Molecular Detection of Phytophthora colocasiae of Taro Leaf Blight Based on PCR

The present PCR assay was conducted to develop rapid and sensitive detection of Phytophthora colocasiae,in order to provide a robust and reliable tool for healthy seedling production of taro and limiting the transmission and spread of the causal organism of taro leaf blight in taro planting regions.The samples were used to extract total DNA and to be detected by PCR with P.colocasiae specific primer pairs PCSP-RL F/PCSP-RL R and PCSP-T F/PCSP-T R,respectively.Distinct fragments of about 200 bp and 240 bp were amplified by PCR using primers PCSP-RL F/PCSP-RL R and PCSP-T F/PCSP-T R,respectively.The analysis of the nucleotide sequence of the PCR products were found to be 99% identical to sequence of RAS-related protein (Ypt1) and phospho-ribosylanthranilate isomerase (TRP1) in P.colocasiae,respectively.It is concluded that rapid and sensitive developed PCR assay for detection of P.colocasiae could be used in routine diagnosis and aid in management practices to mitigate taro leaf blight.

芋[Colocasiaesculenta(L.) Schott]的倍性水平及基因组特征解析

采用流式细胞术,对2个芋[Colocasiaesculenta(L.)Schott]品种苏芋1号和香蕉芋的倍性水平和DNA含量进行了测定,2倍基因组DNA含量分别为7.71 pg和5.25 pg,倍性分别为三倍体和二倍体。同时通过HiSeq 2500测序,分别获得240.53 Gb和87.69 Gb数据,芋基因组大小为2.08~2.38 Gb,GC含量为42.57%~42.75%,杂合度为0.52%~2.29%。根据苏芋1号的基因组数据,组装得到基因组序列为2.27 Gb,Contig N50为359 bp;共注释获得24302个基因,99.43%的基因可注释到6个公共数据库;筛选出928987个SSR位点。

芋(Colocasia esculenta)的民族植物学

应用民族植物学的基本原理和方法,选择中国云南和山东为试点,兼顾其他省区,开展芋〔Colocasiaesculenta(L. )Schott〕的民族植物学研究。结果表明:在云南传统栽种芋的菜园和农地被高附加值的经济作物所代替,芋在不同民族家庭中的地位也从传统作为主食变成蔬菜或杂粮;在山东已形成芋的产业化、标准化生产的格局,芋在汉族农家经济中的地位得到提升。在云南分布有芋的野生近缘种、半栽培种、栽培品种,种质资源丰富;在山东未发现芋的野生类型,以旱芋类型的多子芋栽培品种为主。由于经济的发展和主流文化的影响,民间对芋的植物崇拜及植物崇拜文化丢失的速度大大加剧。在古朴的传统食芋文化中,蕴含着丰富的关于芋植物资源利用和保护的传统知识和朴素的科学内涵,需要进行深入的挖掘和探讨。

Transcriptome analysis of purple pigment formation in Colocasia esculenta

Taro(Colocasia esculenta(L.)Schott)is an important crop in Africa,Southeast Asia,and subtropics and is used as a food and medicine.The purple color pigmentation is an appealing character in taro.We sampled taro corms of the cultivar‘Lipu Taro’at four developmental stages,including LPYS1(without purple pigment,50 days of development(DOD)),LPYS2(very few purple pigments,75 DOD),LPYS3(moderate purple pigments,115 DOD)and LPYS4(high purple pigments,205 DOD).The purpose of our study was to identify the key genes underpinning the purple pigmentation in taro based on RNA-sequencing.Through RNA-Seq,6453 differentially expressed transcripts(DETs)were identified between purple and non-purple pigmented samples.We identified 41 and 12 flavonoid and anthocyanin related DETs transcripts,respectively.These DETs were upregulated at LPYS2,LPYS3,and LPYS4 as compared to LPYS1,indicating their positive contribution to the color formation in taro.Moreover,we identified several DETs encoding for transcription factors,including MYB and bHLH,known to be major regulators of structural genes involved in the flavonoid-anthocyanin pathway.Finally,we reported several plant hormones(ethylene,auxin,gibberellin,jasmonic acid,and cytokinin)related DETs,which are predicted to play important roles in the corm coloration.Different regulation of transcripts representing the flavonoid-anthocyanin biosynthesis pathway,plant hormone transduction pathway,and transcription factors may have key roles in purple pigmentation in taro.Our findings will facilitate future research on improving the quality and appeal of taro.

