桃叶片再生不定芽的研究

以桃栽培品种曙光、金童5号和甜桃王试管苗叶片为外植体,研究了基本培养基、植物生长调节剂种类及质量浓度组合、基因型和试管苗继代次数等因素对叶片再生的影响。结果表明,适宜暗培养的培养基为LP+1.5mg·L-1TDZ+0.15mg·L-1NAA;光培养基为LP+0.5mg·L-1TDZ+0.3mg·L-1KT+0.3mg·L-1NAA和LP+0.6mg·L-1TDZ+0.3mg·L-1KT+0.4mg·L-12,4-D;金童5号和曙光具有较强的再生能力,再生率分别为21.8%和14.5%;曙光和金童5号试管苗继代第1～11次叶片的再生能力与继代前3次愈伤组织相比形成率较高;曙光再生苗在培养基1/2MS+0.5mg·L-1NAA上生根率为100%,平均生根条数为7.9。     

无花果的引种表现及栽培技术

引进A212、A1213、A132、A134、B110和日本紫果无花果优良品种,经4年栽培观察看出,被引品种早果、丰产、适应性强。定植当年结果,A132和A212的4年生树666.7m2产量分别达1259kg和1032kg,果实品质极佳。6个品种均适宜在临沂推广种植。冬季防冻害是丰产稳产的关键,宜采用低干矮冠的杯状或丛状树形,通过修剪控制树高,冬季及时包草、埋土防寒。     

满园香梨叶片不定芽的诱导

采用正交试验设计研究了不同基本培养基、TDZ、生长素种类及其浓度4个因素组合对满园香梨(Pyrusus鄄suriensisMaxim.cv.Manyuanxiang)离体叶片不定芽再生的影响。结果表明,基本培养基的种类是影响叶片不定芽再生最重要的因素,其次是生长素的种类。通过叶片不定芽再生频率和平均每叶再生不定芽数分析表明,NN69+TDZ3.0mg/L+IAA0.2mg/L为满园香梨叶片不定芽发生的最佳培养基组合,在该培养基组合上,满园香梨叶片再生率和平均再生芽数分别达到了74.8%和4.40。同一叶片的基部和顶端比中部更有利于不定芽的再生。     

Suitability of isozyme,RAPD and AFLP markers to assess genetic differences and relatedness among fig (Ficus carica L.) clones

Sarilop is the main and standard cultivar for commercial dried fig (Ficus carica L.) production in Turkey. Eleven of the most promising Sarilop clones and one clone of Sarizeybek, all selected from a former agronomic evaluation, were analysed by three molecular marker techniques, isozymes, RAPDs and AFLPs. The resolution power and the accuracy of these three analytical techniques, in distinguishing among fig clones, were determined. The analysis of five isozyme systems permitted the discrimination between the two cultivars, Sarilop and Sarizeybek. Besides the discrimination between the two fig cultivars, the use of 31 10-mer primers in RAPD analysis allowed splitting the 11 Sarilop clones into two groups of genetic similarity, but not to distinguish between all the clones. The AFLP™ technology showed a much higher multiplex ratio than the RAPD technique (42.4 vs. 6.1) and eight combinations of EcoRI/MseI primers were enough to clearly distinguish between all the Sarilop clones.     

Factors controlling high efficiency adventitious bud formation and plant regeneration from in vitro leaf explants of roses (Rosa hybrida L.)

