Name: protocol for mosquito rearing (a. gambiae)
Uploaded: 2007-07-04T01:00:00
Description: Watch this Scientific Journal Video about Protocol for Mosquito Rearing A. gambiae at JoVE.com

蚊子的食物： <ol><li>食品答：接地的鱼食（Aquaricare）。需要添加一小捏。 </li><li>食品乙：接地猫粮（普瑞纳）。需要添加一小捏。 </li><li>食品：猫食品（普瑞纳）。两片需要添加。 </li></ol>血型： 动物或人体血液中可用于后方蚊子。收集了新鲜的血液是用注射器，并把在无菌15毫升猎鹰，每10毫升血液中含有的CPD（抗凝）1毫升管，然后轻轻地混合，在室温下...

<ol>
	<li>Bangs, M. J., Soelarto, T., Barodji, B. P., Wicaksana, D. T., Boewono, <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Colonization+of+Anopheles+maculatus+from+Central+Java,+Indonesia.">Colonization of Anopheles maculatus from Central Java, Indonesia.</a> J Am Mosq Control Assoc. 18, 359-363 (2002).</li><li>Beier, M. S., Beier, J. C., Merdan, A. A., el Sawaf, B. M., Kadder, M. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Laboratory+rearing+techniques+and+adult+life+table+parameters+for+Anopheles+sergentii+from+Egypt.">Laboratory rearing techniques and adult life table parameters for Anopheles sergentii from Egypt.</a> J Am Mosq Control Assoc. 266, 266-270 (1987).</li><li>Benedict, M. Q., Crampton, J. M., Beard, C. B., Louis, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Care and maintenance of anopheline mosquitoes, The molecular biology of disease vectors: A methods manual. , 3-12 (1997).</li><li>Casanges, A. H., Mcgovran, E. R., Chiles, J. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rearing+of+Anopheles+quadrimaculatus+Say+and+Aedes+aegypti+(L.)+in+the+laboratory.">Rearing of Anopheles quadrimaculatus Say and Aedes aegypti (L.) in the laboratory.</a> Mosq News. 9, 112-117 (1949).</li><li>Merritt, R. W., Dadd, R. H., Walker, E. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Feeding+behavior,+natural+food+and+nutritional+relation+ships+of+larval+mosquitoes.">Feeding behavior, natural food and nutritional relation ships of larval mosquitoes.</a> Annual Rev. Entomol. 37, 349-376 (1992).</li><li>Nasirian, H., Ladonni, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Artificial+bloodfeeding+of+Anopheles+stephensi+on+a+membrane+apparatus+with+human+whole+blood.">Artificial bloodfeeding of Anopheles stephensi on a membrane apparatus with human whole blood.</a> J Am Mosq Control Assoc. 22, 54-56 (2006).</li><li>Ng'habi, K. R., John, B., Nkwengulila, G., Knols, B. G., Killeen, G. F., Ferguson, H. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effect+of+larval+crowding+on+mating+competitiveness+of+Anopheles+gambiae+mosquitoes.">Effect of larval crowding on mating competitiveness of Anopheles gambiae mosquitoes.</a> Malar J. 30 (4), 49 (2005).</li><li>Stobart, R. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Factors+affecting+the+control+of+body+volume+in+the+larvae+of+the+mosquitoes+Aedes+aegypti+and+Aedes+detritus.">Factors affecting the control of body volume in the larvae of the mosquitoes Aedes aegypti and Aedes detritus.</a> Edw. J. Exp. Boil. 54, 67-82 (1971).</li></ol>

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		<table border="1">
		<thead>
		<tr>
		<th>Material Name</th>
		<th>Type</th>
		<th>Company</th>
		<th>Catalogue Number</th>
		<th>Comment</th>
		</tr>
		</thead>
		<tbody>
		