Synthesis of silver nanoparticle with Colocasia esculenta(L.) stem and its larvicidal activity against Culex quinquefasciatus and Chironomus sp

Objective:To synthesize silver nanoparticles with Colocasia esculenta as a reducing agent and to evaluate their effect against Culex quinquefasciatus and Chironomus sp.Methods:The aqueous extract of Colocasia esculenta stem was used for nanosynthesis.The synthesized nanoparticles were characterized by UV-Vis spectrophotometry,Fourier-transform infrared spectroscopy,scanning electron microscope,transmission electron microscopy,energy-dispersive X-ray spectroscopy,X-ray diffraction and Zeta potential studies.The toxicity of Colocasia esculenta stem extract and the synthesized silver nanoparticles was evaluated against the larval stages of target human filarial vector Culex quinquefasciatus and non-target Chironomus sp.Results:Scanning electron microscopy and transmission electron microscopy studies revealed almost spherical shape of the synthesized silver nanoparticles with size ranging from 13-50 nm.After 24 hours of exposure,the LC50 and LC90 of the plant extract against 4th instars larvae of Culex quinquefasciatus were 745.56 mg/L and 1258.28 mg/L,respectively,which were higher than those of synthesized silver nanoparticles(5.17 mg/L and 17.32 mg/L after 24 h;1.58 mg/L and 13.01 mg/L after 48 h).In addition,the LC50 and LC90 of silver nanoparticles against Chironomus sp.were 9.71 mg/L and 23.15 mg/L after 24 h as well as 2.38 mg/L and 19.49 mg/L after 48 h,respectively.Conclusions:The aqueous stem extract of Colocasia esculenta is a good agent for synthesis of silver nanoparticles,which are almost spherical with size less than 30 nm.The synthesized nanoparticles show good larvicidal activity without any harmful effect on non-target species.

Improvement of Two-Dimensional Gel Electrophoresis for Study of Corm Formation Related Proteins in vitro from Taro (Colocasia esculenta)

Efficient and reproducible sample preparation prior to 2D-PAGE (two-dimensional polyacrylamide gel electrophoresis) is a critical step in achieving accurate and reliable data. In this paper, we described a method to prepare protein samples of taro that was compatible with subsequent analysis using 2D-PAGE. We compared proteins from shoot basal region from 0 d and 2 d after the beginning of tuberization. By this method we got about (2 000) spots and high reproducibility. Additionally some changes of protein expression were found.

<i>In Vitro</i>Organogenesis of <i>Colocasia esculenta</i>cv. <i>Antiquorum</i>L.

In vitro organogenesis of an upland species of Colocasia esculenta cv. antiquorum L. was examined in relation to different explants like meristem and parenchymatous storage tissues with or without anthocyanin layer, four levels of each of Kn, 2,4-D, NAA and BAP and four incubation environments such as: 1) 16 h 3 Kl light intensity + 24°C ± 2°C;2) 24 h dark + 24°C ± 2°C;3) 24 h dark + 30°C ± 3°C and 4) 12 h diffuse light + 30°C ± 3°C. Only meristems showed proliferation with various degree of intensity both at 16 h 3 Kl light + 24°C ± 2°C and 24 h dark + 24°C ± 2°C conditions and poor response with different levels of Kn + NAA either in light or in the dark. Cultures with NAA + BAP were proliferated very quickly with very high degree of intensity. The cultures under dark did not proliferate for 20 days which upon transfer to light showed high degree of proliferation. Cultures with NAA + BAP formed calluses more pronouncedly at dark than that occurred in the light. Parenchymatous tissues with or without anthocyanin did not proliferate but the tissues with anthocyanin lost pigmentation after 25 - 30 days and turned to grey colour after 50 days while tissues without anthocyanin turned to green colour with shinny pimples indicating that protocorm may be developed. No culture under high temperature environment (30°C ± 3°C) neither survived nor proliferated. The meristems in culture were died within 15 - 20 days while others within 25-30 days. In conclusion, a combination of NAA (0.5 - 3.0 mg/l) and BAP (0.5 - 2.0 mg/l) and an incubation photoperiod of 16 h coupled with temperature of 24°C ± 2°C were found most suitable for in vitro culture of Colocasia esculenta cv. antiquorum L.

