Species-specific regulatory pathways of small RNAs play sophisticated roles in flower development in Dimocarpus longan Lour.

Flower development plays vital role in horticultural plants.Post-transcriptional regulation via small RNAs is important for plant flower development.To uncover post-transcriptional regulatory networks during the flower development in Dimocarpus longan Lour.‘Shixia’,an economically important fruit crop in subtropical regions,we collected and analyzed sRNA deep-sequencing datasets and degradome libraries Apart from identifying miRNAs and phased siRNA generating loci(PHAS loci),120 hairpin loci,producing abundant sRNAs,were identified by in-house protocols.Our results suggested that 56 miRNA-target pairs,2221-nt-PHAS loci,and 111 hairpin loci are involved in posttranscriptional gene silencing during longan reproductive development.Lineage-specific or species-specific post-transcriptional regulatory modules have been unveiled,including miR482-PHAS and miRN15.miR482-PHAS might be involved in longan flower development beyond their conserved roles in plant defense,and miRN15 is a novel miRNA likely associated with a hairpin locus(HPL-056)to regulate strigolactone receptor gene DWARF14(D14)and the biogenesis of phasiRNAs from D14.These small RNAs are enriched in flower buds,suggesting they are likely involved in post-transcriptional regulatory networks essential for longan flower development via the strigolactone signaling pathway.

Phytochemical constituents and biological activities of longan (Dimocarpus longan Lour.) fruit: a review

Longan(Dimocarpus longan Lour.),as an edible fruit and traditional Chinese medicine,has been consumed for thousands of years.Longan pulp has abundant nutritional phytochemicals such as protein,carbohydrate,vitamin C,polysaccharides,polyphenols,which shows multiple biological activities including antioxidant,immunomodulatory and antitumor effects.Longan pericarp also demonstrates biological activities because of its rich content of polysaccharides and polyphenols.This review summarizes the bioactive compounds and bioactivities of longan pulp and aims to provide comprehensive information for future development of longan as a functional health food.

Genome-wide analysis of the CCCH zinc finger family in longan:Characteristic identification and expression profiles in Dimocarpus longan Lour

CCCH(C3 H) Zinc finger(Znf) transcription factors(TFs), as a novel type of Znf gene, regulate the expression of genes by binding to their mRNAs and play important roles in plant growth and development and abiotic stress resistance.Longan(Dimocarpous longan) is a tropical/subtropical fruit tree of great economic importance in Southeast Asia.However, genomic information on C3 H and their functions in longan are still unknown. In this study, a comprehensive analysis of the longan C3 H(DlC3 H) gene family was carried out. A total of 49 DlC3 H genes in three clades were identified from the longan genome database. Characteristics of the genes were analyzed with respect to gene structure,motif composition, phylogenetic tree and potential functions. The analysis of alternative splicing(AS) events suggested that AS events in DlC3 H genes were related to the transformation from longan non-embryonic to embryonic cultures.Promoter analysis indicated that most of the DlC3 H genes included cis-acting elements associated with hormones and stresses responses. Quantitative real-time PCR(qRT-PCR) analysis indicated that 26 of the 49 DlC3 Hs, which possess methyl jasmonate(MeJA) and abscisic acid(ABA) responsive cis-acting elements, showed differential expression patterns under treatment with ABA, MeJA and their endogenous inhibitors, suggesting that DlC3 Hs might be involved in the ABA and MeJA signaling pathways. The expression profiles of 17 of the 49 DlC3 Hs in non-embryonic callus and three tissues of embryonic cultures showed that only five of the 17 DlC3 Hs had the same expression trends as the FPKM trends in transcriptome data;the expression levels of DlC3 H07/14/16/36/49 in embryogenic callus and DlC3 H04/38 in globular embryos were high, suggesting that they have different functions in embryonic development. Further, we verified that DlC3 H01/03/05/11/19/39 were regulated by sRNAs by a modified 5’ RLM-RACE method. This study provides the first systematic analysis of C3 H genes in longan, and found that C3 H genes may be involved in hormone and stress responses, and somatic embryogenesis. Our preliminary investigation may provide clues to further studies on the characteristics and functions of this family in longan.

