荔枝(Litchichinensis Sonn.)花芽分化过程的细胞超微结构观察

应用透射电镜对荔枝茎尖生长锥由营养生长转变为生殖生长过程的细胞超微结构进行观察 ,结果表明 ,荔枝圆锥花序发育前期细胞排列紧密 ,细胞质浓 ,液泡小 ,细胞核大并占据着细胞大部分位置 ;细胞质内有丰富的线粒体和膜状系统 ,整个膜状系统内连细胞核 ,外接胞间连丝 ;细胞内囊泡大量形成、融合与迁移 .这些均显示出细胞内结构物质在此期经历着活跃的周转和合成代谢 ,细胞内部在基因的控制下有序地表达 ,为发育过程质的转变积累足够的信息和物质 .生长锥由尖长变为宽平的花序原基时 ,其组织细胞结构也发生了相应的变化 ,最显著的特征是在此过程中 ,有一些细胞发生了程序性死亡 .正在凋亡的细胞核染色质固缩、趋边、核膜破裂或皱缩、细胞质囊泡化并将降解的物质运到周围细胞 。

荔枝(Litchi chinensis)细胞培养的初步研究

用荔枝无菌苗作为外植体，在添加了ＡｇＮＯ３或蜂王浆的含１ｍｇ／Ｌ６－ＢＡ和１ｍｇ２，４－Ｄ的ＭＳ培养基中诱导出愈伤组织，在同样培养基中继代培养，得到了稳定的培养细胞系．这种细胞在含１ｍｇ／Ｌ６－ＢＡ＋２ｍｇ／Ｌ２，４－Ｄ的ＭＳ培养基中悬浮培养，培养周期２５ｄ．结果表明：维生素Ｃ和ＮａＣｌ在悬浮培养中有良好的防褐作用．

Cloning and Sequence Analysis of Expansin Genes from Litchi chinensis Fruit

Using PCR degenerate primers, designed with reference to the sequences of the conserved amino acids of known expansins, to amplify cDNA fragments in litchi fruit by RT-PCR, two different cDNA fragments , named as Lc-Exp1 and Lc-Exp2 , were cloned. Lc-Exp1 and Lc-Exp2 was respectively composed of 531 bp encoding 177 amino acids and 537 bp encoding 179 amino acids. Eight cysteine residues and three tryp-tophan residues, which is supposed to be the characteristics of expansins, are conserved in both Lc-Exp1 and c-Exp2. In addition, the homology between the two expansins is 71. 6% at nucleotide acid sequences and 76.3% at amino acid sequences. The homology of Lc-Exp1 with Fa-Exp2 or Pp-Exp1 was 92.7% or 92.1%, but that of Lc-Exp2 with Fa-Exp2 or Pp-Exp1 was only 77. 4% or 76.3% at amino acid sequences.

Molecular cloning,characterization and expression profile of the sucrose synthase gene family in Litchi chinensis

Sucrose synthase(SUS,EC 2.4.1.13)is widely considered as a key enzyme involved in plant sucrose metabolism,and the gene family encoding different SUS isozymes has been identified and characterized in several plant species.However,to date scant information about the SUS genes is available in Litchi chinensis Sonn.Here,we identified five SUS genes in litchi.These Lc SUSs shared high levels of similarity in both nucleotide and amino acid sequences.Their gene structure,phylogenetic relationships,and expression profiles were characterized.Gene structure analysis indicated that the Lc SUSs have similar exon-intron structures.Phylogenetic analysis revealed that the five members could be classified into three groups(LcSUS1 and LcSUS2 in SUSⅡ,LcSUS4 and LcSUS5 in SUSⅢ,and LcSUS3 in SUSⅠ),demonstrating evolutionary conservation in the SUS family across litchi and other plant species.The expression levels of Lc SUSs were investigated via real-time PCR in various tissues and different developmental stages of aril.For tissues and organs,Lc SUSs exhibited distinct but partially redundant expression profiles in litchi,being predominantly expressed in young leaves(sink).During aril development,the expression pattern of LcSUS1 was consistent with the trend of sugar accumulation,indicating it may play important roles in determination of sink strength in aril.Moreover,transcript levels of LcSUS2,LcSUS4,and LcSUS5 varied between cultivars with different hexose/sucrose ratios,which may regulate the sugar composition in aril.Our results provide insights into physiological functions of SUS genes in litchi,especially roles in regulating sugar accumulation in aril.

