Differences in Primary Callus Induction among Different Anthurium andraeanum Varieties

[Objective] This research aimed to optimize continuously the highly efficient regeneration system of Anthurium andraeanum. [Method] The leaves and petioles of four A. andraeanum varieties were used as explants to investigate the differences in primary callus induction among different A. andraeanum varieties. [Result]The callus formation capacity of SAM and SST was stronger than that of SDM and SHG. Among the four varieties, the leaf regeneration capacity of SAM, SDM and SHG was stronger than the corresponding petiole regeneration capacity. However,the petiole regeneration capacity of SST was stronger. The optimum medium for petiole callus induction of SST was 1/2 MS ＋ TDZ 4.0 mg/L ＋ 2, 4-D 0.2 mg/L with induction rate of 87.5%; the optimum medium for leaf callus induction of SAM was 1/2 MS ＋ TDZ 2.0 mg/L ＋ 2, 4-D 0.2 mg/L with induction rate more than90%; the optimum medium for leaf callus induction of SDM and SHG was all 1/2MS ＋ ZT 2.0 mg/L ＋ 2, 4-D 0.2 mg/L with induction rates of 59.34% and 79.63%,respectively. [Conclusion] In addition to variety differences, the differences in differentiation ability among different types of calluses should be also taken into account in the establishment and optimization of tissue culture and rapid propagation technology system of A. andraeanum.

Key Techniques of Industrial Seedling for Anthurium andraeanum

[Objective] The aim was to study the techniques for the tissue culture,rapid propagation and transplantation of Anthurium andraeanum.[Method] The leaf,leaf stalk,spathe and spadix of the pot flower variety and cut flower variety of A.andraeanum were used as explants,in order to investigate the influences of the explants selection,disinfection conditions and different media on the callus induction,proliferation and rooting.[Result] The leaf stalk of A.andraeanum was the optimum explant,with the highest callus induction rate（78.3%）.The best disinfecting effect could be obtained when the explants was sterilized with 0.1% HgCl2 for 8 min.The optimum media for the callus induction of Ping Champion and MIDORI were respectively 1/2 MS＋1.50 mg/L 6-BA＋0.50 mg/L 2,4-D＋0.10 mg/L NAA and 1/2 MS＋1.50 mg/L 6-BA＋1.00 mg/L KT＋0.10 mg/L 2,4-D,while the optimum media for callus subculture of Ping Champion and MIDORI were respectively MS＋1.00 mg/L 6-BA＋0.50 mg/L NAA＋0.30 mg/L 2,4-D and MS＋0.50 mg/L 6-BA＋0.50 mg/L KT＋0.10 mg/L NAA.It was beneficial for the proliferation and growth of cluster buds while 10%（w/v）of coconut water or banana jam was added to MS medium at the proliferation stage,and the proliferation rate could be increased by 2.83 fold.At the rooting stage,the medium 1/2 MS＋0.10 mg/L NAA＋0.10 mg/L IAA was the optimum medium for rooting,and the rooting rate was up to 100%.[Conservation] The research results will lay a foundation for the establishment of the industrial seedling breeding system of A.andraeanum,and it will be significant for the development of A.andraeanum industry.

Research on the Vase Preservative of Anthurium seherzerianum Cut Flowers

[Objective] The research aimed to study the vase preservative of Anthurium seherzerianum cut flowers and provide reference for the planting and preservative method selection of A.seherzerianum.[Method] Using Tropical＇s spathe,one kind of common cut blossoms of A.seherzerianum as experimental materials,three different formula of vase solutions were used.And it was compared with common preservative formula of A.seherzerianum to study the vase preserved liquid of A.seherzerianum.[Result] Formula 3 was the optimum vase preservative,which was composed of 4% sucrose,0.08% NaCl,0.01% Ca（H2PO4）2·H2O,0.01% Chinese medicine antiseptic （ethonal extract from coptis）,0.1 mol/L NaOH,0.1 mol/L citric acid and 10 mg/L 6-BA.The preserved period reached 23 days,13 days longer than CK.The appreciative period reached 31 days,18 days longer than CK,with significant effects.In formula 3,water loss in A.seherzerianum spathe was decreased obviously,which was favorable for maintaining water content in tissues.The cell membrane permeability was reduced and the peroxidation of membrane ester was inhibited.The accumulation of MDA was decreased and SOD activity was increased.The protective enzyme activity of cells was enhanced and the content of proline and soluble sugar were increased and the respiration rate was reduced.[Conclusion] The formula（4% sucrose ＋ 0.08% NaCl＋ 0.01% Ca（H2PO4）2＋0.01% Chinese medicine antiseptic（ethonal extract from coptis）＋ 0.1 mol/L NaOH＋0.1 mol/L citric acid and 10 mg/L 6-BA）achieved the purpose of prolonging the preservative period of A.seherzerianum spathe and prolonging the preservative period of A.seherzerianum effectively.