Optimization of Extraction Process for Longan(Dimocarpus longan Lour.)Leaves Using Multi-indicator Comprehensive Evaluation Method

[Objectives]This study aimed to optimize the extraction process of longan(Dimocarpus longan Lour.)leaves by multi-indicator comprehensive evaluation method.[Methods]The contents of gallic acid ethyl ester,astragalin,quercetin,luteolin and kaempferol in longan leaves were determined simultaneously by HPLC.Using the multi-indicator comprehensive evaluation method,the methanol concentration,solid to liquid ratio and extraction time for extraction of longan leaves were optimized by orthogonal test.[Results]The optimal extraction process of longan leaves are as follows:methanol concentration of 100%,solid to liquid ratio of 1∶5,and extraction time of 50 min.[Conclusions]The optimized process is simple and feasible,and it can be used to determine the contents of different ingredients in longan leaves.

A Preliminary Analysis of the Relationship between Longan Canopy Temperature and Air Temperature during Overwintering Period

[Objective] The study aimed to provide supports for developing chilling and freezing injury monitoring and disaster damage assessment of longan(Dimocarpus Longan Lour.).[Method] Based on field observation data,the relationships between longan canopy temperature and air temperature under different weather types(sunny,cloudy to sunny,cloudy,rainy,radiation chilling injury and advection chilling injury)in 2007-2008 winter were analyzed.[Result] Diurnal variations of longan canopy temperature under sunny and radiation chilling injury weather conditions were most dramatic,followed with those under cloudy to sunny condition,while variations under cloudy,rainy and advection chilling injury conditions were mild.Diurnal variations of orchard air temperature were also closely related to weather types.By using linear and curvilinear regression methods,the relationship models between longan canopy temperature and observation station air temperature were established.The models for cloudy,rainy and advection chilling injury had better effects than those for sunny,cloudy to sunny and radiation chilling injury;the models for night were better than those for daytime and the whole day.[Conclusion] To some extent,applying the relationship models between longan canopy temperature and observation station air temperature could make up the shortcoming of meteorological data which were higher than the real values.

Indicator Analysis of Off-season Longan Flower Reversal

[Objectives] This study was conducted to analyze the internal causes of flower reversal in Longan （ Dimocarpus longan Lour.） trees. [Methods] With flowering trees and flower reversal trees as experimental materials, the variations in sugar and starch in mature leaves, tender leaves, mature branches, twigs and terminal buds after flower forcing were analyzed. [Results] During flowering process, sugar content showed the greatest difference between flowering and flower reversal trees, and the difference was the greatest in mature leaves. Trees with mature leaves having a sugar content above 44.71 mg/g were found to be more prone to flowering, while those with leaf sugar content lower than 27.80 mg/g were susceptible to flower reversal. In addition, longan trees with a higher sugar content in tender leaves were not prone to flower reversal. [Conclusions] In future, whether off-season flower forcing can be performed on longan trees could be judged through the detection of tree leaves, which is of great significance to prevention of flower reversal in off-season longan production.

反季节龙眼不同发育时期果实脱落响应差异与果实呼吸耗氧量的关系

反季节龙眼在果实发育过程中落果现象非常严重。前期的研究表明,果实中碳水化合物水平的下降与果实脱落密切相关。为了进一步了解不同发育时期果实的碳水化合物代谢变化与脱落差异的关系,以反季节储良龙眼为试材,果穗从树上剪下后迅速插入清水中并去除叶片诱发果实脱落,通过测定胁迫处理下3个发育时期果实落果率、呼吸耗氧量、果柄分离力变化,检测不同光强对3个时期果实呼吸的影响,发现不同发育时期龙眼果实对脱落的敏感性不同。饥饿胁迫迅速启动了各发育时期果实的脱落响应,但3个时期果实脱落响应差异较大,幼果期果实对饥饿胁迫的响应较中期和后期延后2 d,果实发育中期和后期脱落响应更剧烈;幼果期果实呼吸耗氧量较低,而果实发育中期和后期呼吸耗氧量较高;随着光强的增加,仅幼果期果实呼吸作用未造成氧气的下降,还出现了氧气的释放。幼果期果柄分离力下降也较果实发育中期和后期缓慢。因此,本研究结果表明,在胁迫条件下果实呼吸作用与果实脱落密切相关,果实呼吸强度越强;在饥饿胁迫条件下果柄分离力下降越快,果实脱落越快。龙眼幼果期可通过果皮进行光合作用部分代偿呼吸对碳水化合物的消耗。幼果期饥饿胁迫下落果慢的主要原因是低的呼吸消耗和具有一定的光合能力,因此,龙眼幼果是一个部分自养的器官。

