果蝇细胞凋亡核心机制的基因组比较

基因组比较研究是从基因组序列推测调控网络的主要途径。细胞凋亡信号网络是调控网络的一个典型代表。ＥＧＬ１、ＣＥＤ３、ＣＥＤ４和ＣＥＤ９及其同源蛋白质的线虫和哺乳动物构成保守的凋亡核心机制。目前果蝇细胞凋亡核心机制尚不完整,还未找到ＥＧＬ１和ＣＥＤ９类似蛋白质。通过一系列基于生物信息学的基因组比较分析,在果蝇的基因组数据库中发现了两个ＢＣＬ２／ＣＥＤ９和一个ＥＧＬ１的同源蛋白质的编码基因,并重构了果蝇     

黑腹果蝇的性别控制

性别的形成包括两个过程,即性别决定和性别分化。果蝇的性别控制研究包括性别决定、性别分化、性别鉴定、性别诱导和性别控制5个方面。性别决定是在两种不同发育途径之间的选择,它提供了一个研究基因调控的模式系统。果蝇的性别决定问题已经研究得相当详细［1］。性别分化是使胚胎向着雌性或雄性发育的过程,决定了性别表型。果蝇的性别分化也取得了不少研究成果。近年来,许多重要的性别调控基因已被克隆和鉴定。随着果蝇基因组全序列测定的完成,果蝇的性别控制研究将会更为深入而完善。本文对与黑腹果蝇性别决定和性别分化相关的一些问题进行综述。     

黑腹果蝇细胞谱系分析方法进展

对动物体内单个细胞的谱系进行分析有助于追踪其在发育过程中的作用,但是体内各种组织都是由很多形态、结构、功能各不相同的细胞构成的复杂系统,这种复杂性严重阻碍了对单个细胞的研究。嵌合克隆技术(Mosaic technique)和标记技术(Labeling technique)的出现为这一研究提供了强有力的手段。文章介绍了近几年来黑腹果蝇(Drosophila melanogaster)研究中常用的7种嵌合克隆标记方法,包括FRT介导的有丝分裂重组(FRT-mediated mitotic recombination)、MARCM(Mosaic analysis with a repressible cell marker)、TSG(Twin spotgenerator)、Twin-spot MARCM、Q-MARCM(Q system-based MARCM)、Coupled MARCM和G-TRACE(Gal4technique for real-time and clonal expression)技术,详述了这些技术的原理及应用,并对不同技术进行了对比。运用这些技术研究者可以从单细胞水平进行遗传学标记和操作,特别是在神经系统等复杂系统中追踪单个细胞的发育过程。果蝇中的这些技术也将为其他模式生物追踪细胞谱系提供参考。     

黑腹果蝇先天免疫研究进展

先天免疫是昆虫适应复杂环境的关键,也是新型害虫防治的重要研究方向。昆虫通过模式识别受体识别环境中不同的病原物,激活先天免疫系统以清除病原物。昆虫的先天免疫系统主要包括体液免疫与细胞免疫,体液免疫包括免疫信号通路诱导产生抗菌肽、活性氧以及黑化作用等,细胞免疫包括血细胞的吞噬、包囊和凝结。本文将重点总结黑腹果蝇Drosophila melanogaster在模式识别受体、免疫信号通路和细胞免疫相关方面的研究进展,为进一步研究其他经济昆虫与农业害虫的免疫机制,提高生产经济效益提供参考。     

霍氏肠杆菌对黑腹果蝇生长和发育的影响

【背景】共生菌群可以影响宿主的生理、代谢以及神经行为,黑腹果蝇是研究宿主与共生菌作用机制的优秀遗传模型。【目的】分离和鉴定黑腹果蝇体内的共生菌,并研究其对果蝇生长和发育的影响。【方法】利用YG固体培养基分离果蝇肠道菌;革兰氏染色、生化鉴定和16S rRNA基因比对鉴定菌种;体内定殖和世代传递实验验证细菌与果蝇的共生关系;建立无菌和悉菌模型,通过发育历期和生长速率来验证其促生长作用;RT-PCR检测促生长分子基因表达;免疫荧光染色检测果蝇肠道细胞的增殖;葡萄糖氧化酶法检测葡萄糖浓度。【结果】分离到一株能够明显促进果蝇生长发育的细菌,经鉴定为霍氏肠杆菌(Enterobacter hormaechei),该菌可以稳定定殖于黑腹果蝇肠道以及培养基内,而且可以世代垂直传递。该菌可明显地促进果蝇的生长,并通过加速促胸腺激素的分泌来发挥促生长作用,同时可以促进果蝇肠道细胞的增殖。霍氏肠杆菌相较于无菌果蝇可以降低体内葡萄糖水平。【结论】霍氏肠杆菌是黑腹果蝇的有益共生菌,实验证实其可以促进果蝇的生长和发育。     