Studies were conducted to improve adventitious bud regeneration in roses (Rosa hybrida L.), specifically to extend the protocol to different genotypes and to initiate production of multiple shoots per explant. The best results were obtained by using a two-stage procedure where excised leaflets were incubated on Murashige and Skoog (MS) (1962) induction medium with 6.8 μM TDZ plus 0.49 μM IBA in the dark for 7 days and subsequently transferred to an MS-based regeneration medium with 2.22 μM BA plus 0.049 μM IBA exposed to a PPFD of 15 μmol m⁻² s⁻¹ PAR. Bud formation capacity was also significantly affected by the genotype and the environment, such as the use of bottom cooling creating a lower RH in the vessel. The addition of silver nitrate to the induction medium also significantly improved the percentage of regeneration in three genotypes tested. Regenerated shoots failed to elongate when transferred to MS proliferation medium containing 0.5 mg l⁻¹ BA, however maximum bud development and elongation were achieved when kinetin in the range 1–2 mg l⁻¹ was used. Elongated shoots were excised and rooted best on zero growth regulator half-strength MS modified medium. Rooted plantlets were acclimatized under greenhouse conditions for evaluation of somaclonal variation.     

甜樱桃品种吉列玛叶片再生不定梢的研究

在基本培养基NN69或WPM上附加BA和低浓度的NAA，IAA，IBA作培养基，诱导甜樱桃品种吉列玛试管苗的叶片产生了不定梢。以WPM附加BA 3mg/L和IAA0．3mg/L的处理，不定梢再生率最高为33．3％。     

Biodiversity of figs (Ficus carica l.) in Coruh valley of Turkey

The Coruh valley lies within the Caucasus ecological zone, which is considered by the World Wild Fund for Nature and by Conservation International as a one of the biodiversity hotspot in the world. The valley is also recognized by Turkish conservation organizations as an important plant area, an important bird area, a key biodiversity area and has been nominated as a high priority area for protection. This valley is rich in plants and contains 104 nationally threatened plant species of which 67 are endemic to Turkey. Fig (Ficus carica L.) is one of the most important wild edible fruit tree along with pomegranate and olive in the valley. Figs have been used for fresh consumption for centuries and also an important element of natural landscape of the valley. The valley abundantly has black, purple and yellow-green fruited fig trees. In this study we determined some important tree, leaf and fruit morphological characteristics of 50 selected wild grown fig accessions naturally found in the valley. The results showed that the investigated morphological traits of fig accessions displayed significant differences each other. Principal coordinate analysis showed that diversity among the accessions was high and the accessions had black and yellow-green colored fruits were the most diverse groups.     

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.     

无花果细胞系耐盐性与抗氧化酶活性的变化

经＾６０Ｃo－γ射线辐射后再由羟脯氨酸筛选出的无花果抗羟脯氨酸细胞ＡＨ具有更强的 耐盐力。将这些细胞进一步分成不同的细胞系,培养在含有ＮaＣ１１６７mmol/Ｌ的ＭＳ培养基上,７d后,不同细胞间存活率和相对生长量表现出一定差异,最高的是ＡＨ２２和ＡＨ３３,而ＡＨ２细胞几乎全部死亡。盐胁迫诱导绝大多数细胞ＳＯＤ、ＰＯＸ、ＣＡＴ和ＡＰＸ活性增加,而且盐分对细胞生长的抑制程度与ＰＯＸ、ＣＡＴ和ＡＰＸ活性增加倍数     

砂梨叶片再生不定梢的研究

研究了3个砂梨栽培品种翠冠、)青、西子绿离体叶片不定梢再生能力。翠冠再生能力最强,)青其次,而西子绿未能再生。一定时期的暗培养是翠冠叶片再生的必要条件。对翠冠再生过程不同时期的叶片样品进行扫描电子显微镜观察表明,在试验开始的10d后就出现了类分生组织结构,在2个月后观察到充分发育的梢,且最终形成的梢的数量远少于所形成的分生组织数量。     

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.     

Adventitious shoot regeneration and Agrobacterium tumefaciens-mediated transformation of leaf explants of sweet cherry (Prunus avium L.)

