龙眼miR396分子进化特性及响应蓝光的表达分析

关于蓝光对龙眼胚性愈伤组织miRNA组学研究中发现大量响应蓝光的miRNAs,挑选关键且差异表达的miR396a-3p_2和miR396b-5p进行分子进化特性分析,以进一步明确它们在龙眼响应蓝光过程中的作用。该研究对前体和成熟体进化树、前体二级结构、启动子顺式作用元件、靶基因预测及其响应蓝光表达模式等进行分析。结果表明:(1)由5p臂上形成的miR396成熟体序列保守性较高,由3p臂上形成的miR396成熟体序列特异性较大;茎序列的保守性高于环序列,5p臂上保守性高于3p臂。(2)miR396a-3p_2和miR396b-5p的启动子主要包括光、激素、生物和非生物胁迫响应元件,可能通过这些顺式元件在龙眼响应蓝光中起调控作用;其响应蓝光的靶基因或蛋白主要包括生长素调控因子(GRF2)、LRR类受体丝氨酸(FLS2)、乙烯不敏感蛋白(EIN3)和DNA定向RNA聚合酶α亚基(rpoA)等。(3)实时荧光定量PCR结果表明,miR396a-3p_2和miR396b-5p在不同光质的表达模式存在差异;在不同光强的蓝光条件下,miR396b-5p与其靶基因的表达趋势基本呈负相关,而miR396a-3p_2与rpoA的表达趋势未呈负相关,可能存在其他miR396家族成员或miRNAs同时调控,从而实现靶基因转录水平的调控。     

Cytokinins from terminal buds of Euphoria longana during different growth stages

The presence of endogenous cytokinins were detected in the terminal buds of longan ( Euphoria longana Lam.) after purification by ion exchange and Sephadex LH-20 chromatography, and bioassay, enzymic degradation, high-performance liquid chromatography and gas chromatography-mass spectrometry. Permethylated derivatives of two highly active cytokinin glucoside compounds from dormant buds were: 6-(4-O-β-D-glucosyl-3-methyl-but-2-enylamino) purine (zeatin-O-glucoside) and 9-β-D-ribofuranosyl-6-(4-hydroxy-3-methyl-but-2-enylamino) purine (zeatin riboside-O-glucoside). Simultaneously, four active cytokinins from buds at the stages of leaf flush and flower bud initiation were identified as 6-(4-hydroxy-3-methyl-but- trans -2-enylamino) purine (zeatin), zeatin-9-β-D-ribofuranosylpurine (zeatin riboside), 6-(3-methyl-2-butenyl) aminopurine (isopentenyladenosine, 2iPA) and N-(3-methyl-2-butenyl) adenine (isopentenyladenine, 2iP). The total cytokinin levels were low at leaf flush, with the zeatin and zeatin riboside in the buds about 70% of the total. In the transition of the terminal bud from leaf flush to dormancy, the principal cytokinins were zeatin-O-glucoside and zeatin riboside-O-glucoside. However, significant decreases in the levels of zeatin-O-glucoside and zeatin riboside-O-glucoside and increases in those of zeatin, zeatin riboside, 2iPA and 2iP were observed at flower bud initiation. It is suggested that in longan, the cytokinins that are translocated to the shoots are accumulated in the buds at the dormant stage, and that later there is a considerable increase in free cytokinins during flower bud initiation, leading to the promotion of flower bud development.     

低温胁迫下龙眼(Dimocarpus longana Lour.)幼叶细胞内Ca2+水平及细胞超微结构

用焦锑酸钾沉淀的电镜细胞化学方法研究低温胁迫下龙眼幼叶细胞内Ｃａ＾２＋水平的变化。未经低温处理的幼叶,细胞壁,细胞间隙,质膜和液泡存在大量的Ｃａ＾２＋沉淀颗粒,叶绿体,细胞质和细胞核中也有一些Ｃａ＾２＋沉淀颗粒分布,表明液泡和细胞间隙是植物细胞的钙库。     

龙眼内生菌的分离与脂肪酸鉴定

对‘福眼’龙眼果实的内生菌进行分离纯化和脂肪酸鉴定,结果表明,在龙眼果实中检测到分属于肠杆菌属(Enterobacter)、果胶杆菌属(Pectobacterium)、克吕沃尔菌属(Kluyvera)和沙门氏杆菌属(Salmonella)的内生细菌,以及分属于枝孢霉属(Cladosporium)、柱顶孢霉属(Scytalidium)、外瓶霉属(Exophiala)的内生真菌。     

Association of Eriophyes dimocarpi (Acari: Eriophyidae) with longan witches’ broom disease in Vietnam

Longan witches’ broom (LgWB) syndrome was observed in all longan-growing provinces of Vietnam. Key symptoms include: small, stunted shoots with curved, rolled up margins, deformed leaves with blisters and hairy patches on the underside; abnormal development of flower structures, flower abortion, failure to produce fruits or developing small fruits. During 2012–2015, field surveys and study of the relationship between LgWB symptoms and Eriophyes dimocarpi mites presence were carried out. Moreover, molecular analysis, electron microscopy observations, E. dimocarpi inoculations, pruning of affected spikelets and miticide experiments were performed. An average of 36.58% of longan seedlings inoculated with E. dimocarpi mites showed typical symptoms of LgWB at about 27.5 days after inoculation. The use of several pesticides in different combinations showed high efficacy on E. dimocarpi mites elimination and of symptomatology incidence reduction. These results suggest a significant association of E. dimocarpi with LgWB syndrome in Vietnam.     