<tr>
<td>Food A</td>
<td>&nbsp;</td>
<td>Aquaricare</td>
<td>&nbsp;</td>
<td>Grounded fish food. A small pinch needs to be added.</td>
</tr>
<tr>
<td>Food B</td>
<td>&nbsp;</td>
<td>Purina</td>
<td>&nbsp;</td>
<td>Grounded cat food. A small pinch needs to be added.</td>
</tr>
<tr>
<td>Food C</td>
<td>&nbsp;</td>
<td>Purina</td>
<td>&nbsp;</td>
<td>Cat food. Two tablets needs to be added</td>
</tr>
<tr>
<td>RBC</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>prepared red blood cells: 40% hematocrit in human serum</td>
</tr>
<tr>
<td>RPMI</td>
<td>Medium</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>CPD</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>anticoagulant</td>
</tr>
<tr>
<td>serum</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>O+ human serum</td>
</tr>
<tr>
<td>Mosquito feeder</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>parafilm</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>Anopheles gambiae</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>mosquitos</td>
</tr>
<tr>
<td>10% sucrose</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>mosquito food</td>
</tr>
<tr>
<td>insectary</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>		</tbody>
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protocol for mosquito rearing (a. gambiae)

这个协议描述在insectary蚊虫饲养。 insectary客房均保持在28 ° C和〜80％湿度，12小时，。日/夜循环。对于此过程，你需要蚊子的笼子里，无菌10％的蔗糖溶液，擦手纸，烧杯，滤纸过滤纸，玻璃进料器，人体血液和血清，水浴，封口膜，蒸馏水，干净的塑料托盘，蚊子的食物（描述下文），蚊帐覆盖的托盘，真空，并收集室，收集成人。

Watch this Scientific Journal Video about Protocol for Mosquito Rearing A. gambiae at JoVE.com

Protocol for Mosquito Rearing A. gambiae

该视频演示了在实验室中使用背面按蚊的一般技术。为照顾实验室蚊子的方法证明通过从幼虫到蛹吸血成年人机体的生命周期的所有阶段。

蚊子养育的议定书“（冈比亚按蚊）

This protocol describes mosquito rearing in the insectary. The insectary rooms are maintained at 28&deg;C and ~80% humidity, with a 12 hr. day/night ...

Research

JoVE Journal

Biology

Protocol for Mosquito Rearing (A. gambiae)

Protocol for RNAi Assays in Adult Mosquitoes (A. gambiae)

Reverse genetic approaches have proven extremely useful for determining  which genes underly resistance to vector pathogens in mosquitoes.  This video protocol illustrates a method used by the Dimopoulos lab to inject dsRNA into Anopheles gambiae mosquitoes, which harbor the malaria parasite.  The technique manipulating the injection setup and injecting dsRNA into the thorax is illustrated.

Protocol for RNAi Assays in Adult Mosquitoes A. gambiae

Reverse genetic approaches have proven extremely useful for determining which genes underly resistance to vector pathogens in mosquitoes. This video protocol illustrates a method used by the Dimopoulos lab to inject dsRNA into Anopheles gambiae mosquitoes, which harbor the malaria parasite. The technique manipulating the injection setup and injecting dsRNA into the thorax is illustrated.

成蚊（冈比亚按蚊的RNAi实验议定书）

Reverse genetic approaches have proven extremely useful for determining  which genes underly resistance to vector pathogens in mosquitoes.  This video ...

科研

生物学

Building a Better Mosquito: Identifying the Genes Enabling Malaria and Dengue Fever Resistance in A. gambiae and A. aegypti Mosquitoes

In this interview, George Dimopoulos focuses on the physiological mechanisms used by mosquitoes to combat Plasmodium falciparum and dengue virus infections.   Explanation is given for how key refractory genes, those genes conferring resistance to vector pathogens, are identified in the mosquito and how this knowledge can be used to generate transgenic mosquitoes that are unable to carry the malaria parasite or dengue virus.

In this interview, George Dimopoulos focuses on the physiological mechanisms used by mosquitoes to combat Plasmodium falciparum and dengue virus infections. Explanation is given for how key refractory genes, those genes conferring resistance to vector pathogens, are identified in the mosquito and how this knowledge can be used to generate transgenic mosquitoes that are unable to carry the malaria parasite or dengue virus.