Construction of high-density SNP genetic maps and QTL mapping for dwarf-related traits in Litchi chinensis Sonn

Litchi chinensis Sonn is widely cultivated in subtropical regions and has an important economic value.A high-density genetic map is a valuable tool for mapping quantitative trait loci(QTL)and marker-assisted breeding programs.In this study,a single nucleotide polymorphism(SNP)-based high-density linkage map was constructed by a genotyping-by-sequencing(GBS)protocol using an F1 population of 178 progenies between two commercial litchi cultivars,‘Ziniangxi’(dwarf)and‘Feizixiao’(vigorous).The genetic map consisted of 3027 SNP markers with a total length of 1711.97 cM in 15 linkage groups(LGs)and an average marker distance of 0.57 cM.Based on this high-density linkage map and three years of phenotyping,a total of 37 QTLs were detected for eight dwarf-related traits,including length of new branch(LNB),diameter of new branch(DNB),length of common petiole(LCP),diameter of common petiole(DCP),length of internode(LI),length of single leaf(LSL),width of single leaf(WSL),and plant height(PH).These QTLs could explain 8.0 to 14.7%(mean=9.7%)of the phenotypic variation.Among them,several QTL clusters were observed,particularly on LG04 and LG11,which might show enrichment for genes regulating the dwarf-related traits in litchi.There were 126 candidate genes identified within the QTL regions,55 of which are differentially expressed genes by RNA-seq analysis between‘Ziniangxi’and‘Feizixiao’.These DEGs were found to participate in the regulation of cell development,material transportation,signal transduction,and plant morphogenesis,so they might play important roles in regulating plant dwarf-related traits.The high-density genetic map and QTLs identification related to dwarf traits can provide a valuable genetic resource and a basis for marker-assisted selection and genomic studies of litchi.

Agrobacterium tumefaciens-mediated transformation of embryogenic callus and CRISPR/Cas9-mediated genome editing in‘Feizixiao'litchi

Litchi(Litchi chinensis Sonn.)is a type of commercially prevalent subtropical and tropical fruit.Since litchi has a highly heterozygous genetic background and a long reproductive cycle,conventional breeding methods(such as hybridization)have limited ability to nurture new litchi cultivars.Here,an efficient and stable Agrobacterium tumefaciens-mediated genetic transformation of embryogenic callus was established in‘Feizixiao’litchi.Transgenic materials were verified using polymerase chain reaction(PCR)analysis,β-glucuronidase(GUS)assay,and green fluorescent protein(GFP)assay.To implement the technology of the Clustered Regularly Interspaced Short Palindromic Repeats(CRISPR)/associated protein 9(CRISPR/Cas9)technology in‘Feizixiao’litchi and verify the validity of these transformation systems,the litchi polyphenol oxidase gene(LcPPO,JF926153)was knocked out.Various categories of mutations,covering base insertions,deletions,and substitutions,were found in transgenic materials via sequence analysis.The transformation system achieved high feasibility and efficiency,and the system of CRISPR/Cas9 was successfully employed to edit genes in‘Feizixiao’litchi.This work provides an essential foundation for investigating the functions of genes and accelerating litchi genetic improvement.

Effects of saponin from the seed of Litchi chinensis Sonn on TGF-β1, FN and SOCS-1 in renal tubular epithelial cells under high glucose

目的:探讨荔枝核皂苷对高糖培养的人肾小管上皮细胞凋亡的影响.方法:用含30mmol/L葡萄糖的DMEM/F12培养基培养HKC细胞,加入或者不加入SLS干预.空白对照组加入等体积的含5.5mmol/L葡萄糖的DMEM/F12培养基.用ELISA检测细胞转化生长因子β1(TGF-β1)、纤维连接素(FN)的分泌,用Annexin V-FITC/PI双染法检测细胞凋亡,用Western blot检测细胞因子信号转导抑制因子-1(SOCS-1)的表达.结果:ELISA结果显示,SLS可以降低TGF-β1、FN的分泌,与30mmol/L的葡萄糖组相比具有显著性差异(P<0.01).Annexin V-FITC/PI双染法结果显示,与正常组比较,30mmol/L的葡萄糖组细胞凋亡明显增加(P<0.01);SLS各剂量组明显降低HKC细胞的凋亡(P<0.01),其SLS浓度与HKC细胞凋亡率呈负相关.Western blot结果显示:与空白对照组相比,30mmol/L葡萄糖组SOCS-1的蛋白表达量升高(P<0.01),SLS能显著降低SOCS-1蛋白的表达(P<0.01).结论:SLS可以降低HKC细胞TGF-β1、FN的分泌,并明显降低HKC细胞的凋亡,从而延缓与减轻肾小球硬化.