Morphological Comparison of Five Varieties of <i>Colocasia esculenta</i>(L.) Schott in Anambra State, Southeastern Nigeria

Comparative morphological study of five varieties of Colocasia esculenta (L.) Schott present in Anambra State, Nigeria was carried out, in order to furnish plant taxonomists with information which could be of great help in delimitation of the varieties. Significant difference was established at p < 0.05. The result showed that adaxial surface of “kochuo” had a purplish dot on the centre, abaxial leaf surface had a conspicuous purplish colour at the point of attachment to the leaf, and the cormels had numerous striking vertical purple stripes on the surface. There was presence of foliaceous (leaf-like) appendages at the veins of the abaxial surface of the leaf of “ogeriobosi”. The leaf length ranged from 35.6 ± 7.70 cm (“kochuo”) to 49.9 ± 3.55 cm (“ogeriobosi”). Petiole length of “ogeriobosi” was the highest (63.3 ± 3.83 cm), whereas the least was Colocasia esculenta var. antiquorum (26.67 ± 2.20 cm). The corm length ranged from 4.10 ± 0.10 cm (Colocasia esculenta var. antiquorum) to 8.60 ± 0.35 cm (“ogeriobosi”), while the cormel length ranged from 3.70 ± 0.96 cm (Colocasia esculenta var. antiquorum) to 7.03 ± 0.36 cm (“ogeriobosi”). This work has revealed diagnostic and differential morphological characters, which could be useful for identification and description of varieties of C. esculenta. In addition, it provided additional information which might be helpful in resolving the on-going controversy in the taxonomy of Colocasia, which would, in turn, probably lead to possible delimitation of C. esculenta.

Characterization of a Highly Potent Insecticidal Lectin from <i>Colocasia esculenta</i>Tuber and Cloning of Its Coding Sequence

Hemipteran insects are the most devastating pest for different crops of high economic value. Colocasia esculenta tuber agglutinin (CEA), a mannose binding monocot lectin from araceae family was previously reported by the present group to be effective against some members of this class of pests. In the present study, efficacy of this potent lectin has been extended to cotton aphid (Aphis gossypii) which is becoming a highly damaging pest of cotton in recent days. Because, like other aphids, A. gossypii not only extracts the phloem fluid but also transmit disease causing viruses and add to the high degree of yield loss. Efficacy of the lectin on cotton aphid as well as other hemipteran insects prompted us further to clone the protein coding gene. Very little sequence information of this gene was available in the database. Hence, attempt had been made to study the protein through liquid chromatography-tandem mass spectrometry (LC-MS/MS) to have the detailed peptide information. On the basis of the peptide homology information obtained from LC-MS/MS the complete coding sequence of CEA was determined. The coding sequence corresponding to CEA was cloned further using primers designed on the basis of above information and genome walk technology for its potential utilisation in insect management programme.