建设一个更美好的蚊子：确定基因在冈比亚按蚊和埃及斑蚊蚊子启用疟疾和登革热的电阻

In this interview, George Dimopoulos focuses on the physiological mechanisms used by mosquitoes to combat Plasmodium falciparum and dengue virus ...

A Protocol for the Production of KLRG1 Tetramer

Killer cell lectin-like receptor G1 (KLRG1) is a type II transmembrane glycoprotein inhibitory receptor belonging to the C type lectin-like superfamily. KLRG1 exists both as a monomer and as a disulfide-linked homodimer. This well-conserved receptor is found on the most mature and recently activated NK cells as well as on a subset of effector/memory T cells.
Using KLRG1 tetramer as well as other methods, E-, N-, and R-cadherins were identified as KLRG1 ligands. These Ca2+-dependent cell-cell adhesion molecules comprises of an extracellular domain containing five cadherin repeats responsible for cell-cell interactions, a transmembrane domain and a cytoplasmic domain that is linked to the actin cytoskeleton. 
Generation of the KLRG1 tetramer was essential to the identification of the KLRG1 ligands. KLRG1 tetramer is also a unique tool to elucidate the roles cadherin and KLRG1 play in regulating the immune response and tissue integrity.

This protocol describes the production of KLRG1 tetramer, which is a powerful tool for the analysis of KLRG1 ligands.

KLRG1四聚体的生产协议

Killer cell lectin-like receptor G1 (KLRG1) is a type II transmembrane glycoprotein inhibitory receptor belonging to the C type lectin-like superfamily. ...

An Optimized Protocol for Rearing Fopius arisanus, a Parasitoid of Tephritid Fruit Flies

Fopius arisanus (Sonan) is an important parasitoid of Tephritid fruit flies for at least two reasons. First, it is the one of only three opiine parasitoids known to infect the host during the egg stage1. Second, it has a wide range of potential fruit fly hosts. Perhaps due to its life history, F. arisanus has been a successfully used for biological control of fruit flies in multiple tropical regions2-4. One impediment to the wide use of F. arisanus for fruit fly control is that it is difficult to establish a stable laboratory colony5-9. Despite this difficulty, in the 1990s USDA researchers developed a reliable method to maintain laboratory populations of F. arisanus10-12. There is significant interest in F. arisanus biology13,14, especially regarding its ability to colonize a wide variety of Tephritid hosts14-17; interest is especially driven by the alarming spread of Bactrocera fruit fly pests to new continents in the last decade18. Further research on F. arisanus and additional deployments of this species as a biological control agent will benefit from optimizations and improvements of rearing methods. In this protocol and associated video article we describe an optimized method for rearing F. arisanus based on a previously described approach12. The method we describe here allows rearing of F. arisanus in a small scale without the use of fruit, using materials available in tropical regions around the world and with relatively low manual labor requirements.

An Optimized Protocol for Rearing Fopius arisanus, a Parasitoid of Tephritid Fruit Flies

Fopius arisanus is an egg-larval parasitoid of Tephritid fruit flies that is successfully used in biological control of these important tropical pests. We describe here an optimized protocol for rearing F. arisanus on a small scale using readily available materials.

一个优化的协议饲养 Fopius arisanus Tephritid果寄生蝇

Fopius arisanus (Sonan) is an important parasitoid of Tephritid fruit flies for at least two reasons. First, it is the one of only three opiine ...

SIVQ-LCM Protocol for the ArcturusXT Instrument

SIVQ-LCM is a new methodology that automates and streamlines the more traditional, user-dependent laser dissection process.&#160;It aims to create an advanced, rapidly customizable laser dissection platform technology. In this report, we describe the integration of the image analysis software Spatially Invariant Vector Quantization (SIVQ) onto the ArcturusXT instrument. The ArcturusXT system contains both an infrared (IR) and ultraviolet (UV) laser, allowing for specific cell or large area dissections. The principal goal is to improve the speed, accuracy, and reproducibility of the laser dissection to increase sample throughput. This novel approach facilitates microdissection of both animal and human tissues in research and clinical workflows.

