温郁金萜类合酶基因CwTPS4的克隆和表达特征

温郁金是著名的"浙八味"之一,药用价值高、应用广泛,萜类化合物是其主要的药用成分.萜类合酶是植物萜类化合物生物合成途径中的关键酶.依据温郁金根茎的转录组数据,采用反转录PCR获得了1个萜类合酶基因CwTPS4(GenBank登录号:MW774935),其开放阅读框长1515 bp,编码504个氨基酸,含有萜类合酶特有的结构域和保守序列RXR、DDXXD等;生物信息学分析表明CwTPS4编码的蛋白定位在细胞质中、无跨膜区域,为水溶性的稳定蛋白,属于萜类合酶的TPS-a亚家族成员.实时荧光定量分析表明,CwTPS4基因主要在生长旺盛的叶片中表达,其次在根茎膨大初期的根茎中表达量较高,而在成熟或衰老的组织中表达量较低.本研究从温郁金中克隆得到1个新的萜类合酶基因CwTPS4,经生物信息学分析表明CwTPS4可能参与了温郁金中倍半萜类化合物的合成,为下一步研究其在温郁金萜类物质合成中的功能奠定了一定的基础.     

姜黄属植物过氧化物酶同工酶的研究

用聚丙烯酰胺凝胶电泳分析姜黄属14种(28个样品)植物的过氧化物酶同工酶。结果表明属内种间有明显的酶谱差异,各种都有特征酶谱。根据酶谱特征及酶谱距离,结合根茎颜色,可把14个种分成三群:第一群,根茎黄色至深红色,有姜黄、毛姜黄、郁金、印尼莪术、C.petiolata和C.sP.(2);第二群,根茎灰白色,有广西莪术、大莪术和温郁金;第三群,根茎浅黄而间淡蓝色或深蓝色,有莪术、顶花莪术、细莪术、C.aeruginosa及C.zedoaria。研究结果还表明:1.国产莪术的酶谱与从美国引入的C.zedoaria和C.aeruginosa均不相同,而与引自新加坡的C.Phaeocaulis一致;2.不同形态的广西莪术具有完全一致的酶谱;3.温郁金与郁金的酶谱有差异。     

HPLC测定不同产地温郁金中吉马酮和莪术二酮的含量

高效液相色谱法(HPLC)测定10批不同产地道地性温郁金药材中的吉马酮和莪术二酮的含量,比较不同产地的含量差异,为质量评价和临床合理应用提供合理依据。运用水蒸气蒸馏法,采用Purospher STAR~LP C18色谱柱(250mm×4.6mm,5μm),以乙腈-水为流动相梯度洗脱,流速为0.8m L/min,检测波长225nm,柱温35℃,进样量10μL。虽同为道地性药材,但不同产地温郁金中化学成分的含量仍有差异,吉马酮和莪术二酮含量最高的产地是均为乐清,平均含量分别为0.020%和0.039%。该方法简便易行,采用重复蒸馏,测量结果相对准确,可作为测定温郁金药材中吉马酮、莪术二酮含量的相对准确的方法。     

温郁金超临界萃取物的化学成分研究

采用超临界流体萃取技术萃取温郁金中的挥发性成分,通过L9(34)正交设计对萃取条件进行优化,结果表明55℃,35 MPa,240 min为较优条件。GC-MS联用技术对挥发性成分进行定性及半定量分析,首次从温郁金中检测到香橙烯氧化物、喇叭烯氧化物、卡拉烯环氧化物等11种化合物。与水蒸汽蒸馏工艺相比,超临界萃取工艺的收率较之高出3倍,萃取时间缩短4倍,抗癌有效活性成分榄香烯、莪二酮、莪术醇和吉马酮的得率分别高出32%,23%,105%和15%。     