Effect of Different Levels of Nitrogen on Yield of Colocasia (<i>Colocasia esculenta</i>) at District Malakand Dargai

A field experiment was conducted to evaluate the response of colocasia (Colocasia esculenta) to different levels of 0, 60, 90, 120 and 150 kg N ha-1 under farmer’s field condition at Garhi Usmani Khel, District Malakand Dargai during 2013. The experiment was laid out in Randomized Complete Block design with three replications and treatment plot size of 2.74 × 2.43 m2. All levels of N in the form of urea along with uniform basal doze of 90 kg P2O5 ha-1 as Triple Super Phosphate (TSPPP) were applied to soil at time of seed bed followed by thorough mixing. Seeds of colocasia c.v. local variety were sown in these plots with row spacing of 30 cm and plant to plant distance of 12 cm in February, 2013. The results showed that application of N produced significantly higher colocasia tuber yield, number of tubers plant-1, 1000-tubers weight and size of tubers (mean length and diameter) over control but the differences among levels of N were nonsignificant. However, some parameters like tuber yield was maximum at 60 kg N ha-1 and tuber size especially the length of colocasia tuber was maximum at 150 kg N ha-1 suggesting that the response of each parameter was different to N levels. Based on maximum relative yield (100%) and increase over control (46.1%) still at lower N levels of 60 kg N ha-1, this level seems to be appropriate level for colocasia under the prevailing soil and climatic conditions.

Evaluation of the Value Nutritious Leaves Five Varieties Taro (Colocasia esculenta) Cultivated in Burkina Faso

Taro is a perennial herbaceous plant whose large leaves are mainly used as vegetables in human food in several tropical countries. However, young taro leaves are not eaten very much in Burkina Faso unlike other countries which have made them a staple diet. In the present work, we collected leaves of taro varieties cultivated in the provinces of Comoe and Kenedougou. Our study aimed to determine the biochemical composition of these leaves in order to detect their nutritional quality. For this purpose, we first determined the total sugars in our different samples;then quantify the proteins and finally assay the lipids contained in the leaves of the different varieties of taro harvested. Analysis of the organic constituents gave the following results: proteins (186.29 to 265.23 μg EQ/100mg fresh leaves), fats (0.28% to 1.90%), carbohydrates (183.03 to 238.57 μg EG/100mg fresh leaves). The highest energy value was obtained with the variety BF/CO/06 (1728.71 kcal/kg) and the lowest with BF/CO/04 (272.15 kcal/kg). This study allowed us to conclude that the taro leaves (Colocasia esculenta) studied are of nutritional interest with regard to their biochemical composition.

Comparison of the Effectiveness of Different Traditional Soaking Processes on the in Vitro Digestibility of Taro(Colocasia esculenta L.SCHOTT)Flour

A traditional process used by farmers in Chad consists in soaking slices of taro (Colocasia esculenta L. SCHOTT) in tamarind infusion, or in corn solution or in water over a 24-hour period to reduce the acridity of taro and facilitate cooking. The aim of this study was to assess the effect of traditional soaking on the in vitro digestibility of taro flour using or not using an α-amylase enzyme. The digestion without the enzyme has shown that the soaking processes improve the digestibility of taro flour (from 39.30% for the control sample to 75.11% (after tamarind infusion) and 78.67% (treatment with water) after 24 hours of soaking). Soaking over a 6-hour period and preferentially in tamarind infusion or in corn solution obtains highly digestible flour (around 95% of digestibility rate after 3 hours of enzymatic digestion).

芋可溶性淀粉合成酶CeSS基因家族的克隆和表达分析

为探索芋淀粉合成机理,本研究从靖江香沙芋中克隆获得了3个芋可溶性淀粉合酶(CeSS)家族基因。3个基因CeSSI、CeSSII、CeSSIII碱基序列全长分别为1932 bp、2409 bp、3606 bp,编码形成3个亲水性蛋白质。系统进化分析结果显示,3个蛋白质CeSSI、CeSSII、CeSSIII与海枣、芦笋、药用稻等单子叶植物的亲缘关系较近。表达分析结果表明,这3个CeSS基因广泛存在于芋的叶片、叶柄、母芋和子芋中。CeSSI和CeSSII的表达水平随芋球茎的发育而显著增加,在种植150 d达到峰值,而CeSSIII在成熟后期(种植180 d)的芋球茎中表达量较高。