龙眼DlSWC5基因克隆、亚细胞定位及表达特性分析

【目的】克隆龙眼ATP依赖的染色质重塑相关基因DlSWC5,并分析其亚细胞定位及表达特性,为研究该基因在龙眼生长发育及早期体胚发生过程中的调控功能提供理论依据。【方法】以龙眼品种红核子体胚发生早期胚性愈伤组织(Embryogenic callus,EC)为试验材料,结合龙眼三代基因组数据,采用RT-PCR克隆DlSWC5基因cDNA序列,对其进行生物信息学分析,基于STRING数据库预测DlSWC5蛋白互作调控网络,并通过激光扫描共聚焦试验验证蛋白亚细胞定位情况。分别利用转录组测序技术和实时荧光定量PCR检测DlSWC5基因在体胚发生早期和不同组织及不同浓度吲哚乙酸(IAA)处理下EC中的表达情况。【结果】成功克隆出DlSWC5基因的cDNA序列,与基因组中DlSWC5基因CDS序列相似性达99.80%,共编码340个氨基酸残基,该蛋白不含信号肽和跨膜结构,共含有44个磷酸化位点,分子式为C_(1622)H_(2599)N_(459)O_(542)S_(8),相对分子质量为37458.71 Da,理论等电点(pI)为5.74。DlSWC5蛋白结构的预测结果显示,其二级结构包含48.53%的α-螺旋、40.29%的无规则卷曲、6.47%的延伸链、4.71%的β-转角,其三级结构与漾濞槭同源蛋白高度一致。蛋白互作预测结果显示,DlSWC5蛋白与PIE1、SWC2、SWC4等10个蛋白互作。亚细胞定位显示DlSWC5蛋白定位于细胞核中。DlSWC5基因的相对表达量在早期体细胞胚胎发生中无明显差异。DlSWC5基因在体胚发生早期及种子、根、茎、叶、花蕾、花、果皮、果肉、幼果等组织中均检测到表达,与对照组(0 mg/L)相比,DlSWC5基因的相对表达量在0.5、1.0、1.5和2.0 mg/L IAA处理24 h后均显著降低(P<0.05)。【结论】成功克隆了DlSWC5基因,其编码蛋白为不稳定的亲水性蛋白,呈酸性,定位于细胞核;DlSWC5基因在龙眼体胚发生早期EC阶段被IAA诱导下调表达。

龙眼拟茎点霉(Phomopsis longanae Chi)的生物学特性研究

龙眼拟茎点霉（Phomopsis longanae Chi）是导致采后龙眼果实腐烂的主要病原菌,研究了温度、湿度、pH值、光照、营养、越冬越夏等条件对龙眼拟茎点霉菌丝生长、产孢和孢子萌发的影响。结果表明：龙眼拟茎点霉生长适温为25℃,最适pH6~7;产生子座的最适温度为22~25℃,最适pH6,还需要一定的光照;以蔗糖、葡萄糖、可溶性淀粉和D-果糖为碳源和以蛋白胨、硫酸铵和硝酸铵为氮源的培养基最有利于龙眼拟茎点霉的生长发育;分生孢子的大量萌发要求高湿度和营养刺激,最适萌发温度25~28℃,最适pH6~7,在分生孢子器中,分生孢子的存活时间长于1年;该菌菌丝的致死温度为59℃（10 min）,分生孢子为50℃（10 min）。