温郁金内生拮抗细菌B-11的分离及其抑菌活性

【背景】植物内生菌广泛分布在自然界中,具有巨大的潜在开发与应用价值。【目的】对温郁金内生细菌进行分离鉴定,并从中筛选出具有生防潜能的菌株。【方法】采用常规组织分离法对温郁金根茎内生细菌进行分离并利用16S rRNA基因序列分析进行初步鉴定;采用平板对峙法以铁皮石斛炭疽病菌(Colletotrichum gloeosporioides)为供试菌株对温郁金内生细菌进行拮抗菌的筛选;通过形态学鉴定、生理生化鉴定以及16S rRNA基因序列分析,确定拮抗菌株B-11的分类地位;以6个不同属的植物病原真菌为供试菌株对拮抗菌株B-11的抑菌谱进行测定,并研究其对病原菌菌丝的影响;利用特异性平板和MALDI-TOF-MS检测技术对拮抗菌株B-11产生的抑菌物质进行检测;采用离体叶片接种法研究拮抗菌株B-11对铁皮石斛炭疽病菌的防治效果。【结果】从温郁金根茎中分离得到25株内生细菌菌株,这些细菌菌株分属于12个属,其中芽孢杆菌属为优势菌属,占分离菌株的28%;通过初筛获得8株对C.gloeosporioides有抑菌活性的菌株,其中菌株B-11的抑菌活性最强,经鉴定该菌为贝莱斯芽孢杆菌(Baclliusvelezensis);抑菌谱测定发现菌株B-11能够对供试的属于不同属的6种植物病原真菌的菌丝生长具有抑制作用,显微观察发现经对峙培养6 d后这6种病原菌的菌丝体出现畸形膨大、分枝增多等现象;特异性平板检测结果表明拮抗菌株B-11能够产生蛋白酶、β-葡聚糖酶和嗜铁素,但不产生几丁质酶;MALDI-TOF-MS检测结果表明拮抗菌株B-11能够产生伊枯草素、丰源素和表面活性素3种脂肽类抗生素,其中伊枯草素的产量最高;离体叶片接种实验表明,拮抗菌B-11的离心去菌发酵液对铁皮石斛炭疽病的防治效率可达64%。【结论】温郁金根茎含有丰富的内生细菌资源,其内生细菌菌株B-11有潜力作为开发抗真菌代谢物和新药物的重要微生物资源。     

榄香烯治疗非小细胞肺癌的作用机制研究进展

榄香烯注射液是从天然中草药姜科植物温郁金中提取获得的萜烯类化合物,其主要生物学活性为降低肿瘤细胞有丝分裂能力,诱导肿瘤细胞凋亡,抑制肿瘤细胞的生长,从而改善NSCLC患者的临床症状,增强患者免疫力.本文主要从杀灭癌细胞、诱导癌细胞凋亡、抑制癌细胞血管形成、增强患者免疫力等几方面综述榄香烯注射液对非小细胞肺癌的作用机制.     

榄香烯——一种新型的抗肿瘤药

榄香烯为姜科植物温郁金中提取的有效成分,是近年来我国自行开发的新型抗肿瘤药物,经具有疗效好,副作用小等特点,受到了广泛的重视。本文综述了榄香烯的一般药理特性,体内和体外抗肿瘤作用、临床应用及其作用机理等几方面的研究工作     

温郁金的组织培养与快速繁殖

1植物名称温郁金(Curcuma wenyujin Y.H.Chen et C.Ling)。2材料类别主根茎。3培养条件以MS为基本培养基。芽萌动与生长培养基:(1)MS 6-BA 1.5 mg·L~(-1)(单位下同)。不定芽增殖与愈伤组织诱导培养基:(2)MS 6-BA 1.0 KT 0.5;(3)MS 6-BA 2.0 KT 1.0;(4)MS 6-     

郁金樱愈伤组织诱导及植株再生

为建立郁金樱(Cerasus serrulata var. lannesiana cv. ‘Grandiflora’)再生体系,以多年生母株小叶、一年生嫁接苗小叶、腋芽诱导小叶和增殖一代小叶为外植体,探讨不同外植体和植物激素组合对郁金樱愈伤组织诱导、不定芽分化、增殖和生根的影响。结果表明,4种外植体均可诱导出愈伤组织,...     

中药郁金的挥发油成分研究

本文报道了对5种植物来源的中药郁金中挥发油的比较研究结果,测定了不同采收期以及不同加工方法郁金中挥发油含量,并采用GC-MS-DS法分析比较了郁金挥发油的组成及各组分含量。     

近年来中国热带地区外来有害昆虫的种类特征及入侵分析

我国热带地区是外来入侵物种发生与危害的重灾区,近年来外来有害昆虫入侵所引发的生物灾害及生态安全问题日趋严峻。在海南省、广东省、广西壮族自治区、云南省、福建省外来入侵昆虫的调查结果及文献资料的基础上,整理分析了中国热带地区外来入侵昆虫的种类构成、分布、原产地及首次发现地。据统计,2000-2014年我国热带地区的外来入侵昆虫种类共计23种,隶属于6目11科,数量最多的为半翅目(13种,56.52%),其中以粉蚧科和粉虱科居多;热带地区的外来入侵昆虫在不同地区之间的物种数量差异较大,其中海南省最多(19种,82.61%),其后依次为广东省(16种,69.57%)、云南省(14种,60.87%)、广西壮族自治区(12种,52.17%),而福建省最少(9种,占39.13%);在入侵我国热带地区的外来有害昆虫中,原产于亚洲、北美洲的种类最多均为6种,占26.09%,南美洲占21.74%,而从非洲、大洋洲、欧洲传入的分别占8.70%、4.35%、4.35%;外来入侵昆虫的首次发现地位于广东省的有9种,占39.13%,出现在海南省和云南省的各占30.43%、21.74%,出现在广西壮族自治区的为8.70%。阐明我国热带地区外来有害昆虫的种类特征及入侵现状,可为该地区外来入侵昆虫的监测与防控提供一定的依据。     