长期CO_2加富对苗期红掌(Anthurium andraeanum L.)植株生长和光合作用的影响

以开顶式塑料薄膜温室为设施,研究了红掌(AnthuriumandraeanumL.)幼苗植株生长、叶片净光合速率和光合酶活性对长期高CO2浓度的响应。结果表明:处理90d时,处理组T1((700±100)μmol·mol-1CO2)的株高、单叶面积、株鲜重分别比对照组((360±30)μmol·mol-1)增加了15.76%、14.30%、29.62%,而处理组T2((1000±100)μmo·lmol-1CO2)的株高、单叶面积、株鲜重分别比对照增加了15.00%、9.63%、36·22%;处理150d时T1的株高、单叶面积、株鲜重与对照相比分别增加了16·08%、17.30%、49.09%,而T2增加了16.61%、10.10%、48.87%。在各自生长环境下处理组T1、T2的净光合速率在整个处理期间均高于对照,处理150d时,T1、T2的净光合速率分别比对照高8.25%、20.62%;但处理90d时,在对照CO2浓度下测定的净光合速率处理组开始低于对照组,可能此时处理组的红掌叶片开始出现光合适应现象;CO2浓度升高促进了叶片中可溶性糖和淀粉积累,处理90d时T1、T2处理组中淀粉含量分别比对照高52.60%、67.66%;处理150d时,T1组红掌叶片中淀粉与可溶性糖含量比对照高53.43%、6.32%,T2比对照高58.44%、8.07%,叶绿素含量在处理90d时也开始低于对照组;整个实验过程中,Rubisco活性前期增加,90d以后开始下降;乙醇酸氧化酶活性则明显下降,T1、T2处理组试验结束时与对照组相比分别下降了41.28%、45.35%。一定处理时间(90d)的高浓度CO2处理提高了红掌叶片的净光合速率和碳水化合物的积累,促进了营养生长,但随着处理时间的延长,这种促进作用逐渐降低。

Plant Regeneration by Callus-Mediated Protocorm-Like Body Induction of Anthurium andraeanum Hort.

This research aims at developing a plant regeneration system from leaf and petiole explants of Anthurium andraeanum Hort., thereby establish a foundation for mass production and transformation. Using tissue culture technique, the conditions for callus induction, protocorm-like body （PLB） formation and plant regeneration from leaf explants and petiole of A. andraeanum, such as basal medium and plant growth regulator, were investigated. Totipotent callus was induced on a 1/2-strength MS medium containing 0.90 μmol L^-1 2,4-dichlorophenoxyacetic acid （2,4-D） and 8.88μmol L^-1 N6-benzyladenine （BA）. The callus exhibited complete hormone autonomy for growth and differentiation of PLBs. This callus proliferated well and was maintained by subculturing on 1/2 MS medium containing 0.90 μmol L^-1 2,4-D and 4.44 μmol L^-1 BA. On average, 8 protocorm-like bodies could be obtained from a piece of 4 mm callus after being transferred to the 1/2 MS medium with 4.44 μmol L^-1 BA after 8 wk of culture. The regenerated PLBs formed shoots and roots on 1/2 MS medium. After 24 wk of culture on these medium, well-developed plantlets for potting were produced. An efficient micropropagation method was established for indirect PLB formation and plant regeneration from leaf and petiole ofA. andraeanum.