Synthesis of silver and gold nanoparticles from leaf of Litchi chinensis and its biological activities

Objective: To synthesize and isolate silver and gold nanoparticles from Litchi chinensis leaf methanolic extract, and to evaluate its comparative biological activities including muscles relaxant, analgesic, anti-inflammatory and antidiarrheal. Methods: The gold and silver nanoparticles were synthesized by dissolving methanolic extract in gold chloride and silver nitrate solution separately which were confirmed by colour change and UV-Vis spectroscopy, and pellets were collected through centrifugation. Biological activities of the extract were conducted on BALB/c mice through various standard methods and the data were subjected to One-way ANOVA. Results: The colorless gold chloride solution changed to purple soon after the addition of plant extract, demonstrating that the reaction took place and gold ions were reduced to gold nanoparticles, while colorless silver nitrate solution changed to light and dark brown that was indicative of silver nanoparticles. The muscles relaxant activity showed that silver nanoparticles were more effective than gold nanoparticles and methanolic extract in traction test. The analgesic activity showed that silver and gold nanoparticles showed highest percentage decrease in acetic acid induced writhing at the doses of 50, 100 and 150 mg/kg b.w. The highest anti-inflammatory activity was produced by gold nanoparticles followed by silver nanoparticles, while low activity was observed in methanolic leaf extract. Only the crude methanolic extract showed significant antidiarrheal activity as compared to the standard drug atropine sulphate, while antidiarrheal activities of gold and silver nanoparticles were non-significant. Conclusions: The present work concludes that isolated silver and gold nanoparticles from leaf methanolic extract shows strong muscles relaxant, analgesic and anti-inflammatory activities while crude methanolic extract possesses good antidiarrheal activity.

Study on Improving Litchi chinensis Production in Adverse Weather

Being sensitive to climatic conditions,the growth of Litchi chinensis（ lychee） can be greatly affected both in rainy warm winter which is conductive to winter shoot elongation,seriously affecting the formation of flower buds,and in rainy spring or cold late spring when lychee is flowering,seriously affecting pollination and fruit set. Thus,meteorological conditions play a decisive role in determining the distribution,growth and development,yield formation and quality of lychee. Based on meteorological conditions,some measures can be taken to reduce the impacts of adverse weather and take advantage of favorable weather conditions for the stable and high production of lychee. Therefore,the yield of lychee could be increased by improving the accuracy of weather forecasts and long-term cultivation and management techniques.

Expression of SAMDC Gene for Enhancing the Shelf Life for Improvement of Fruit Quality Using Biotechnological Approaches into Litchi (Litchi chinensis Sonn.) Cultivars

Polyamines play an important role in plant response to abiotic stress. S-adenosyl-1-methionine decarboxylase (SAMDC) is one of the key regulatory enzymes in the biosynthesis of polyamines. In order to better understand the effect of regulation of polyamine biosynthesis on the shelf life improvement of litchi fruit, SAMDC cDNA isolated from Datura stramonium cloned in pBI121 was introduced into litchi genome by means of Agrobacterium tumefaciens through zygote disc transformation. Transgene and its expression are confirmed by Southern and Northern blot analyses, respectively. Transgenic plants expressing Datura SAMDC produced 1.7- to 2.4-fold higher levels of spermidine and spermine than wildtype plants under normal environmental condition, which indicated that the transgenic litchi presented an enhanced polyamines synthesis compared to wildtype plants. Our results demonstrated clearly that increasing polyamine biosynthesis in plants may be a means of creating improved fruit shelf life germplasm.