江西铅山红芽芋叶绿体基因组特征及系统发育分析

为分析江西铅山红芽芋叶绿体基因组的结构组成情况,判定其在芋属中的进化位置及与同芋属叶绿体基因组的区别,为芋属物种鉴定、遗传多样性分析和资源保护提供相关依据。使用Illumina NovaSeq 6000测序平台对江西铅山红芽芋叶绿体基因组进行测序,通过生物信息学分析方法进行序列组装、注释和特征分析,并利用Geneious、MISA、MAFF、FASTTREE、CUSP、Chips和Codon W等生物信息学软件对其基因组结构和数目、密码子偏好性、序列重复、SSR位点和系统发育进行分析。结果表明,江西铅山红芽芋叶绿体基因组大小为16 2544 bp,呈现四分体结构。共包含130个基因,其中84个编码蛋白质的基因,37个t RNA基因,8个核糖体rRNA基因,1个假基因。通过密码子偏好性分析,平均有效密码子数为45.60,ENC值小于45的基因39个,说明其密码子偏好性强。通过SSR分析,检测到101个SSR位点,其中单核苷酸重复最多(约占83.17%),且单核苷酸以A/T型为主。与近缘种比较,发现其叶绿体基因组序列高度保守,尤其蛋白质编码序列相似度极高。此外,系统发育分析发现江西铅山红芽芋与芋属、岩芋属聚为一支。本研究得到了江西铅山红芽芋的叶绿体基因组基本情况与系统发育位置,为江西铅山红芽芋的物种辨别、天然种群遗传多样性与功能基因组学提供前期研究铺垫。

江西槟榔芋疫病病原菌鉴定及室内药剂筛选

为明确江西省九江市槟榔芋疫病的病原菌并有针对性地对该病害进行防治,本研究采集具有典型芋疫病症状的芋叶片和叶柄对病原菌进行分离纯化,结合形态学观察、致病性测定及基于ITS、Ypt1和β-tubulin基因联合系统发育分析对该病原菌进行鉴定,并测定了其对7种杀菌剂的敏感性。根据病原菌菌落、卵孢子形态等特征初步鉴定为芋疫霉Phytophthora colocasiae,基于多基因联合系统发育分析结果与形态学结果一致,确定该病原菌为芋疫霉。室内药剂筛选结果表明,23.4%双炔酰菌胺SC对菌丝生长抑制效果最好,EC_(50)为0.006μg/mL;96.8%烯酰吗啉TC、0.3%丁子香酚SL、20%氟吗啉WP、500 g/L氟啶胺SC、97%霜脲氰TC对菌丝生长的抑制效果也较好,EC_(50)介于0.094~1.272μg/mL之间,研究结果可为槟榔芋疫病的化学防治提供参考。

红芽芋及荔浦芋叶绿体基因组测序及比较分析

为明确红芽芋、荔浦芋叶绿体基因组的基本特征及其差异,本研究以江西铅山红芽芋和广西荔浦芋为研究对象,采用改良CTAB法提取2种芋新鲜叶片基因组DNA,利用高通量测序技术获得2种芋全长叶绿体基因组序列,并进行SSR检测、序列比对、遗传多样性滑窗分析和系统发育树构建分析。结果表明,红芽芋和荔浦芋的叶绿体基因组全长分别为162478 bp和162453 bp,均呈现典型的环状双链四分体结构,二者均注释到131个基因,包括蛋白编码基因86个,tRNA基因37个,rRNA基因8个,其中具有2个拷贝的基因18个,具有内含子的基因23个。简单重复序列(SSR)位点分析发现,红芽芋和荔浦芋基因组序列分别含有130和124个SSR,二者SSR序列中A/T碱基含量分别占63.08%和61.29%,基于其SSR位点分析开发出15条具有多态性的SSR引物。红芽芋和荔浦芋叶绿体基因组共检测到236个SNP位点,26.7%的SNP位点发生在编码区,其中有25个基因发生错位突变,这些基因的变异可能促进了2种芋之间的性状变异。与天南星亚科已公布的13个属代表植物叶绿体基因组进行比较,其基因结构、种类和数量都比较保守,trnS-trnG和ndhF-rpl32分别是SSC区和LSC区的最高变异位点。最大似然法构建的系统发育树显示芋属与皂七属亲缘关系最为相近,与斑龙芋属亲缘关系较远。本研究结果将为我国特色芋种质资源开发利用、遗传多样性评价和系统发育分析奠定理论基础。