Antibacterial Effects of Nanometallic Materials in Genetic Transformation of Anthurium by Agrobacterium-mediated Method

[Objective] This study was conducted to investigate the application of nanometallic materials in inhibiting Agrobacteriurn contamination in genetic transformation of Anthurium. [Method] Different nanometallic materials were added into Agrobacterium medium and Anthurium callus medium, to investigate the effects of their effects on Agrobacterium growth, callus growth and differentiation, and Agrobacterium contamination. [Result] Among the 4 nanometallic materials, NanoAg-2 showed a significant inhibitory effect on the growth of Agrobacterium, with a minimal inhibitory concentration of 25 mg/L. Even for the Anthurium calli or transgenic material contaminated by the Agrobacterium, a good antibacterial effect could be achieved after treating with 25 mg/L NanoAg-2 for 1 d with oscillation, the antibacterial rate reached 100%, and the Anthurium calli could grow and differentiate normally. [Conclusioa] NanoAg-2 could effectively inhibit Agrobacterium contamination, and its an- tibacterial effect is significantly better than cephalosporin and carbenicillin.

Preliminary Study on the Mechanism of Flower Color Variation in White Mutants of Anthurium andeaeanum

[Objectives] This study was conducted to explore the physiological mechanism of flower color variation in the white mutants of Anthurium andeaeanum. [Methods] The seven white mutants of 'Alabama' and 'Turenza' were used as materials to analyze the pigment types, flavonoid types and content and anthocyanin content in the wild type and mutants. [Results] The white spathe mainly contained flavonoids, flavonols, dihydroflavonols and dihydroflavonols; the white mutants of 'Alabama' had a higher total flavonoid content than the wild type, while the white mutants of 'Turenza' showed an opposite trend; and the spathe of the wild type had the highest anthocyanin content, and the pink part of the two-color mutant or the spathe of the binary color mutant contained trace anthocyanins, while no anthocyanins were detected in the white part of the mutants. [Conclusions] The main cause of the white mutants of A. andeaeanum is related to anthocyanin metabolism.

红掌(Anthurium andreanum Dakota)幼叶诱导愈伤组织研究

采用红掌(Anthurium andreanum Dakota)幼叶叶片中部(带主叶脉叶片),叶片基部(叶柄与叶片连接处),叶尖部位作为外植体,接种在1/4 MS附加不同浓度的6-BA+2,4-D+KT的培养基上.结果表明,红掌(Anthurium andreanum)愈伤组织的诱导率不仅与叶片不同部位有关,而且与添加外源激素的浓度及配比有关.经过60 d愈伤组织诱导培养,叶片基部作为外植体诱导愈伤组织效果最好,愈伤组织诱导率最高的培养基为1/4 MS+6-BA0.9 mg/L+2,4-D 0.9 mg/L+KT0.5 mg/L.

Molecular-based Integrated Identification of Bacterial Blight(Xanthomonas axonopodis pv. dieffenbachiae) in Anthurium and Detection of Latent Infection

Bacterial blight, caused by Xanthomonas axonopodis pv. dieffenbachiae （Xad）, is the most destructive disease of anthurium worldwide, and no effective control technique has been developed currently. The comprehensive survey and precise detection of the pathogen is essential for evaluating disease progress and strengthening management to avoid a serious epidemic. In this study, a total of 253 blight-suspected samples of anthurum and other Araceae species were collected across the country, and 166 potential pathogenic bacteria strains were isolated and purified, after combined analysis on the characteristics of morphology, pathoge- nicity, 16S rDNA sequences and amplicans of Xad-specific SCAR markers. Finally, 93 of which were considered as X. axonopod/s pv. dieffenbachiae. In addition, by using a nested-PCR in repeated detections, 17 out of 21 prevalent anthurium cultivars without blight symptom exhibited latent infection even in young leaves. The results indicated that the anthurium bacterial blight distributed commonly in growing areas in China, and most of the commercial cuhivars had no strong resistance. The identification of Xad infection （latent） would be beneficial for the disease forecasting and management improving in anthttrium production.

