg／L＋白糖3％＋琼脂0．65％培养基中，诱导产生不定芽，其诱导率为72％，不定芽在MS＋6-BA3mg／L＋NAA0．1mg／L＋白糖5％＋琼脂0．8％培养基中，诱导产生丛芽，其诱导倍数是1．57，对丛芽进行继代培养、实现丛芽平均每代的增殖率228％。用MS＋NAA1．0mg／L＋IBA0．3mg／L＋活性炭0．2％＋白糖3％培养基诱导生根，其生根率为82％；将生根苗移至菇渣5：塘泥3：表土1：河沙1配方基质的苗床假植，移栽平均成活率为84％。该项技术的研究成功，为今后剑麻种苗的工厂化生产提供了技术支撑。也为开展剑麻的生物工程育种提供了技术基础。     

Visual inspections of nursery stock fail to protect new plantings from Blueberry scorch virus infection

Blueberry scorch virus (BlScV) is one of the most pervasive pathogens of highbush blueberry. The virus is aphid-vectored and exhibits a latent period between infection and symptom expression in the host plant of up to 5 years. In many cases, we have observed BlScV symptom expression in new fields that appears inconsistent with aphid-vectored introduction and spread. It was, therefore, speculated that the virus may be introduced through infected nursery stock. To examine this possibility, we first surveyed selected nurseries to determine if mother plants, used for propagation by cuttings, were BlScV-infected. Two nurseries were found to harbor symptomless, infected, mother plants (cv. Duke). Cuttings from one nursery were collected from infected and non-infected plants and rooted in propagation beds. The survival and infection of cohorts from each mother plant were determined one year after planting. A significantly greater proportion of cuttings survived from non-infected mother plants (0.7) than from infected mother plants (0.5). Of the cohort from infected mother plants that survived, 40% tested positive for BlScV. We also surveyed the distribution of infected, symptomatic plants in recent ‘Duke’ plantings that originated from nurseries with BlScV-infected mother plants and compared distribution with older plantings with more advanced BlScV outbreaks. In all cases, the distribution of BlScV symptom development in young fields was random, which is consistent with introduction from planting stock. Older plantings showed a strong clustered distribution, which is consistent with aphid transmission. This study identifies infected nursery stock as an important source of BlScV dissemination and underscores the importance of having symptomless mother plants virus tested.     

生根剂处理对于茶树嫩枝扦插内源激素水平和繁殖的影响

为进一步探究茶树不同嫩度的嫩枝插穗根系和地上部生长适宜的生根剂处理时间及内源激素水平,本试验以3年生台茶12号嫩枝为试验材料,采用酶联免疫吸附测定法（ELISA）分析了嫩梢不同部位IAA、ZR、GA₃和ABA含量,同时研究了生根剂处理时间对插穗茎基部激素含量和茶苗扦插快繁的影响。结果表明,在茶树嫩枝顶部一芽一叶至一芽二叶初展梢（BL1）、半功能叶（L2）、第1片功能叶（L3）及其茎段（S1）、第2片功能叶（L4）及其茎段（S2）、第3片功能叶（L5）及其茎段（S3）、第4片功能叶茎段（S4）中,叶片内IAA和ABA以第2片功能叶含量最高,ZR以第3片功能叶含量最高,GA₃含量则随叶片成熟度增加而增大。茎段S1—S3中IAA含量显著高于S4,而ABA含量则为S1显著高于S2—S4。浸泡100βmg·L^-1 ABT-1号生根剂会显著提高插穗茎基部IAA含量,并使IAA/ABA和(IAA+ZR+GA₃)/ABA值增至1.0左右,从而有利于生根。其中,保留顶梢带1片和2片功能叶的插穗（F1-1、F2-1）在浸泡生根剂前期1βh内IAA含量快速增加,且IAA/ABA和(IAA+ZR+GA₃)/ABA值可增至1.0以上,其他插穗则在浸泡生根剂4βh后上述3个指标达到最大值。F1-1、F2-1浸泡生根剂0.5~1βh,摘除顶梢带1片功能叶的插穗（F1-2）浸泡生根剂4βh,F2-2、F3-1、F3-2浸泡生根剂3~4βh,IAA含量达到较高水平,而且IAA/ABA和 (IAA+ZR+GA₃) /ABA值可增至1.0左右。出苗时,平均根系活力、平均生根条数和平均茎粗增加值的最大值出现在浸泡生根剂0~2βh内,其他根系和地上部生长情况以浸泡生根剂4βh表现较好。     

弄岗马兜铃组培苗的生根培养

对弄岗马兜铃组培苗进行生根诱导研究,以达到可移栽的目的。在1/2改良MS固体培养基(KH2PO4 170 mg/L、蔗糖30 g/L,pH 5.8)中添加不同浓度的6-苄胺基腺嘌呤(6-BA)和吲哚丁酸(IBA)筛选最佳生根培养基配方。结果表明：在1/2改良MS固体培养基(蔗糖30 g/L,pH 5.8)加入1 mg/L的IBA具有较好的生根诱导作用,而6-BA对弄岗马兜铃组培苗不具有生根作用。     

Comparison of inoculation methods on ring rot development in potatoes

Symptoms of bacterial ring rot (Corynebacterium sepedonicum) developed earlier and were more severe in root-inoculated potato plants grown from stem cuttings treated with indole-3-butyric acid (IBA) than in those grown from stem cuttings not so treated or from tuber seed pieces. Disease expression was more rapid and severe in root-inoculated plants than in plants from inoculated seed pieces. The use of IBA-treated stem cuttings provided uniform plant material for ring-rot studies.     