黑腹果蝇的分离变相因子

众所周知,一对基因在杂合状态中保持相对的独立性,而在配子形成时,又按原样以相同比例分离到不同的配子中去,这是生物中最基本的遗传规律———孟德尔分离定律。但实际上并不是所有基因的分离都严格遵循孟德尔分离定律,存在于黑腹果蝇中的分离变相因子（ＳｅｇｒｅｇａｔｉｏｎＤｉｓｔｏｒｔｅｒ,以下简称ＳＤ）就是一种具有减数分裂驱动（ｍｅｉｏｔｉｃｄｒｉｖｅ）性质的、违反孟德尔分离定律的特殊遗传因子,从六十年代发现至今引起了人们的广泛关注,本文从其遗传背景,结构特征及进化规律等方面分别介绍一些近期的研究结果。１…     

樱桃新害虫黑腹果蝇的生物学特性

果蝇是近几年发现危害樱桃果实的一类重要害虫,在国内外樱桃产区均有发生。天水地区危害甜樱桃的果蝇有3个种,分别是黑腹果蝇Drosophila melanogaster Meigen、铃木氏果蝇Drosophila suzukii(Matsumura)和海德氏果蝇Drosophila hydei(Sturtevant),黑腹果蝇为优势种。作者记述黑腹果蝇对甜樱桃果实的危害情况、寄主范围及其生活史、生活习性、发育历期与温度的关系等,调查发现蚂蚁是樱桃果蝇的天敌之一。     

黑腹果蝇抗真菌肽基因Drs和Drs-lC 的原核可溶性表达及抗真菌活性测定

Drosomycin （Drs）是第1个从黑腹果蝇Drosophila melanogaster体内鉴定发现的昆 虫抗真菌肽因子。它对细菌无明显的抗性,但对丝状真菌具有高效广谱的抑杀作用。此外, 在黑腹果蝇基因组还存在着Drs的另外6个同系物的基因序列,其中同系物Drosomycin-lC（Drs-lC）的抗真菌谱仅次于Drs。将Drs抗真菌肽基因(Drs)和同系物Drs-lC基因（Drs-lC）进行可溶性表达,对果蔬等农产品防腐保鲜的研究有应用前景。本实验将Drs和Drs-lC分别克隆到硫氧还蛋白（Trx）融合表达载体pThiohis A中,转化宿主菌TOP10,进行可溶性表达,并从诱导表达的菌液起始浓度、IPTG的诱导浓度及诱导时间等方面进行了表达条件的优化。结果表明2种融合蛋白Trx-Drs和Trx-Drs-lC大部分以可溶形式表达,可溶性表达的Trx-Drs在上清液中约占菌体总蛋白的22％。2种融合蛋白的表达产物经 Ni-NTA亲和层析得到纯化。生测结果表明, 2种融合蛋白分别对8种供试真菌中的5种真菌显示明显的抗性。     

黑腹果蝇头部中RNA结合蛋白RBP9的互作蛋白筛选

【目的】筛选RNA结合蛋白RBP9在黑腹果蝇Drosophila melanogaster头部中的互作蛋白。【方法】利用CRISPR/Cas9基因组编辑技术,分别将3×Flag和V5标签编码序列插入到黑腹果蝇成虫头中RBP9和FNE基因起始密码子ATG的后面,构建黑腹果蝇转基因品系3×Flag-RBP 9/3×Flag-RBP9(3FRBP9)和V5-FNE/V5-FNE(VFNE);利用免疫沉淀和质谱法鉴定黑腹果蝇3FRBP9和野生型品系成虫头部中RBP9的互作蛋白并进行生物信息学分析。利用免疫共沉淀方法检测RBP9与FNE蛋白之间的相互作用。【结果】质谱法从黑腹果蝇成虫头部共鉴定了190个与RBP9相互作用的蛋白,其中包括ELAV和FNE。KEGG富集分析显示这些蛋白的基因主要参与核糖体、碳代谢、三羧酸循环、氨基酸生物合成等通路。免疫共沉淀实验结果验证了RBP9与FNE蛋白之间存在相互作用。【结论】RNA结合蛋白ELAV家族成员在黑腹果蝇成虫头部中具有相互作用。本研究为RBP9在果蝇神经系统发育中的功能研究提供了重要实验依据。     

