Metabarcoding技术在植物鉴定和多样性研究中的应用

目前植物学研究已进入后植物志时代,iFlora的实现需要以传统植物分类学及相关研究为基础,整合基于高通量测序的DNA条形码（DNAbareoding）技术并开发便携式快速鉴定仪器及构建信息平台。传统植物鉴定多基于形态学分类,而近年来快速发展的DNA条形码快速鉴定技术被各界分类学家认可,在植物鉴定中也被广泛应用。但DNA条形码技术仍存在一些问题亟待解决,如种的鉴定需要多个条形码的解析、Sanger测序平台无法处理混合样品。本文介绍了传统植物分类技术和DNA条形码技术在植物研究中的应用和遇到的瓶颈;并重点介绍了基于高通量测序的metabarcoding技术在植物鉴定及多样性研究中的应用及前景,及其与iFlora的关系。     

Response of soil microbial activity to grazing, nitrogen deposition, and exotic cover in a serpentine grassland

Background and aims

Exotic species, nitrogen (N) deposition, and grazing are major drivers of change in grasslands. However little is known about the interactive effects of these factors on below-ground microbial communities.

Methods

We simulated realistic N deposition increases with low-level fertilization and manipulated grazing with fencing in a split-plot experiment in California’s largest serpentine grassland. We also monitored grazing intensity using camera traps and measured total available N to assess grazing and nutrient enrichment effects on microbial extracellular enzyme activity (EEA), microbial N mineralization, and respiration rates in soil.

Results

Continuous measures of grazing intensity and N availability showed that increased grazing and N were correlated with increased microbial activity and were stronger predictors than the categorical grazing and fertilization measures. Exotic cover was also generally correlated with increased microbial activity resulting from exotic-driven nutrient cycling alterations. Seasonal effects, on abiotic factors and plant phenology, were also an important factor in EEA with lower activity occurring at peak plant biomass.

Conclusions

In combination with previous studies from this serpentine grassland, our results suggest that grazing intensity and soil N availability may affect the soil microbial community indirectly via effects on exotic cover and associated changes in nutrient cycling while grazing directly impacts soil community function.     

Cloning,characterization and expression of \emph{OsFMO}_{{\mathbf{(}\emph{t}\mathbf{)}}} in rice encoding a flavin monooxygenase

Flavin monooxygenases (FMO) play a key role in tryptophan (Trp)-dependent indole-acetic acid (IAA) biosynthesis in plants and regulate plant growth and development. In this study, the full-length genomic DNA and cDNA of OsFMO _(t), a FMO gene that was originally identified from a rolled-leaf mutant in rice, was isolated and cloned from wild type of the rolled-leaf mutant. OsFMO _(t) was found to have four exons and three introns, and encode a protein with 422 amino acid residues that contains two basic conserved motifs, with a ‘G×G××G’ characteristic structure. OsFMO_(t) showed high amino acid sequence identity with FMO proteins from other plants, in particular with YUCCA from Arabidopsis, FLOOZY from Petunia, and OsYUCCA1 from rice. Our phylogenetic analysis showed that OsFMO_(t) and the homologous FMO proteins belong to the same clade in the evolutionary tree. Overexpression of OsFMO _(t) in transformed rice calli produced IAA-excessive phenotypes that showed browning and lethal effects when exogenous auxins such as naphthylacetic acid (NAA) were added to the medium. These results suggested that the OsFMO_(t) protein is involved in IAA biosynthesis in rice and its overexpression could lead to the malformation of calli. Spatio-temporal expression analysis using RT-PCR and histochemical analysis for GUS activity revealed that expression of OsFMO _(t) was totally absent in the rolled-leaf mutant. However, in the wild type variety, this gene was expressed at different levels temporally and spatially, with the highest expression observed in tissues with fast growth and cell division such as shoot apexes, tender leaves and root tips. Our results demonstrated that IAA biosynthesis regulated by OsFMO _(t) is likely localized and might play an essential role in shaping local IAA concentrations which, in turn, is critical for regulating normal growth and development in rice.     