Chemical control of Cylindrocladium and Rhizoctonia root rots in blueberry propagation

Cylindrocladium and Rhizoctonia root rots, caused by Cylindrocladium parasiticum and binucleate Rhizoctonia spp., respectively, are common soilborne diseases observed during blueberry propagation in Georgia, USA. Six trials with artificially inoculated propagation medium were completed in a shade-house to determine the efficacy of select fungicides to control these diseases. For Cylindrocladium root rot, two trials with single-drench fungicide applications were made at the time the cuttings (southern highbush blueberry cultivar Rebel) were stuck. In two additional trials, a second drench application was added two weeks later. In all trials, fludioxonil (Medallion) significantly reduced lesion incidence and lesion length. Neither triflumizole (Terraguard) nor thiophanate-methyl (Cleary's 3336) consistently reduced lesion incidence with one application, indicating that a single application of these materials was not sufficient for disease suppression. The second fungicide application improved disease control for all fungicides. However, fludioxonil always performed statistically better than thiophanate-methyl and numerically better than triflumizole as indicated by lesion incidence, lesion length and girdling incidence. For Rhizoctonia root rot, two trials were completed, each with two fungicide applications (when cuttings were stuck and 2 weeks later). Fludioxonil and azoxystrobin (Heritage) significantly reduced lesion incidence and lesion length in both trials. Flutolanil (Prostar) reduced lesion incidence in both trials and also reduced lesion length in the second trial. Pending labeling for this use pattern, fludioxonil, which has provided good control of both diseases in our trials, may form the basis of a fungicide rotation program to supplement sanitation practices for managing Cylindrocladium and Rhizoctonia root rots in blueberry propagation.     

Evaluation of fungicides to control Cylindrocarpon liriodendri and Cylindrocarpon macrodidymum in vitro, and their effect during the rooting phase in the grapevine propagation process

The efficacy of 14 selected fungicides against Cylindrocarpon liriodendri and Cylindrocarpon macrodidymum was evaluated in vitro by testing their effect on mycelial growth and conidial germination. Carbendazim, hydroxyquinoline sulphate, imazalil, and prochloraz were the most effective fungicides in reducing mycelial growth in both Cylindrocarpon species. Captan, copper oxicloride, didecyldimethylammonium chloride and thiram were the most effective to inhibit conidial germination of both species. A pot assay was also conducted with captan, carbendazim, copper oxychloride, didecyldimethylammonium chloride, hydroxyquinoline sulphate, imazalil and prochloraz in order to determine their potential to prevent infections caused by C. liriodendri and C. macrodidymum during the rooting phase in the grapevine propagation process. All fungicides significantly decreased the root disease severity values in both species compared with control treatment, with the exception of imazalil in C. macrodidymum. In the case of the percentage of reisolation, all values were lower than those obtained for the control treatment, but only captan, carbendazim and didecyldimethylammonium chloride were significantly different in the case of the cuttings inoculated with C. liriodendri, and prochloraz in the case of those inoculated with C. macrodidymum.     

芦笋组织培养快繁技术

[目的]探索芦笋的组织培养快繁技术.[方法]利用丰岛芦笋3—4月抽出的嫩茎为外植体,在不同培养基上诱导、增殖和生根.[结果]筛选出芦笋适宜的诱导培养基为MS+1.0 mg/L BA+0.1 mg/L NAA,增殖培养基为MS+0.5 mg/L BA+0.1 mg/L NAA,生根培养基为MS+1.0 mg/L IBA+...     

邓恩桉微扦插技术

通过对邓恩桉不同季节、不同种源的穗条采用不同激素处理进行微扦插生根试验，结果表明，3个因素对其生根率的影响表现为：种源〉扦插季节〉激素；不同种源的影响依次为：澳大利亚昆士兰州〉澳大利亚新南威尔士〉巴西柏拉拿州：不同扦插季节的影响表现为：夏季〉春季〉秋季；不同激素的影响表现为：GGR〉IBA〉ABT1^#，但3种激素对邓恩桉微扦插生根率的影响差异不显著。春季澳大利亚昆士兰州种源的母树采集的邓恩桉穗条，经4mg·kg^-1ABT1^#。生根粉或GGR或IBA与滑石粉调成糊状，速蘸后扦插生根率最高，达73．55％。