SIVQ-LCM is an innovative approach that harnesses a computer algorithm, Spatially Invariant Vector Quantization (SIVQ), to drive the laser capture microdissection (LCM) process. The SIVQ-LCM workflow greatly improves the speed and accuracy of microdissection, with applications in both the research and clinical settings.

为ArcturusXT仪器SIVQ-LCM协议

SIVQ-LCM is a new methodology that automates and streamlines the more traditional, user-dependent laser dissection process.&#160;It aims to create an ...

An Experimental and Bioinformatics Protocol for RNA-seq Analyses of Photoperiodic Diapause in the Asian Tiger Mosquito, Aedes albopictus

Photoperiodic diapause is an important adaptation that allows individuals to escape harsh seasonal environments via a series of physiological changes, most notably developmental arrest and reduced metabolism. Global gene expression profiling via RNA-Seq can provide important insights into the transcriptional mechanisms of photoperiodic diapause. The Asian tiger mosquito, Aedes albopictus, is an outstanding organism for studying the transcriptional bases of diapause due to its ease of rearing, easily induced diapause, and the genomic resources available. This manuscript presents a general experimental workflow for identifying diapause-induced transcriptional differences in A. albopictus. Rearing techniques, conditions necessary to induce diapause and non-diapause development, methods to estimate percent diapause in a population, and RNA extraction and integrity assessment for mosquitoes are documented. A workflow to process RNA-Seq data from Illumina sequencers culminates in a list of differentially expressed genes. The representative results demonstrate that this protocol can be used to effectively identify genes differentially regulated at the transcriptional level in A. albopictus due to photoperiodic differences. With modest adjustments, this workflow can be readily adapted to study the transcriptional bases of diapause or other important life history traits in other mosquitoes.

An Experimental and Bioinformatics Protocol for RNA-seq Analyses of Photoperiodic Diapause in the Asian Tiger Mosquito, Aedes albopictus

RNA-Seq analyses are becoming increasingly important for identifying the molecular underpinnings of adaptive traits in non-model organisms. Here, a protocol to identify differentially expressed genes between diapause and non-diapause Aedes albopictus mosquitoes is described, from mosquito rearing, to RNA sequencing and bioinformatics analyses of RNA-Seq data.

实验和生物信息学议定书光周期滞育的亚洲虎蚊的RNA序列分析，<em&gt;白纹伊蚊</em

Photoperiodic diapause is an important adaptation that allows individuals to escape harsh seasonal environments via a series of physiological changes, ...

RNAi Trigger Delivery into Anopheles gambiae Pupae

RNA interference (RNAi), a naturally occurring phenomenon in eukaryotic organisms, is an extremely valuable tool that can be utilized in the laboratory for functional genomic studies. The ability to knockdown individual genes selectively via this reverse genetic technique has allowed many researchers to rapidly uncover the biological roles of numerous genes within many organisms, by evaluation of loss-of-function phenotypes. In the major human malaria vector Anopheles gambiae, the predominant method used to reduce the function of targeted genes involves injection of double-stranded (dsRNA) into the hemocoel of the adult mosquito. While this method has been successful, gene knockdown in adults excludes the functional assessment of genes that are expressed and potentially play roles during pre-adult stages, as well as genes that are expressed in limited numbers of cells in adult mosquitoes. We describe a method for the injection of Serine Protease Inhibitor 2 (SRPN2) dsRNA during the early pupal stage and validate SRPN2 protein knockdown by observing decreased target protein levels and the formation of melanotic pseudo-tumors in SRPN2 knockdown adult mosquitoes. This evident phenotype has been described previously for adult stage knockdown of SRPN2 function, and we have recapitulated this adult phenotype by SRPN2 knockdown initiated during pupal development. When used in conjunction with a dye-labeled dsRNA solution, this technique enables easy visualization by simple light microscopy of injection quality and distribution of dsRNA in the hemocoel.