PARTICLE SIZE OF INSECTICIDAL SUSPENSIONS AND THEIR CONTACT TOXICITY III. TEMPERATURE COEFFICIENTS AND TESTS BY INJECTION

Aqueous DDT suspensions containing particles of different sizes gave different temperature coefficients of mortality in dipping tests against adult Oryzaephilus surinamensis and Tribolium castaneum. In the range 12–30°C., colloidal DDT showed a large negative coefficient, but the coefficient for a suspension of DDT crystals of 400μ was smaller ( T. castaneum ) or negligible ( O. surinamensis ). Thus, although a suspension of crystalline DDT was more toxic than colloidal DDT to warm insects, it was less toxic than the colloid to cool insects.
A test was made of rotenone suspensions against adult milkweed bugs ( Oncopeltus fasciatus Dall.) by application of measured droplets under the wings. Colloidal rotenone was more toxic than a crystalline suspension. Milkweed bugs could not be killed in this way by DDT suspensions.
Tests were also made by injecting suspensions of rotenone, DDT and its fluorine analogue (DFDT) into milkweed bugs. A suspension of rotenone crystals was equitoxic with colloidal rotenone against insects kept for 3 days at 27°C. after treatment. The same was true of crystalline and colloidal DDT suspensions. If the bugs were kept at 10°C., colloidal rotenone was much more toxic than crystalline rotenone 2 days after treatment. But if the insects were kept for about 3 weeks and inspected at intervals, the first apparent difference in toxicity grew smaller, and in the end it nearly disappeared. With DDT the time for this to occur was about 10 days; and suspensions of colloidal and crystalline DFDT were equitoxic after only 2 days, even at 10°C.
In the application test, the difference between colloidal and crystalline rotenone was a real difference in toxicity. It did not disappear with the lapse of time; but in the injection tests the two forms of each poison differed only in speed of action, and not in ultimate toxicity.     

Aggregation of entomopathogenic nematodes, heterorhabditis spp. and steinernema spp., among host insects at 9 and 20 degrees C and effects on efficacy

The negative binomial distribution (NBD) with density-dependent k was used to test and describe the distribution of nematodes in Otiorhynchus sulcatus, Galleria mellonella, and Spodoptera exigua at 9 and 20 degrees C. Aggregation was greater in O. sulcatus than in the other two hosts. In case of G. mellonella exposed to the heterorhabditid HF85, the distribution of nematodes in the insects was random. There was only a small effect of low temperature on aggregation; this effect was stronger for O. sulcatus than for the other two hosts. Aggregation of nematodes among insects in a population increases the infection chance of already infected insects and decreases the infection chance of uninfected insects. Consequently, more nematodes will have to penetrate O. sulcatus than G. mellonella or S. exigua to cause the same proportion of infection in a population. The relevance of aggregation of nematodes for control O. sulcatus is discussed. Copyright 1999 Academic Press.     

Stressors and rearing diseases of Trichoplusia ni: evidence of vertical transmission of NPV and CPV.

Inability to consistently rear healthy Trichoplusia ni led to a study of its rearing diseases. Four diseases were designated after preliminary research which included electron microscopy: cytoplasmic polyhedrosis (due to cytoplasmic polyhedrosis virus, or CPV), nuclear polyhedrosis (due to nucleopolyhedrovirus, or NPV), "neonate death" syndrome (mortality in first or second instars), and "late-instar" syndrome (death in late instars accompanied by bacterial decomposition). Infectious agents were not detected by electron microscopy in insects with the latter two diseases. Prevalence of CPV and NPV, but not the neonate-death or late-instar diseases, in progeny was significantly associated with pairs of mating adults. In conjunction with egg-surface decontamination, this indicated that both viruses may be transmitted transovarially. Pupae killed by CPV had virions in the cytoplasm of infected cells, but polyhedra were empty, not occluding virions. None of the diseases had a consistent pattern of prevalence associated with the date on which eggs were laid after oviposition began. Prevalence rates of cytoplasmic polyhedrosis, nuclear polyhedrosis, and the late-instar disease were significantly greater at 95-100% relative humidity (RH) than at RH levels of 75% or below. These same three diseases killed significantly more insects in crowded rearing conditions (four or five larvae per cup with 10.2-cm(2) diet surface) than in uncrowded conditions (one to three larvae per cup). As a result of these experiments, healthy T. ni have been reared for 10 generations by use of a modified Pasteur method and rearing cups containing no more than two larvae.     