芋扩展蛋白基因家族的全基因组鉴定及其在球茎膨大中的表达分析

扩展蛋白是一类具有促进细胞壁伸长和增加细胞壁柔韧性作用的蛋白。本研究基于芋全基因组范围鉴定扩展蛋白基因,并分析该家族基因结构特征、系统发育及其在芋球茎膨大中的表达模式。结果表明,芋全基因组包含31个扩展蛋白基因,分布在芋8条染色体及2个重叠群上。在芋和拟南芥、水稻、马铃薯等4个物种中的扩展蛋白基因进行系统发育树分析中,所有的扩展蛋白基因划分到4个亚家族中,其中EXPA有23个,EXPB有5个,EXLA有1个,EXLB有2个。共线性分析表明,芋和拟南芥之间存在7对共线性扩展蛋白基因。基因表达分析发现,芋扩展蛋白基因家族不同成员在不同组织中有不同的表达模式,其中CeEXPA 2、CeEXPA 11、CeEXPA 13、CeEXPA 23和CeEXLA 1等5个基因在球茎膨大过程中高表达且差异显著,表明它们可能对芋球茎的膨大发挥了重要的调控作用。本研究结果将有助于了解芋扩展蛋白基因家族的特征及其在球茎膨大中的分子功能,为揭示芋球茎生长发育的分子调控机理提供理论依据。

基于气相离子迁移谱技术的芋头产地鉴别方法

分析不同产地芋头挥发性有机化合物组成差异,并构建芋头产地溯源的可视化指纹图谱。采用气相离子迁移谱法对不同产地芋头的挥发性物质进行测定,结合主成分分析(PCA)实现样品产地的快速区分,进一步筛选芋头中差异挥发性物质。结果表明,在不同产地芋头中,共检测到45个信号峰,鉴定出26种化合物,包括单体和部分化合物的二聚体。主成分分析将芋头分为3类,其中,靖江香沙芋和奉化芋头的挥发性物质较为相似,而与荔浦芋头、沙沟芋头差别很大。异丙醇、2-甲基-乙酸丁酯、辛酸甲酯是区别不同芋头的特征标记物。该方法直观、快速,为地方特色芋头的区分提供了新方法和技术支持。

芋球茎贮藏物质积累与食味品质形成的关系

为探明芋球茎贮藏物质积累与食味品质的关系,本研究以靖江‘香沙芋’和泰兴‘香荷芋’为实验材料,运用树脂半薄切片、顶空气相色谱-质谱联用等方法比较了两芋品种球茎中淀粉和挥发性成分等物质的积累情况,并进行食味品质鉴定。结果表明,‘香荷芋’含水量较高,而总淀粉含量和直链淀粉含量较低。‘香荷芋’的淀粉平均粒径明显小于‘香沙芋’,前者淀粉粒度分布峰值为1~2μm,后者为2~3μm。‘香荷芋’中主要检测到10种挥发性成分,其中芳樟醇比例高达48.82%,‘香沙芋’中检测到28种,但大部分属于无色无味的物质。食味品质分析发现‘香荷芋’气味得分较高,但粘度、硬度、外观、口感得分都较低。综上所述‘,香荷芋’挥发性香味物质含量多,使得其气味评分高;‘香沙芋’含水量低、淀粉和直链淀粉含量高、淀粉粒径大使得其硬度、口感和和综合得分高。该结果说明芋球茎含水量、总淀粉含量、直链淀粉含量、挥发性成分含量等指标都影响食味品质的形成。