Biological Features of Flowering and Development of Male Gametophyte in Anthurium andreanum

The aim of this study is to follow each development stage of inflorescence in order to understand the biological feature of flowering and the development of male gametophyte in Anthurium andreanum “Arizona' and to try to find the optimum conditions for its pollination. The methods of dissection and paraffin section were adopted to examine the structural characteristics of anthurium’s tiny floret and the development of the microspore. All the florets of the anthurium arrange on the rhachis helically sub- tended by a colorful bract. Each tiny floret has one gynoecium, four tepals and four stamina. The bract and the florets show different colors during the whole blooming period. The ovary is bicarpellary and has two locules, each of which has one anatropous ovule. The placenta is of a central placentation type. The stylar canal cells not only can produce the secretory mucilage but also can release their own cytoplasm caused by their self-disintegration before the pistil reaches its maturity. The wall of the anther is composed of four layers: epidermis, endothecium, middle layer and tapetum. The tapetal cells and the middle layers’ cells degenerated completely dur- ing meiosis of microsporocytes. The pollen grains were 2-celled at the time of anther dehiscence. Early morning, when the inflores- cences stay at their fifth development stage, is the optimum opportunity for pistil to get pollen grains. The pollen-collection should be done at the end of the seventh stage.

Advances in Transgenic Breeding of Anthurium andraeanum

At present, transgenic technologies have become important means of plant breeding, and the application and promotion of transgenic technologies have created huge economic and social benefits. Transgenic plant products have significantly affected human life. Anthurium andraeanum is the second major tropical potted flower and its transgenic breeding has a promising prospect of application. In this paper, acceptors, transformation methods and introduced exogenous genes （ including reporter genes, selectable marker genes and target genes） of Anthurium andraeanum were summarized; in addition, several issues related to transforma- tion of Anthurium andraeanum were analyzed, aiming at providing reference for transgenic breeding of Anthurium andraeanum.

1-DE/MS Analysis of the Proteins Related to Spathe Color Variation in Anthurium andraeanum 'Madural'

In order to understand the mechanism of spathe color variation in Anthurium andraeanum at the protein level, the leaves, inflorescences and spathes of the wild type and two mutants of A. andraeanum 'Madural' were used as research objects in which the differential expression of proteins related to flower color mutants was analyzed by one-dimensional electrophoresis and mass spectrometry(1-DE/MS). The 1-DE patterns showed that the protein components expressed highly in spathes were mainly concentrated in the molecular weight range of 20-42 kD, and differential bands were detected between the wild type and the mutant, while no significant differences were detected in the leaf and inflorescence proteins. According to the results of mass spectrometry analysis of the differential bands, 21 known functional proteins involved in life processes such as glucose metabolism, resistance, cytoskeleton, gene regulation and signal transduction were identified. It showed that in addition to the influences from anthocyanidins, the spathe color variation of A. andraeanum 'Madural' is also regulated by a variety of metabolic pathway-related proteins.

1-DE/MS Analysis of Proteins Related to the Spathe Color Variation in an Anthurium andraeanum Mutant

[Objectives] This study was conducted to explore the color variation mechanism of Anthurium andraeanum spathe at the protein level. [Methods]The differential proteins of wild type and its white mutant were separated and identified by using one-dimensional gel electrophoresis and mass spectrometry( 1-DE/MS). [Results] Compared with leaves and spadices,the 1-DE patterns of two kinds of spathe proteins were significantly different,and two different bands were detected in wild type spathes and mutant spathes respectively. The four significantly differential bands were selected and analyzed by mass spectrometry,and 138,111,70 and 427 proteins were identified respectively. The results of GO functional annotation analysis showed that the molecular functions of the proteins were mainly catalytic activity and binding,and the main biological processes involved were cellular process and metabolic process. Many proteins involved in the synthesis of anthocyanins and flavonoids,sugar metabolism and some resistance proteins were screened,indicating that the spathe color difference of A. andraeanum‘Pink champion'is not only related to anthocyanin anabolism,but also regulated by various metabolic pathways. [Conclusions]The study provides a new experimental basis for elucidating the molecular mechanism of the regulation of A. andraeanum flower color.