海南岛4市县人群口腔假丝酵母菌分布情况的初步调查

目的：调查海南岛海口、三亚市、五指山市和保亭县四市县人群口腔假丝酵母菌的分布情况,为口腔假丝酵母菌分子生物学和遗传学特点研究提供基础资料。方法：分离出的假丝酵母菌经过菌落特征、制片乳酸酚棉蓝染色、芽管形成试验、厚膜孢子形成试验等初步鉴定,并用科玛嘉显色培养基培养确定。结果：海口、三亚市、五指山市和保亭县人群口腔假丝酵母菌总分离率为25．23％．其中白假丝酵母菌占54．48％,光滑假丝酵母菌占14．04％,热带假丝酵母菌占11．06％,克柔假丝酵母菌占9．36％。住院病人和健康人口腔假丝酵母菌的分离率分别为39．95％和11．61％,差异有统计学意义（P〈0．01）,结论：海南地区人群口腔假丝酵母菌的总分离率高于国内其它省份,分离出的假丝酵母菌以白假丝酵母菌为首位,病人口腔假丝酵母菌分离率明显高于健康人。     

Isolation and characterization of polymorphic microsatellite loci in Aleurodicus dispersus (Hemiptera, Aleyrodidae)

Ten microsatellite markers were isolated and characterized from Aleurodicus dispersus, the spiraling whitefly, an exotic pest species that is considered to be one of the most serious agricultural pests on Hainan Island, China. The polymorphism of these loci was examined in individual whiteflies from Hainan Island and from the Canary Islands. All loci were polymorphic, with two to four alleles per locus. Mean observed and expected heterozygosity values were 0.773 and 0.585, respectively. These microsatellite markers provide powerful tools for ecological, epidemiological and population genetic studies on this highly invasive insect. Thirty insects were collected and studied at each location. There were no differences between the two locations.     

海南坡鹿的起源、进化及保护

坡鹿是世界濒危物种,三个亚种分布在东南亚大陆,仅海南坡鹿种群分布在中国海南岛.2003年,国际社会的专家和学者提出了将海南坡鹿引入泰国亚种原分布区,重建已经绝灭野生种群的建议.在此种情况下,明确海南坡鹿的起源、与其它亚种间的系统发生关系、以及遗传多样性水平对有效保护坡鹿具有重要意义.本研究以线粒体DNA D-loop区490 bp基因片段为分子标记,比较分析了海南坡鹿、泰国亚种和缅甸亚种共35个样本的序列差异.我们所测的样本中,总共发现4种单倍型.所有21个海南坡鹿样品共享1种单倍型.利用最大似然法(ML)、最大简约法(MP)、邻接法(NJ)和贝叶斯法(Bayesian)构建的系统进化树表明海南坡鹿种群与泰国亚种的关系较近.但是,二者也发生一定程度的遗传分化.海南坡鹿与泰国亚种的遗传距离均值为0.026.我们推测海南坡鹿可能是在更新世冰期(69万年前)通过陆桥由东南亚大陆迁入中国海南岛.我们的结论说明海南坡鹿的遗传多样性很低,并且已独立进化很长时间.因此,我们不支持将海南坡鹿引入泰国亚种的原分布区,重建已经绝灭的野生种群的设想和建议.我们建议将海南坡鹿与泰国亚种分别作为两个独立的进化显著单元(ESUs)进行管理.     

Essential fatty acid for the mosquito Culex pipiens: arachidonic acid.

A nutrient associated with animal-derived phospholipids has previously been found essential for newly-emerged adults of the mosquito Culex pipiens to fly and survive more than a few days. Pure arachidonic acid was completely effective in supporting the emergence of viable flying adults; in combination with synthetic dipalmitoyl lecithin, which slightly improves larval growth rate without inducing adult flight, it wholly adequately replaces animal phospholipids. Linoleic and linolenic acids, which have satisfied the needs of all insects hitherto shown to require an essential fatty acid, were ineffective for C. pipiens, with or without synthetic lecithin. An optimal effect on adult flight was obtained with 0.05 mg of arachidonic acid per 100 ml of dietary medium, a concentration much lower than the linoleic/linolenic concentrations needed by other insects with an essential fatty acid requirement. The relationship of this unique mosquito fatty acid requirement to the essential fatty acid needs of both vertebrates and insects in general is discussed.