棕果蝠对龙眼的取食行为及进食量

棕果蝠(Rousettus leschenaulti)在云南西双版纳地区是比较常见的果蝠。本文研究了棕果蝠对当地经济作物龙眼(Dimocarpus longan)的取食模式和取食时间规律,并在实验室条件下研究了其对龙眼的整夜进食量。野外观察发现:棕果蝠叼取龙眼后不在母树上停留,而是携带龙眼到其他进食地点进食。棕果蝠在19:30～5:30时有3个取食高峰期,分别是在20:30～21:00时、22:00～22:30时和1:00～1:30时之间。此外,室内笼养实验结果表明,棕果蝠对龙眼的取食个数因体重不同而有差异,个数在6～20之间,实际进食量相当于其体重的1.0～1.8倍。     

外源NO对酸雨胁迫下龙眼幼苗生理特性的影响

研究了外源NO供体硝普钠(SNP)对pH3.0酸雨胁迫下龙眼幼苗叶绿素含量、抗氧化酶活性和渗透调节物质含量的影响.结果表明:酸雨胁迫下,龙眼幼苗叶片中的超氧化物歧化酶(SOD)、过氧化物酶(POD)和过氧化氢酶(CAT)活性降低,叶绿素、蛋白质和可溶性糖含量下降,丙二醛含量升高,呈现出一定的毒害效应.外源NO对酸雨胁迫下龙眼幼苗的生理代谢的作用具有双重性,0.1 ～0.5.mmol·L-1SNP显著提高叶绿素、可溶性糖和蛋白质含量,增强SOD、POD和CAT活性,降低丙二醛含量;其中0.5 mmol·L-1 SNP处理效果最好,叶绿素、可溶性糖和蛋白质含量分别比单独酸雨胁迫处理增加了76.0％、107.0％和216.1％,SOD、POD和CAT活性分别提高150.0％、350.9％和97.1％,丙二醛含量降低了46.4％.低浓度外源NO能通过刺激抗氧化酶活性来清除活性氧,减轻氧化胁迫,缓解酸雨胁迫对龙眼幼苗的毒害作用;而高浓度外源NO对酸雨胁迫的缓解作用减弱.     

不同生态类型龙眼种质亲缘关系的ISSR分析

利用ISSR分子标记技术对不同生态类型的39份龙眼种质进行亲缘关系分析。研究结果表明,从100条ISSR引物中筛选出12条重复性好、条带清晰的引物,对39份龙眼种质基因组DNA进行扩增,得到152个位点,其中多态性位点117个,多态性比例为76.97%。供试龙眼种质间遗传相似系数变幅为0.57～0.92,说明ISSR标记能够揭示材料间较高的遗传多样性。UPGMA聚类结果表明,在0.65相似水平可以将39份龙眼种质分为3个类群,类群Ⅰ均为来自中国的南亚热带生态型龙眼;类群Ⅱ包括石硖和大乌圆2个南亚热带生态型龙眼品种,以及热带生态型龙眼四季蜜类型的品种和单株;类群Ⅲ包括来自越南和泰国的龙眼种质。不同生态类型对龙眼的亲缘关系影响不大,热带生态型和南亚热带生态型龙眼相互聚在一起,说明两种不同生态类型龙眼具有较多相同的遗传背景。本试验结果将有助于进一步开展龙眼的分类、遗传与进化研究。     

龙眼体胚发生早期SAUR 63/64基因克隆及表达分析北大核心CSCD

SAUR(small auxin up-regulated RNA)是一类生长素早期响应基因,与植物的生长发育密切相关。为揭示SAUR 63和SAUR 64在龙眼体胚发生早期的调控作用,该研究采用TA克隆龙眼SAUR 63和SAUR 64基因,对其进行亚细胞定位、qRT-PCR表达分析和荧光原位杂交分析验证。结果显示:(1)龙眼SAUR 63和SAUR 64序列长度分别为386 bp和312 bp;系统发育树显示SAUR63、SAUR64与拟南芥亲缘关系较近;亚细胞定位结果显示SAUR63蛋白定位在细胞核与细胞外基质,SAUR64蛋白定位在叶绿体。(2)预测发现SAUR 63和SAUR 64同时作为LTCONS_00038949、LTCONS_00038950及LTCONS_00038952(lnc38949、lnc38950及lnc38952)的靶基因。(3)qRT-PCR检测发现SAUR 63、SAUR 64与3个lncRNAs均在不完全胚性紧实结构阶段高表达;SAUR 63与3个lncRNAs在100和500μmol·L^(-1)赤霉素(GA 3)处理下上调表达,且在500μmol·L^(-1) GA 3时表达量达到峰值;SAUR 63、SAUR 64、lnc38949和lnc38950均在35℃处理下被显著诱导表达,而lnc38950在35℃下受到抑制;此外,SAUR 63、SAUR 64和3个lncRNAs还响应SA、MeJA、盐胁迫和干旱胁迫的调控。(4)荧光原位杂交结果显示,SAUR 63与lnc38949均没有定位在合子胚特定部位,且lnc38949表达量高于SAUR 63。研究表明,SAUR 63、SAUR 64基因可能受到lncRNAs的调控,共同参与龙眼体胚发生早期的调控。     

滨海风沙地龙眼园土壤特性的研究

滨海风沙地土壤结构性差,质地疏松,氧气充足,有利龙眼菌根生长;沙地夏秋土温高,温差变幅大,持续时间久,对幼根生长 有害,克服措施是盖草降温和浇灌,而３５ｃｍ以下沙层温度终年适宜龙眼根系的生长。风沙地渗漏性大,蓄水力差,有机质贫乏,各种营养元素含量低;经过耕作培肥改土,土壤养分水平迅速提高,土壤速效羊分的增加和土壤微生物量和土壤酶活性有着密切的关系。在一年中土壤各元素含量的变化是冬高夏低,夏季表土上