基于HPLC指纹图谱和多组分含量测定结合化学模式识别研究的龙眼核质量评价

目的 建立龙眼Dimocarpus longan Lour.核HPLC指纹图谱及含量测定方法。方法 建立13批龙眼核指纹图谱,结合指纹图谱相似度评价及化学模式识别研究对龙眼核质量进行分析,并测定其中没食子酸、柯里拉京、鞣花酸的含量。结果 13批龙眼核样品相似度在0.919~0.999,标定共有峰11个,并指认出3种成分;聚类分析(HCA)将13批龙眼核聚为3类,主成分分析(PCA)提取到3个主成分,正交偏最小二乘判别分析(OPLS-DA)筛选出5个差异性成分;定量分析得到13批龙眼核中没食子酸、柯里拉京金、鞣花酸的含量范围依次为0.65~1.8 mg·g^(-1)、3.02~5.46 mg·g^(-1)、3.18~4.49 mg·g^(-1)。结论 建立的HPLC指纹图谱及含量测定方法重复性、稳定性良好,能够为龙眼核质量评价提供参考。

龙眼叶黄酮类有效组分配伍体外降糖作用的研究

基于体外降糖活性优选龙眼(Dimocarpus longan Lour.)叶黄酮有效成分山柰酚、槲皮素、槲皮苷的配伍比例。建立HepG2细胞胰岛素抵抗模型,分别考察不同浓度的胰岛素及不同诱导时间对胰岛素抵抗模型的诱导情况,并以山柰酚、槲皮素、槲皮苷三者用量为考察因素,以对胰岛素抵抗的HepG2细胞葡萄糖消耗率为指标,运用星点设计-效应面法(CCD-RSM)优化三者配伍比例。结果表明,建立HepG2细胞胰岛素抵抗模型的最佳条件为1×10^(-7)mol/L胰岛素诱导24 h;山柰酚、槲皮素、槲皮苷体外降糖最佳配伍比例为山柰酚105.0μmol/L、槲皮素78.9μmol/L、槲皮苷194.0μmol/L,配伍比例优化试验拟合方程为Y=20.94+1.55×A-0.44×B-2.05×C-0.66×AB+1.68×AC-1.53×A^(2)-3.2×B^(2)-2.68×C^(2)。山柰酚、槲皮素、槲皮苷3种单体的联合使用对HepG2细胞的葡萄糖消耗率预测值是21.52%,实验值是(21.05±0.52)%。优选出的最佳配伍比例的龙眼叶黄酮类有效组分,对HepG2细胞的葡萄糖消耗率最高,降糖活性较好,可为龙眼叶治疗糖尿病的有效性提供科学方法与实验依据。

不同疏花方式对龙眼开花坐果及果实品质的影响

通过对龙眼不同疏花方式处理组和对照组的观察和数据分析,探究了疏花方式不同对开花坐果及果实品质的影响。结果显示:完全疏花组的开花数量和果实数量显著减少,坐果率降低,但果实品质较好;部分疏花组保留了适量的花蕾,开花数量相对较多,坐果率相对较高,保持一定的产量和较好的果实品质;延迟疏花组保留了所有花蕾,开花数量最多,但坐果率受到影响,果实品质相对较差。研究结果可为龙眼种植者提供一定的参考和指导,为龙眼品种改良和栽培技术的发展提供理论基础和实践指导。

基于UPLC-Q-TOF-MS/MS技术的龙眼叶入血成分分析

目的:基于血清药物化学方法,对龙眼叶乙酸乙酯部位进行入血成分分析,探讨龙眼叶的药效物质基础。方法:采用UPLC-Q-TOF-MS/MS高分辨液质联用法,对乙酸乙酯部位及给药后大鼠血清样品进行分离后在负离子检测模式下进行总离子检测、二级碎片离子扫描,初步鉴定龙眼叶提取物在大鼠血中原型成分及代谢成分。结果:从含药血清中初步鉴定了共17个药源性成分,其中有1个原型成分曾从龙眼叶的提取物中得到分离,其余多为原型成分的苷元代谢产物。结论:初步确定了没食子酸乙酯、槲皮素、槲皮苷、木犀草素等成分可能为龙眼叶降血糖活性的物质基础。

Classification of Thai Honey Origins by Their Mineral Contents and Color Parameters