黑腹果蝇黑条体突变体的求偶行为

黑腹果蝇Drosophila melanogaster黑条体果蝇(e^bsr)与黑檀体果蝇(e)为同一个基因(ebony)的不同突变体, 两者具有相似的形态表型, 但行为特征表现出明显的差异。本研究以黑条体、 黑檀体和野生型果蝇为研究对象, 首先检测果蝇的视力和活跃度, 再采用不同交配组合进行求偶成功率、交配时间和求偶模式的分析。结果表明: 黑条体果蝇视力与活跃度与野生型果蝇比较无显著差异; 黑条体果蝇的交配成功率和交配潜伏期与野生型果蝇不存在显著的差异; 黑檀体果蝇的交配成功率和交配潜伏期与野生型果蝇存在极显著的差异（P<0.000）。黑条体果蝇表现出异于黑檀体果蝇的活跃度和交配活力, 可能是由于黑条体果蝇ebony基因的新突变导致了果蝇体内多巴胺水平异常, 从而形成了黑条体果蝇独特的求偶模式。     

The Drosophila melanogaster genome sequencing and annotation projects: a status report.

The sequence and genome annotations of Drosophila melanogaster were initially published in late 1999 and early 2000. Since then, the Berkeley Drosophila Genome Project (BDGP) and FlyBase have improved the quality of the sequence and reviewed the annotations by hand, respectively, to produce an account of the fruit fly genome that is of the highest quality. This review discusses the main features of this process, both from the point of view of the biology revealed in the end result and in the development of software that has been central to this genome sequencing and annotation project.     

Drosophila melanogaster males respond differently at the behavioural and genome‐wide levels to Drosophila melanogaster and Drosophila simulans females

Drosophila melanogaster are found in sympatry with Drosophila simulans, and matings between the species produce nonfertile hybrid offspring at low frequency. Evolutionary theory predicts that females choose mates, so males should alter their behaviour in response to female cues. We show that D. melanogaster males quickly decrease courtship towards D. simulans females. Courtship levels are reduced within 5 min of exposure to a heterospecific female, and overall courtship is significantly lower than courtship towards conspecific females. To understand changes at the molecular level during mate choice, we performed microarray analysis on D. melanogaster males that courted heterospecific D. simulans females and found nine genes have altered expression compared with controls. In contrast, males that court conspecific females alter expression of at least 35 loci. The changes elicited by conspecific courtship likely modulate nervous system function to reinforce positive conspecific signals and dampen the response to heterospecific signals.     

Effects of auditory stimuli on the lifespan of Drosophila melanogaster

It is well known that music can have calming effects on humans, other mammals and birds. Reducing environmental stress or enhancing the resistance to certain stressors has been shown to extend lifespan in several organisms. Evidence also suggests that mild temporary stress may also enhance stress resistance and ultimately slow the aging process. This study explored the possibility that music may influence the lifespan of the fruit fly Drosophila melanogaster, possibly by affecting responses to stress. Flies received either background sounds (control), or background sounds supplemented with music (experimental). The experimental group had classical music playing constantly at an average of 20 dB above background sound. The median lifespan of females receiving music was 42 days compared to the median of 45 days without music, but the difference was not significant. For males median lifespans were 42 days with music exposure, and 47 days without music and the difference was significant. These results suggest that exposure to classical music decreased the lifespan of male Drosophila. Both experimental and control populations showed age-dependent increases in mortality, indicating that music affects the normal aging process rather than showing overt toxicity. These results suggest that certain auditory stimuli may be stressful and can be used as insect management.     

Patterns of protein synthesis following heat shock in pupae of Drosophila melanogaster

Pupae of Drosophila melanogaster were heat-shocked under conditions required to induce phenocopies in more than 90% of the flies that subsequently emerge. The effects of these treatments on protein synthesis in two tissues (thoracic epithelium and brain) were followed for several hours after the heat treatments. Results from pulse-labeling and protein separations on sodium dodecylsulfate (SDS) acrylamide gels showed a virtually complete cessation of protein synthesis immediately after the shock, followed by a noncoordinate resumption of the starting pattern. Similar experiments following double heat shocks demonstrated a more rapid resumption of synthesis of heat shock proteins after two successive heat treatments than after a single one.     