荔枝VQ基因家族鉴定及其对非生物胁迫的响应

【目的】VQ蛋白是一类含有保守VQ基序(FxxhVQxhTG)的植物特异性蛋白,在植物生长发育和非生物胁迫应答中发挥重要作用。研究鉴定了荔枝VQ基因家族,并分析其在不同组织的表达模式及在低温、高温、干旱和盐胁迫下的应答,为后续研究其抗逆机制奠定了基础。【方法】用生物信息学方法在荔枝全基因组中鉴定LcVQ基因,并对其理化性质、亚细胞定位、基因结构和保守基序等进行分析;用MEGA 6.0软件构建系统发育树,分析荔枝、拟南芥和水稻VQ蛋白的系统发育关系;用实时荧光定量PCR(qRT-PCR)技术验证LcVQs对多种非生物胁迫的响应情况。【结果】荔枝中共鉴定获得可聚类为9个亚族的18个VQ基因(LcVQ1-18),依次分布在荔枝的11条染色体上,其编码蛋白的氨基酸数介于111~427之间,分子质量为12.48~45.49 kD;除LcVQ15和LcVQ17定位于细胞质之外,其余LcVQ蛋白均定位于细胞核。LcVQs启动子区域包含大量植物生长发育响应元件、激素响应元件及逆境响应元件。LcVQs的表达量在不同组织中具有明显差异,总体上分为普遍性表达和特异性表达。LcVQs可快速响应非生物胁迫,在低温、高温、干旱和盐胁迫处理3 h内分别有4,3,3,4个LcVQs明显上调表达。【结论】荔枝全基因组中有18个VQ家族成员,具有典型VQ保守结构域,能差异化响应多种非生物胁迫。

褐毛荔果实表型性状多样性研究

【目的】对云南省的褐毛荔种质资源果实表型性状进行遗传多样性分析,为褐毛荔种质资源的评价和利用提供依据。【方法】在褐毛荔分布较多的云南省10个县调查收集褐毛荔种质资源,采集成熟果实,评价果实的16个描述性性状和8个数量性状,并对资源进行表型多样性分析、聚类分析及主成分分析。【结果】共调查收集到93份褐毛荔资源,其中1份为焦核资源。褐毛荔种质资源果实表型遗传多样性较丰富,数量性状的遗传多样性均高于描述性性状,其中多样性指数最高的是种子质量,为2.98。变异系数最高的是果皮质量,为38.36%。果实的主要特征包括:形状为歪心形,果皮颜色淡红带微黄,果肩和果顶形状多为双肩斜和钝圆;龟裂片多为平滑,裂片峰多为毛尖,缝合线不明显,果肉颜色以蜡白色为主,果肉内膜褐色为无或少,种子形状以椭圆形为主,种皮颜色以浅褐色为主,肉质多为细嫩,香味、风味和涩味以无香、酸甜、微涩为主,种子多为大核。根据果实数量性状分析将93份荔枝资源分为5类,其中第Ⅱ类的可食率最高,为62.49%,第Ⅴ类的果实最大。根据果实描述性性状可将荔枝资源分为4类,其中第Ⅳ类的果实风味较好。主成分分析结果表明,前2个主成分主要体现了果实大小、果实可食率和可溶性固形物含量,依据主成分得分,筛选出2株优异单株。【结论】褐毛荔种质资源表型多样性较为丰富,可用于荔枝新种质创制和新品种选育。

荔枝种子发育过程中有机营养变化与萌发的关系

对比分析“妃子笑”和“紫娘喜”2个荔枝品种种子发育过程中的种子萌发规律,研究其淀粉、可溶性糖、有机酸和氨基酸组分及含量变化,探讨种子萌发与这些有机营养成分之间的相关性。结果表明,随着果实发育成熟,“妃子笑”荔枝种子萌发率呈先上升后下降的趋势,“紫娘喜”荔枝种子萌发率持续上升;盛花后45 d开始,“紫娘喜”荔枝种子萌发率持续高于“妃子笑”。果实发育过程中,“妃子笑”和“紫娘喜”荔枝种子中总可溶性糖含量分别呈“N”和“W”形变化,两者的淀粉含量均逐渐升高;果实完全成熟时,“紫娘喜”荔枝种子中淀粉含量显著高于“妃子笑”;两者种子中总有机酸含量均持续下降,但组分不同,“妃子笑”荔枝种子中含有8种有机酸,“紫娘喜”荔枝种子多了乳酸和琥珀酸。两个品种种子均含21种氨基酸,“妃子笑”荔枝种子总氨基酸含量呈先降低后上升至稳定水平的趋势;而“紫娘喜”种子中总氨基酸含量呈先短时降低后升高再降低的变化。相关性分析表明,两个荔枝品种的种子萌发率与淀粉含量呈极显著正相关,与总有机酸含量呈极显著负相关。说明淀粉和有机酸可能是影响种子发育与萌发的主要有机营养物质。