A Study on the Sensibility of Embryogenic Suspension Cells of Anthurium andraeanum to Two Antibiotics

In this study,embryogenic cell aggregates obtained from the established embryogenic cell suspension culture system of Anthurium andraeanum‘Alabama’were used as experimental materials to investigate the effects of kanamycin and hygromycin on survival rate of cell aggregates,somatic embryogenesis and plantlet regeneration of A.andraeanum.According to the results,at the embryonic cell propagation stage,lethal doses of kanamycin and hygromycin to embryogenic cell aggregates of A.andraeanum were 200 and 60 mg/L,respectively;at the differentiation stage,either 150 mg/L kanamycin or 40 mg/L hygromycin could inhibit somatic embryogenesis;either 100 mg/L kanamycin or 20 mg/L hygromycin could inhibit plantlet regeneration.These results provided important reference for further studies of transgenic A.andraeanum.

红掌(Anthurium andraeanum)细胞工程研究进展及其在育种上的应用

红掌是一种重要的观花和观叶兼具的草本植物。植物离体培养技术已经成为红掌繁殖主要采用的方法,绝大部分品种可以通过组织培养实现再生,其推动了红掌细胞工程育种的发展。本综述总结了红掌细胞组织培养繁殖途径,包括直接获得无菌苗途径、愈伤组织诱导和不定芽再生途径、体细胞胚胎发生途径、原球茎和类原球茎途径、原生质体途径等方面的研究进展,及其花器官培养,多倍体诱导,物理化学诱变,遗传转化,体细胞无性系变异及筛选的研究在育种中的应用。并提出了红掌细胞工程研究当前存在的问题和以后的研究方向,以期为将来红掌的研究提供借鉴。

基于自由曲面和L-system的红掌生长模型

针对红掌花卉生长可视化中参数难以确定的问题进行研究,提出推导控制点的方法,建立红掌器官模型:采用Bezier曲线结合扫描成型的方法来构造叶柄模型,采用双三次NURBS曲线对佛焰苞进行建模;运用红掌的相关表型数据拟合生长函数,提出基于红掌生长规律的微分L-system,可有效模拟红掌的拓扑结构和生长过程;通过虚拟器官表示红掌器官的几何属性,降低微分L-system的复杂度。试验验证提出的方法对红掌各项生长指标拟合度可达0.89以上,并可对每个生长阶段的红掌进行模拟,能有效地对红掌的生长过程进行建模。

Physiological Effect of Kinetin on the Photosynthetic Apparatus and Antioxidant Enzymes Activities During Production of Anthurium

The results observed in the literature raise the hypothesis according to which cytokinin plays important roles in photosynthetic metabolisms and antioxidant enzymes. Thus, the study aimed to evaluate the effect of foliar application of the isolated cytokinin kinetin at the production cycle, seeking to analyze its effects on enzyme activity and photosynthetic parameters. The plants treated with CK presented reduction of leaf CO_2 assimilation rate(Pn) and stomatal conductance(Gs), while that transpiration rate(Tr) was unaffected. The internal CO_2 concentrations decreased with the increase in cytokinin levels, but were maintained under CK 50 mg·L^(-1). The plants treated with CK 75 mg·L^(-1) was verified higher carboxylation efficiency(Pn/Ci), which was associated to values of CO_2 assimilation and transpiration unaltered. Apparent electron transport rate showed variations in the concentration of 25 mg·L^(-1). Considering the study of enzyme activity, on the other hand, it cannot be stated that kinetin has an effective action in delaying oxidative damage. It presents mixed results, since an efficiency in the application of cytokinin was not observed, presenting induction levels of ascorbate peroxidase activity. Thus, further research is needed to determine more precisely the effects of kinetin on gas exchange and antioxidant enzymes in anthurium plants.