NO体外供体硝普钠诱导蚕豆气孔保卫细胞死亡机理研究

为分析NO在植物细胞死亡过程中的作用,以蚕豆表皮条和NO体外供体硝普钠(SNP)及NO信号途径抑制剂为材料,采用表皮条生物法,探讨SNP对蚕豆叶面保卫细胞的毒性机理.结果表明:(1)0.5～9 mmol· L-1的SNP可使蚕豆气孔保卫细胞活性降低,部分细胞死亡,且随着SNP浓度的增高细胞死亡率增高.(2)凋亡抑制剂Z-Asp-CH2-DCB或TLCK可显著降低SNP诱发的保卫细胞死亡率.(3)抗坏血酸(AsA)、过氧化氢酶(CAT)、Ca2+螯合剂EGTA或Ca2+通道抑制剂LaCl3与SNP共同作用时,细胞死亡率显著降低.(4)NO清除剂c-PTIO、MAPK激酶抑制剂PD98059和鸟苷酸环化酶抑制荆ODQ亦能有效阻止SNP诱发的细胞死亡.研究发现,较高浓度的SNP可诱导蚕豆保卫细胞程序性死亡,SNP诱发植物细胞死亡与胁迫组保卫细胞内NO、ROS和Ca2+水平升高有关,cGMP和MAPK参与了SNP诱发的细胞死亡.     

甜瓜抗蔓枯病基因Gsb-3的ISSR分子标记

以甜瓜抗病资源PI511890和感病自交系‘白皮脆’及其F1、BC1P1、BC1P2、F2群体为材料,苗期接种蔓枯病菌(Did ymella bryoniae)进行遗传分析.结果表明:(1)对甜瓜苗期蔓枯病菌接种鉴定结果显示,抗源PI511890的抗病基因Gsb-3由显性单基因控制.(2)利用集团分离分析及ISSR分析技术,引物ISSR-100扩增出的多态性条带与Gsb-3表现连锁关系,该多态性片段大小为900 bp.(3)统计ISSR-100在182个F2单株上的多态性,并用JoinMap 4.0软件分析结果显示,ISSR-100与Gsb-3遗传连锁距离为8.3 cM,定名为ISSR-100900.研究认为,ISSR100900可作为甜瓜抗蔓枯病分子育种的备选分子标记.     

微波法从黄连中提取黄连素的工艺研究

为了研究微波法从黄连中提取黄连素的效果,分别考察不同微波功率、原料浸泡时间、料液比(g/mL,下同)、提取次数、微波辐射时间对黄连素提取率的影响。结果微波法从黄连中提取黄连素的最佳工艺条件为:提取功率500W,原料浸泡时间24 h,料液比1∶120,提取次数2次,微波辐射时间3 min,提取率达5.61%,提取效果较好。与硫酸法、石灰法、乙醇浸取法相比,不仅提高了提取速度,并且更加环保。     

温度对亚热带地区常见树种叶片甲烷排放的影响

以亚热带常见树种米槠、木荷、浙江桂、罗浮栲、杉木和柑橘为对象,利用控制试验研究了温度对树木叶片甲烷(CH4)排放的影响.结果表明:当温度在10℃时,供试的6种树木中,仅木荷、柑橘和罗浮栲的叶片排放CH4;温度高于20℃时,所有树木叶片均可排放CH4.温度高于30℃时,叶片排放CH4的平均排放速率(1.010ngCH4·g-1DM·h-1)是10～30℃时平均排放速率(0.255ngCH4·g-1DM·h-1)的3.96倍.增温对柑橘和杉木CH4排放速率的影响显著高于其他4种树木.培养时间对叶片排放CH4速率有显著影响,温度胁迫对树木排放CH4的影响受植物活性的控制.在低温或高温条件下,树木干叶均不能排放CH4.高温胁迫对树木叶片排放CH4有重要影响,全球变暖可能增加植物的CH4排放.     

冬虫夏草寄主蒲氏钩蝠蛾雄成虫触角感受器的观察

蒲氏钩蝠蛾Thitarodes pui(Zhang et al.)是冬虫夏草寄主昆虫之一,其雄成虫触角感受器在求偶交配过程中起主要作用。本研究应用电子扫描显微镜对蒲氏钩蝠蛾雄成虫触角上的化学感受器进行观察。结果发现,雄成虫触角上有7种感受器,即毛形感受器、刺形感受器、锥形感受器、腔锥形感受器、钟形感受器、Bhm氏鬃毛和鳞形感受器,其中以毛形感受器和鳞形感受器数目最多,腔锥感受器又分为长栓形和短栓形两种。综合本研究结果与已知蝠蛾的触角感受器,发现蝠蛾触角感受器在表面结构、感受器类型等方面与其它鳞翅目昆虫存在差异。