玉林市仙进奉荔枝高产栽培技术要点

为提升荔枝(Litchi chinensis Sonn.)果实的品质和产量,针对仙进奉荔枝在玉林市的物候期表现及果实变异的问题,主要从建园、种苗选择、秋梢培养、控梢促花、疏花保果、适时采摘、采后修剪及清园等方面描述其栽培技术要点,以期为仙进奉荔枝高质量生产提供参考。

含宁夏枸杞果提取物和荔枝果皮提取物眼部精华液的临床功效

探讨一款含宁夏枸杞果提取物和荔枝果皮提取物的眼部精华液的临床功效,选取了35例18~60岁健康受试者使用该产品,通过测试皮肤角质层水分含量、经皮水分流失量、眼角皱纹面积、眼底细纹面积、眼角皮肤紧致度F4值和眼角皮肤弹性R2值,同时结合消费者自我评估,进行人体功效评价试验,验证该产品具有保湿、屏障修护、抗皱、紧致功效。

荔枝提质增效丰产栽培技术研究

茂名盛产荔枝,在当地已有2000年以上的栽培历史,是广东主要的荔枝产地之一。该文结合当地实际,对荔枝的提质增效丰产栽培技术进行简单地总结,包括果园清理、间伐及回缩短截、高接换种、肥水管理、有害生物防治等,对当地荔枝的优质丰产有着一定的积极指导作用。

Effects of simulated acid rain on fertility of litchi

The regulatory role of calcium in fertility of pollen and pistil under simulated acid rain was investigated. The germination percentage of pollen treated with acid rain of pH 4.5 was 9.42% lower than that of control, and that of pH 3.5, pH 2.5 and pH 1.5 were 22.47%, 45.49% and 71.62%, respectively. Simultaneously, the injury character of pollen was obviously observed when flowers were treated with acid rain of pH 3.5. The difference in fruit setting rate between the female flower treated with acid rain of pH 4.0 and the control was significant at p 〈 0.05. Ca（NO3 ）5 of 0.2-0.4 mmol/L could promote pollen germination under the stress of acid rain. The beneficial function was reduced when calcium concentration surpassed 0.8 mmol/L. Spraying 2 mmol/L Ca（NO3 ）5 reduced the injury of acid rain to pistil and increased fruit-setting rate significantly. The physiological importance of calcium during pollen germination and pistil development was also discussed.

氨基酸对荔枝愈伤组织增殖及体胚发生体系优化的研究

【目的】探究氨基酸对荔枝愈伤组织增殖及体胚诱导的影响。【方法】采用L9(34)正交设计,分别在妃子笑荔枝愈伤组织继代及体胚诱导阶段添加不同氨基酸,比较其对愈伤组织增殖及体胚诱导的影响。【结果】在愈伤组织增殖阶段培养基中添加氨基酸均显著提高愈伤组织增殖率,最优处理为基础继代培养基添加0.1 g·L^(-1)γ-氨基丁酸+0.2 g·L^(-1)谷氨酰胺,愈伤组织增殖率为初始接种量的14.04倍。与对照相比,氨基酸处理组的胚性愈伤组织淡黄,颗粒较小,细胞呈椭圆形,细胞质浓厚,多为正在分裂状态;非胚性愈伤组织浅白,水渍严重,着生于顶部或者边缘,易于剔除。在愈伤组织增殖阶段培养基中添加γ-氨基丁酸、谷氨酰胺和丙氨酸对后续的体胚发生及萌发影响极显著(p<0.01),最优配方为基础继代培养基添加0.3 g·L^(-1)γ-氨基丁酸+0.4 g·L^(-1)谷氨酰胺+0.05 g·L^(-1)丙氨酸,每克愈伤组织可获得461个体胚,77株组培苗。在体胚发生阶段培养基中添加氨基酸,获得的体胚发生及萌发最优配方为基础诱导培养基添加0.3 g·L^(-1)γ-氨基丁酸+0.4 g·L^(-1)谷氨酰胺+0.05 g·L^(-1)丙氨酸,每克愈伤组织可获得270个体胚,72株组培苗。【结论】氨基酸可调节愈伤组织胚性,提高愈伤组织增殖率,促进体胚发生及萌发。