Cloning and functional analysis of a novel ascorbate peroxidase(APX) gene from Anthurium andraeanum

An 888-bp ful-length ascorbate peroxidase （APX） complementary DNA （cDNA） gene was cloned from Anthurium andraeanum, and designated as AnAPX. It contains a 110-bp 5′-noncoding region, a 28-bp 3′-noncoding region, and a 750-bp open reading frame （ORF）. This protein is hydrophilic with an aliphatic index of 81.64 and its structure consisting ofα-helixes,β-turns, and random coils. The AnAPX protein showed 93%, 87%, 87%, 87%, and 86% similarities to the APX homologs from Zantedeschia aethiopica, Vitis pseudoreticulata, Gossypium hirsutum, Elaeis guineensis, and Zea mays, respectively. AnAPX gene transcript was measured non-significantly in roots, stems, leaves, spathes, and spadices by real-time polymerase chain reaction （RT-PCR） analysis. Interestingly, this gene expression was remarkably up-regulated in response to a cold stress under 6 °C, implying that AnAPX might play an important role in A. andraeanum tolerance to cold stress. To confirm this function we overexpressed AnAPX in tobacco plants by transformation with an AnAPX expression construct driven by CaMV 35S promoter. The transformed tobacco seedlings under 4 °C showed less electrolyte leakage （EL） and malondialdehyde （MDA） content than the control. The content of MDA was correlated with chilling tolerance in these transgenic plants. These results show that AnAPX can prevent the chilling challenged plant from cellmembrane damage and ultimately enhance the plant cold tolerance.

红掌特伦萨四倍体的离体诱导及其鉴定

为了研究红掌四倍体的离体诱导和鉴定技术体系,为其倍性育种和杂交育种奠定基础,以特伦萨愈伤组织为试验材料,采用0.5、1.0、2.0、4.0 g·L^(-1)的秋水仙素分别处理2、4、6、8 h,之后转入培养基MS+6-BA 2.0 mg·L^(-1)+NAA 0.5 mg·L^(-1)进行愈伤组织的生长和丛生芽分化,75 d后将丛生芽转入培养基1/2 MS+NAA 0.2 mg·L^(-1)诱导生根,待丛生芽生根后统计愈伤组织块存活率、再生植株数,并对再生植株进行染色体倍性鉴定和核型分析。结果表明,随着秋水仙素浓度的增加,红掌愈伤组织块存活率呈下降趋势,但愈伤组织块上诱导的再生植株数无显著变化;在0.5~2.0 g·L^(-1)的秋水仙素浓度范围内,红掌疑似四倍体的诱导率随着处理浓度与时间的增加而增加,2.0 g·L^(-1)秋水仙素处理6与8 h时,诱导率达8.50%与8.77%;特伦萨和疑似四倍体红掌的基因组分别为3.73和7.01 Gb;染色体压片倍性鉴定结果证明了疑似四倍体的倍性为四倍体。红掌二倍体的核型公式为2n=2x=30=8 st+16 sm+6 m,四倍体的核型公式为2n=4x=60=8 st+32 sm+20 m;与红掌二倍体植株相比,四倍体植株叶片变厚、气孔变大、保卫细胞密度变小,佛焰苞直径变大,单个花序花期变长,差异均达显著水平。综上,采用2.0 g·L^(-1)秋水仙素处理6 h可培育红掌四倍体植株,此四倍体可作为红掌倍性育种和杂交育种的重要种质资源。

红掌火烈鸟组培快繁技术

探讨红掌新品系火烈鸟组织快繁技术,为实现其规模化生产提供技术支持。以火烈鸟为材料,研究其组织培养技术。结果表明,火烈鸟的嫩叶可以被诱导分化出不定芽,外植体的最佳消毒时间为10 min,以1/2 MS添加1.5 mg/L 6-BA和0.3 mg/L NAA为诱导培养基,不定芽的分化情况最好;在增殖培养基中添加0.6 mg/L 6-BA和0.2 mg/L IBA,火烈鸟增殖系数最高,达到3.44;在生根培养基中添加0.2 mg/L NAA和0.2 mg/L IBA,培养至30 d时,生根率已高达100%。通过叶片直接诱导出不定芽,可以建立火烈鸟高效快繁技术体系,为红掌的规模化繁殖及工厂化生产提供理论依据。