Honey is a product of the elaboration of flower nectar by bees. The general features and elemental composition of honey depend on its botanical origin. In this study, five color parameters （L＊： lightness, a＊： red color, b＊： yellow color, C＊ab and hab） and five elements （Na, K, Mg, Ca and Zn） were determined and related with 91 Thai honey samples. The origins of four botanic types of （1） longan flower （Dimocarpus sp.）, （2） lynchee flower （Litchi sp.）, （3） sunflower （Helianthus sp.） and （4） wild flower （Eupatorium sp.） using principle component analysis （PCA）. The results showed that five color parameters and five metal contents related with the Thai botanic origins of the honeys using principle component analysis （PCA）. Six major indicators of PC i （variance is 44.33%） from three color parameters are L＊（-0.926）, a＊（0.927） and hue or hab （0.824） and from three metal contents are K（0.833）, Ca（0.816） and Mg（0.595）. Two minor indicators of PC2 （variance is 21.58%） from color parameters are b＊（-0.934） and Chroma or C＊ab （-0.834）. Two indicators of PC3 （%variance is 12.47%） from contents of Na （-0.722） and Zn （0.704）. Thai Lynchee （C） flower honeys classified using both six parameters in PC 1 and two color parameters in PC2. Thai longan flower （G） honeys classified using the contents of Zn and Na in PC3 parameters. Thai sunflower （S） honeys classified using two color parameters in PC2. Thai wild flower （W） honeys classified using the metal contents of K in PC 1 parameter, Zn and Na in PC3 parameters.

茉莉酸甲酯处理对采后龙眼果皮褐变的影响

本实验以10、50、100μmol/L MeJA处理‘储良’龙眼(Dimocarpus longan Lour.)果实,研究其对(20±1)℃、85%相对湿度贮藏条件下的龙眼果皮褐变的影响。结果表明:与对照组相比,MeJA处理可有效降低‘储良’龙眼采后果皮褐变指数、果皮细胞膜透性和果肉自溶指数,而且MeJA浓度越高,抑制效果越明显。与对照组相比,100μmol/L MeJA处理龙眼果实能够保持较高的果皮总酚和类黄酮含量,降低果皮多酚氧化酶活力,有效提高果皮过氧化物酶和苯丙氨酸解氨酶活力;另一方面,MeJA处理可以有效降低超氧阴离子自由基产生速率以及H2O2含量,有效增强抗氧化酶(超氧化物歧化酶、过氧化氢酶)活力以及1,1-二苯基-2-三硝基苯肼自由基清除能力,提高果皮抗氧化能力。综上,100μmol/L MeJA处理可抑制龙眼果实采后部分酚类物质代谢,增强其抗氧化能力,从而延缓龙眼采后果皮褐变的发生,延长贮藏保鲜期。

基于UHPLC-Q-Orbitrap MS技术的龙眼叶乙酸乙酯部位体内代谢研究

目的 采用超高效液相色谱-四级杆/静电场轨道阱高分辨质谱(UHPLC-Q-Orbitrap MS)技术研究龙眼叶乙酸乙酯在大鼠体内的代谢产物。方法 大鼠灌胃给予龙眼叶乙酸乙酯部位后,收集其尿液、血浆以及粪便样品。样品前处理后,采用Waters HSS T3色谱柱(100 mm×2.1 mm, 1.8μm);流动相为0.1%-甲酸水溶液(A)和含0.1%甲酸的100%甲醇(B),梯度洗脱。在正、负离子模式下采集样品数据分析。结果 从大鼠尿液、血清及粪便样品中共鉴定出包括原型成分在内的44个代谢产物,其中分别从尿液、血清及粪便样品中检测到10、20、26个。主要代谢途径有硫酸化、氧化、葡萄糖醛酸化等。结论 通过液质联用技术分析了龙眼叶乙酸乙酯部位在大鼠体内的代谢产物,为龙眼叶药效物质基础的阐明提供了一定的参考依据。