荔枝CDPK基因家族鉴定及其在霜疫病胁迫下的表达分析

【目的】鉴定荔枝(Litchi chinensis Sonn.)钙依赖蛋白激酶(CDPK)基因家族,并分析其在不同组织和荔枝霜疫病胁迫下的表达模式。【方法】基于荔枝基因组数据,利用生物信息学方法鉴定CDPK家族基因,并对其序列特征、基因结构、启动子顺式作用元件、染色体定位和进化关系等进行分析。依赖276份荔枝种质材料,筛选高抗霜疫病的优良荔枝品种。另外,通过RNA-Seq和qRT-PCR方法检测高抗病荔枝品种中LcCDPK基因家族成员在荔枝霜疫病胁迫处理下的表达模式。【结果】从276份荔枝自然群体材料中鉴定到荔枝高抗霜疫病品种裕荣1号(YR1)。从荔枝基因组中共鉴定出19个LcCDPK基因家族成员,分布于11条染色体上。根据保守结构域和系统发育分析将其分为4个亚家族。基因结构分析表明,LcCDPKs外显子数量为7~19。蛋白结构分析发现,所有LcCDPK蛋白均具有1~4个EF-hand结构域。顺式作用元件分析表明,LcCDPKs成员拥有大量的生物和非生物胁迫响应元件。组织表达分析发现,LcCDPK基因在荔枝不同组织存在组织特异性。另外,表达分析发现在高抗霜疫病荔枝裕荣1号(YR1)中,LcCDPK5、LcCDPK17和LcCDPK19在霜疫病胁迫后急剧上调表达,LcCDPK3和LcCDPK8急剧下调表达。【结论】裕荣1号(YR1)是一种高抗霜疫病的荔枝品种。荔枝基因组中共鉴定出19个CDPK基因家族成员,具有明显的组织特异性,LcCDPK5、LcCDPK17、LcCDPK19、LcCDPK3和LcCDPK8可能在荔枝抗霜疫病过程中发挥重要作用。研究结果为进一步探究荔枝CDPK基因家族提供了参考。

拉枝对仙进奉荔枝枝梢生长和成花坐果的影响

【目的】仙进奉荔枝是近年推出的优质晚熟新品种,在中晚熟荔枝产区经济效益和社会效益显著,其高效丰产栽培技术有待研发。拉枝是一种重要的果树整形修剪手段,能有效削弱顶端优势,利于开花结果。探究不同拉枝处理对仙进奉荔枝的影响,为仙进奉荔枝拉枝技术的应用和稳产丰产技术建立提供参考。【方法】以4年生仙进奉荔枝为材料,设置直立(不拉枝,CK)、水平(90°)和下垂(110°)3个处理,统计处理后枝梢生长量、成花率和座果率等指标。【结果】与直立处理(CK)相比,下垂和水平拉枝处理对第1次、第2次梢营养生长影响有限,但能延迟第3次梢(末次秋梢)萌芽7 d以上,同时显著减少末次秋梢萌芽数目,且以下垂处理的抑制效果最强。末次秋梢老熟时,拉枝处理枝条直径略低于直立处理(CK),各处理间差异不显著;下垂和水平拉枝处理均可有效将末次秋梢长度控制在15 cm左右,促生中短枝,利于开花坐果。翌年春季,下垂和水平处理枝组的成花率和座果率显著高于直立处理(CK),其中下垂处理枝组成花率达到100%,且全部为纯花穗、花穗质量高;直立处理(CK)枝组产生较多带叶花穗。各处理枝组的花穗长度无显著差异,但拉枝处理可显著降低花穗基部直径、抑制花穗旺长。雌花盛开后50 d,下垂和水平处理枝组座果率分别为9.81%和7.61%,显著高于直立处理(CK,5.54%)。【结论】下垂和水平拉枝可有效控制仙进奉荔枝结果母枝徒长,改善成花质量,显著提高座果率,以110°下垂拉枝效果最佳。