果蝇蛋白质组学研究进展

蛋白质组学旨在阐明基因组所表达的真正执行生命活动的全部蛋白质的表达规律和生物功能。随着人类基因组学计划的逐渐成熟,分子水平的实验技术不断发展,蛋白质组学的研究被提高到了前所未有的高度。果蝇是生命科学领域最为常用的一种模式生物,长期的系统研究也使果蝇的基因组成为至今注释最好的基因组之一,为功能基因组研究奠定了基础。但由于技术的限制,迄今有关果蝇蛋白质组学研究的报道尚不多见。近年来果蝇蛋白质组学的研究主要包括表达谱、修饰谱、比较蛋白质组学和疾病模型蛋白质组等四个方向,为进一步开展人类疾病临床蛋白质组学研究奠定了基础。     

Targeted gene disruption by CRISPR/xCas9 system in Drosophila melanogaster

Although the Cas9 protein from Streptococcus pyogenes (SpCas9) is the most widely used clustered regularly interspaced short palindromic repeats (CRISPR) variant in genome engineering experiments, it does have certain limitations. First, the stringent requirement for the protospacer adjacent motif (PAM) sequence limits the target DNA that can be manipulated using this method in insects. Second, its complementarity specifications are not very stringent, meaning that it can sometimes cause off-target effects at the target site. A recent study reported that an evolved SpCas9 variant, xCas9(3.7), with preference for various 5′-NG-3′ PAM sequences not only has the broadest PAM compatibility but also has much greater DNA specificity and lower genome-wide off-target activity than SpCas9 in mammalian cells. Here we applied the CRISPR/xCas9 system to target the white gene in Drosophila melanogaster, testing the genome-editing efficiency of xCas9 at different PAM sites. On the GGG PAM site, xCas9 showed less activity than SpCas9. For the non-NGG PAM site TGA, xCas9 could produce DNA cleavage and indel-mediated disruption on the target gene. However, for other non-NGG PAM sites, xCas9 showed no activity. These findings show that the evolved Cas9 variant with broad PAM compatibility is functional in Drosophila to induce heritable gene alterations, increasing the targeting range for the applications of genome editing in insects.     

鱼藤酮对果蝇运动行为及果蝇头部多巴胺合成相关酶基因表达的影响

为了探讨鱼藤酮对黑腹果蝇Drosophila melanogaster运动行为的影响与其头部多巴胺水平之间的关系,我们测定了鱼藤酮对黒腹果蝇成虫运动行为、头部多巴胺水平及酪氨酸羟化酶和多巴脱羧酶基因表达的影响。结果表明：与取食未加入药剂饲料的果蝇相比雌成虫用0.2～0.8 mmol/L、雄成虫用0.1～0.8 mmol/L浓度药液配制的饲料连续饲养6 d后运动能力显著下降,在0.8 mmol/L浓度下雌、雄成虫的运动能力分别仅为对照的55.6%和49.1%。取食用0.8 mmol/L浓度药液配制饲料6,12和21 d的果蝇雌、雄成虫头部多巴胺水平均显著下降,雌成虫头部多巴胺水平分别为对照雌成虫的83.2%,72.3%和59.8%;雄成虫头部多巴胺水平分别为对照雄成虫的79.3%,66.8%和53.2%。用0.8 mmol/L浓度鱼藤酮处理6,12和21d,雌成虫头部酪氨酸羟化酶基因（pale）的表达水平分别为对照的76.3%,51.4%和37.3%,多巴脱羧酶基因（Ddc）的表达水平分别为对照的87.1%,78.2%和63.5%, 均显著下降。结果提示,鱼藤酮可干扰果蝇成虫头部酪氨酸羟化酶和多巴脱羧酶基因的表达,导致果蝇头部多巴胺水平下降,进而影响了果蝇的运动行为。     

不同温度条件下果蝇染色体结构的变化

将黑腹果蝇Drosophila melanogaster置于不同温度环境下培养,应用改良苯酚品红染色法制片.发现其唾腺染色体在不同的温度培养条件下,分布于染色体上的膨松区域有明显变化,在15℃和27℃下染色体的变化区域明显较19℃和23℃下的多,这些区域都与机体调控和表达的基因相关.这可能是机体对不适生存条件采取的保护措施所引起更多的染色体或基因发生变化的结果.对不同温度条件下部分染色体膨松区域的特殊基因作了简单讨论.     

厚朴酚对果蝇寿命和繁殖力的影响

从中药厚朴中提取的厚朴酚（magnolol）具有很强的抗氧化性,对其进行黑腹果蝇抗衰老和繁殖力试验。结果表明,分别采用0．02％、0．03％和0．05％浓度的厚朴酚对果蝇寿命均有不同程度明显的延长作用,平均延寿率从8．7％到18．5％;对果蝇繁殖力均有不同程度的提高,果蝇子代蛹数显著增加,平均增殖率从70．0％到268．2％,成虫果蝇数量也显著增加,平均增殖率从145．3％到541．5％。表明厚朴酚具有延缓衰老和促进果蝇的繁殖能力,提高生育力和生命活力的作用。