龙眼DlSWI3、DlARP、DlSWC和DlBSH亚基的分子特性及其在体胚发生早期的基因表达模式

[目的]探究龙眼Switching 3(SWI3)、Actin-related protein(ARP)、SWR1 complex(SWC)和Bushy(BSH)亚基的潜在功能。[方法]利用生物信息学分析其分子进化特性,用转录组和实时荧光定量PCR(real-time quantitative PCR,q RT-PCR)分析基因在体胚发生早期胚性愈伤组织(embryogenic callus,EC)、不完全胚性紧实结构(incomplete embryogenic compact structures,ICpEC)和球形胚(globular embryos,GE)阶段及在EC中响应激素的表达模式。[结果]龙眼含4个DlS WI3、10个DlA RP、4个DlS WC和1个DlB SH蛋白,它们大多不稳定且主要定位在细胞核、细胞质和叶绿体上。DlS WI3和DlA RP家族扩张受基因复制驱动,而4个DlSWC和1个DlBSH基因由单拷贝产生。DlS WI3、DlA RP、DlS WC和DlB SH结构域各异,进化上与拟南芥、水稻和甜橙一致。启动子预测表明,DlSWI3、DlARP、DlSWC和DlBSH亚基的基因含多种激素、胁迫、光和生长发育相关元件。q RT-PCR分析显示,从EC分化到GE,DlARP4、DlARP5-3、DlARP9和DlBSH逐渐下调,而DlSWI3C、DlARP8和DlSWC2逐渐上调,且DlSWI3D在GE特异高表达;DlARP5-3、DlSWC2、DlSWI3D和DlBSH在脱落酸(abscisic acid,ABA)处理12 h时被显著上调,DlARP4、DlARP5-3、DlSWC2、DlSWI3D和DlBSH在茉莉酸甲酯(methyl jasmonate,MeJ A)处理4 h时被显著上调,5个基因被赤霉素(gibberellin A3,GA3)或水杨酸(salicylic acid,SA)下调,DlARP4被ABA下调。[结论]DlS WI3、DlA RP、DlS WC、DlB SH亚基在进化上高度保守,它们可能通过响应激素诱导影响染色质重塑活性,调控龙眼早期体胚发育。

广西龙眼古树资源调查

龙眼是广西主栽果树种类之一,栽培历史悠久,为了解广西龙眼古树资源情况,对广西龙眼产区的十多个市、县(区)开展龙眼古树资源调查。调查结果表明:调查区域均有龙眼古树分布,其中树龄最大达900年;龙眼古树树龄与胸围显著相关,树高与冠幅显著相关。由于缺乏管理,当前广西龙眼古树资源保护仍存在较多问题,龙眼古树生存状况堪忧,一些古树长势衰弱、濒临死亡。针对存在问题,提出了加强龙眼古树资源保护的建议。

龙眼采后果肉生理生化变化研究

观察了石硖、储良龙眼 (DimocarpuslonganLour.cv .Shixia&Chuliang)在 4和 2 0℃条件下的贮藏效果 ,测定了果肉生理生化变化 .结果表明 :低温能够大大延缓果肉的衰败进程 ,石硖明显比储良耐贮藏 ,储良 33d、石硖 5 0d时果肉几乎全部流汁 .随着贮期延长 ,整果和果肉质量损失率和霉烂率增加 ,好果率降低 ,果皮褐变 .生理生化变化表现为丙二醛 (MDA)含量逐渐增加 ;超氧化物岐化酶 (SOD)活性 (4和 2 0℃ )变化不明显 ,相对较稳定 ,储良活性比石硖低 ;过氧化物酶 (POD)呈上升趋势 ,2 0℃下活性明显比 4℃下高 ;过氧化氢酶 (CAT)活性呈显著下降趋势 。

HPLC法测定龙眼肉中的几种核苷类物质

目的:建立HPLC法测定龙眼肉中腺苷、尿苷和腺嘌呤的含量。方法:采用NucleosilC18反相柱(250mm×4mm),以0.01mol/L的KH2PO4:乙腈=92:8为流动相,流速为1ml/min,检测波长为254nm,柱温40℃,用外标法,测定了龙眼肉中腺苷、尿苷和腺嘌呤的含量。结果与结论:分析中选取腺苷的线性范围0.1～20.0μg/ml,相关系数r=0.9999,回收率94.59%,RSD3.10%;尿苷的线性范围0.05～10.0μg/ml,相关系数r=1,回收率97.49%,RSD4.50%;腺嘌呤线性范围0.05～10.0μg/ml,相关系数r=1,回收率103.11%,RSD1.90%。该方法简便快速,线性关系良好。