Sweet cherry (Prunus avium L.) remains recalcitrant for genetic transformation due to the lack of efficient plant regeneration systems via organogenesis or somatic embryogenesis. In this study, in vitro shoot cultures were derived from a single mature embryo (open pollinated) of ‘Selah’ sweet cherry. Leaf explants were cultured on Woody Plant Medium supplemented with different plant growth regulators to induce shoot regeneration. The optimal regeneration at a frequency of 32.5% and an average of 1.1 shoots per explant occurred on the medium containing 4.54 µM thidiazuron (TDZ) and 2.95 µM indole-3-butyric acid (IBA). Transient transformation showed an efficient delivery of the β-glucuronidase (GUS) reporter gene (gusA) using Agrobacterium tumefaciens strain EHA105. Under the optimal gene delivery conditions, stable transformations were conducted using pGA643 and pBI-VcFT containing a blueberry FLOWERING LOCUS T (VcFT). A total of 500 leaf explants, 250 for each construct, were used for transformation. After 10-week selection, three leaf explants transformed with the pGA643 produced four kanamycin-resistant shoots, in which stable integration and expression of the nptII were confirmed by Southern blot and RT-PCR analysis, respectively. This study demonstrated that it was possible to produce stable transgenic sweet cherry using Agrobacterium tumefaciens-mediated transformation of leaf explants.     

无花果新品种引种及高效栽培技术研究

课题组引进了"布兰瑞克"等无花果新品种,对其在宿迁地区的适应性及栽培表现进行鉴定,筛选出了适宜当地推广的优异品种。并以高产栽培技术集成为切入点,对无花果栽培技术要点进行总结,建立了优质高效的栽培技术体系,以期指导果农生产。     

无花果组培中培养条件对外植体褐化的影响

以无花果为组织培养试验材料,采用改变培养条件和使用不同培养基的方法,研究培养条件对外植体褐化的影响。结果表明:无花果外植体褐化在WPM培养基上比MS培养基上较轻,培养基中琼脂为9g的无花果外植体褐化较轻,经过暗处理的无花果外植体褐化程度较轻,以及低温条件能有效缓解外植体褐化。     

不同药剂处理对无花果根腐病的影响

以2～3a生"布兰瑞克"无花果为试材,研究4种组合药剂对无花果根部病害的防治效果。结果表明:从防治效果和成本看,福美砷+五氯硝基苯+多菌灵或福美砷+根病清+甲霜灵锰锌10～15g的300倍液为最佳组合药剂。     

桃幼胚下胚轴高频植株离体再生

以桃品种京艳和晚蜜不同发育阶段合子胚下胚轴为外植体进行离体培养,用6-BA、CPPU和TDZ等细胞分裂素诱导下胚轴直接再生不定芽试验。结果表明,花后70d下胚轴为诱导的最佳材料,供试细胞分裂素中,诱导活性为TDZ>CPPU>6-BA,苯脲类细胞分裂素TDZ5mg/L与NAA0.01mg/L配合对下胚轴再生具有极强的诱导作用,京艳诱导再生率高达95.2%,平均不定芽数达每外植体25.3个。京艳的诱导率显著高于晚蜜。IBA 0.5mg/L适于诱导再生幼茎的生根,生根率和移栽成活率分别达到92.6%和65.5%。组织学观察表明不定芽起源于皮层薄壁细胞,属直接再生方式。     

甜樱桃砧木离体叶片愈伤组织诱导及不定芽再生

叶片再生效率的高低直接影响目的基因转化的成功率,为建立稳定、高效的樱桃不定芽离体再生体系,以30～40d苗龄的甜樱桃砧木ZY-1的组培继代苗为试材,取上部幼嫩叶片或不含腋芽的茎段,分别从培养基生长调节剂配比、接种材料类型、叶片接种部位、接种方式以及培养条件等方面进行了不定芽再生技术研究。结果表明,培养基中添加7.0mg/L的6-BA与0.5～1.0mg/L的IBA配比时不定芽再生率和出芽数均较高,TDZ和NAA不适于诱导ZY-1叶片再生不定芽;接种继代苗茎段比叶片再生率高;接种叶片以选择嫩叶横切2刀、远轴面向下接触培养基的方式为好;叶片不同部位处理以带叶柄的基部叶块最易再生;叶片接种后在25℃室温条件下,先暗培养3周再照光,利于不定芽的再生。     

An efficient adventitious shoot regeneration system on excised leaves of micropropagated lingonberry (Vaccinium vitis-idaea L.)

Summary

An efficient system to regenerate shoots in vitro on excised leaves of lingonberry (Vaccinium vitis-idaea L.) was developed. Leaf explants from shoot-proliferating cultures produced multiple shoots without an intermediary callus phase on zeatin (ZN)-containing shoot induction media within 3–4 weeks of culture initiation. Cultivars Regal and Splendor, and one clone from a natural stand in Estonia (ECL1), were used in the first experiment. Young expanding leaves with the adaxial side touching the culture medium, and maintained for 7 d in darkness, produced the best results. There were significant genotypic differences in adventitious shoot formation. A second experiment studied the effects of ten concentrations of three cytokinins: ZN at 5, 10, 20, 30 and 40 μM; 1-phenyl-3-(1,2,3-thiadiazol-5-yl) urea (thidiazuron, TDZ) at 0.1, 1, 5 and 10 μm; and 6-(γ-γ-dimethylallylamino) purine (2ip) at 25 μM were compared with leaf segments of different polarity in ‘ECL1’. Zeatin was found to be more effective than TDZ or 2iP as an inductive signal for regenerating many vigorous shoots. Zeatin induced multiple shoot formation at all concentrations tested, but maximum morphogenic response was observed at 20 to 30 μM. The media containing TDZ generally promoted more callus formation and suppresed shoot elongation. In a third experiment with the lingonberry cultivar Erntedank and the clone ‘ECL1’, a new medium developed for lingonberry shoot culture proved more effective than the modified Murashige and Skoog medium for regenerating shoots on leaf explants. Elongated shoots were excised and rooted directly on a 2 peat:1 perlite (v/v) medium after dipping in 0.8% indole-3-butyric acid. Rooted plantlets were acclimatized under greenhouse conditions to evaluate somaclonal variation.     

Adventitious shoot regeneration from hypocotyl slices of mature apricot (Prunus armeniaca L.) seeds: A feasible alternative for apricot genetic engineering

Adventitious shoot regeneration from hypocotyl slices of mature apricot seeds has been achieved with regeneration percentages of 31.7%, 44.4%, and 46.9% for the cultivars ‘Canino’, ‘Dorada’, and ‘Moniqui’, respectively. Regeneration was significantly affected by the parental origin of the explants (P < 0.05) but not by thidiazuron or 3-indolebutyric acid for any of the three cultivars, at the levels tested. None of the other factors studied (basal medium, 2,4 dichlorophenoxy-acetic acid pulses, dark incubation period, or addition of silver thiosulfate) affected significantly shoot regeneration percentage from ‘Canino’ hypocotyl sections. The effect of paromomycin on regeneration was genotype-dependent and different dose–response curves were obtained for each cultivar. While 40 μM paromomycin completely inhibited regeneration from ‘Canino’ sections, some buds were obtained from ‘Dorada’ and ‘Moniqui’ explants. The two aminoglycoside antibiotics tested, kanamycin and paromomycin, showed differing toxicity on ‘Canino’. A lower concentration of kanamycin (20 μM) than of paromomycin inhibited totally adventitious regeneration from ‘Canino’ explants. Agrobacterium-mediated transformation experiments, with the non-oncogenic strain AGL1 harboring the binary plasmid p35SGUSINT, were performed and GUS assays were carried out after four weeks to determinate stable transformation events. The utilization of paromomycin (10 μM) as the selective agent increased significantly both the number of explants that presented at least one transformation event (P < 0.05) and the number of large area or calli expressing the gus gene (P < 0.001), compared with the addition of kanamycin (10 μM). Moreover, when 10 μM paromomycin was added to the medium some massively transformed explants were observed and a chimerical bud was regenerated.