RNAi Trigger Delivery into Anopheles gambiae Pupae

RNA interference (RNAi) is an extremely valuable tool for uncovering gene function. However, the ability to target genes using RNAi during pre-adult stages is limited in the major human malaria vector Anopheles gambiae. We describe an RNAi protocol to reduce gene function via direct injection during pupal development.

RNAi的触发传送到<em&gt;冈比亚按蚊</em&gt;蛹

RNA interference (RNAi), a naturally occurring phenomenon in eukaryotic organisms, is an extremely valuable tool that can be utilized in the laboratory ...

IR-TEx: An Open Source Data Integration Tool for Big Data Transcriptomics Designed for the Malaria Vector Anopheles gambiae

IR-TEx is an application written in Shiny (an R package) that allows exploration of the expression of (as well as assigning functions to) transcripts whose expression is associated with insecticide resistance phenotypes in Anopheles gambiae mosquitoes. The application can be used online or downloaded and used locally by anyone. The local application can be modified to add new insecticide resistance datasets generated from multiple -omics platforms. This guide demonstrates how to add new datasets and handle missing data. Furthermore, IR-TEx can be completely and easily recoded to use-omics datasets from any experimental data, making it a valuable resource to many researchers. The protocol illustrates the utility of IR-TEx in identifying new insecticide resistance candidates using the the microsomal glutathione transferase, GSTMS1, as an example. This transcript is upregulated in multiple pyrethroid resistant populations from C&#244;te D&#39;Ivoire and Burkina Faso. The identification of co-correlated transcripts provides further insight into the putative roles of this gene.

IR-TEx: An Open Source Data Integration Tool for Big Data Transcriptomics Designed for the Malaria Vector Anopheles gambiae

IR-TEx explores insecticide resistance-related transcriptional profiles in the species Anopheles gambiae. Provided here are full instructions for using the application, modifications for exploring multiple transcriptomic datasets, and using the framework to build an interactive database for collections of transcriptomic data from any organism, generated in any platform.

IR-TEx：专为疟疾病媒阿诺菲斯甘巴病设计的大数据转录组学的开源数据集成工具

IR-TEx is an application written in Shiny (an R package) that allows exploration of the expression of (as well as assigning functions to) transcripts ...

A Step-by-Step Guide to Mosquito Electroantennography

Female mosquitoes are the deadliest animals on earth, claiming the lives of more than 1 million people every year due to pathogens they transmit when acquiring a blood-meal. To locate a host to feed on, mosquitoes rely on a wide range of sensory cues, including visual, mechanical, thermal, and olfactory. The study details a technique, electroantennography (EAG), that allows researchers to assess whether the mosquitoes can detect individual chemicals and blends of chemicals in a concentration-dependent manner. When coupled with gas-chromatography (GC-EAG), this technique allows to expose the antennae to a full headspace/complex mixture and determines which chemicals present in the sample of interest, the mosquito can detect. This is applicable to host body odors as well as plant floral bouquets or other ecologically relevant odors (e.g., oviposition sites odorants). Here, we described a protocol that permits long durations of preparation responsiveness time and is applicable to both female and male mosquitoes from multiple genera, including Aedes, Culex, Anopheles, and Toxorhynchites mosquitoes. As olfaction plays a major part in mosquito-host interactions and mosquito biology in general, EAGs and GC-EAG can reveal compounds of interest for the development of new disease vector control strategies (e.g., baits). Complemented with behavioral assays, the valence (e.g., attractant, repellent) of each chemical can be determined.

The present article details a step-by-step protocol for successful and low noise electroantennograms in several genera of mosquitoes, including both females and males.

蚊子电图分步指南

Female mosquitoes are the deadliest animals on earth, claiming the lives of more than 1 million people every year due to pathogens they transmit when ...

A Magnetic-Bead-Based Mosquito DNA Extraction Protocol for Next-Generation Sequencing

A recently published DNA extraction protocol using magnetic beads and an automated DNA extraction instrument suggested that it is possible to extract high quality and quantity DNA from a well-preserved individual mosquito sufficient for downstream whole genome sequencing. However, reliance on an expensive automated DNA extraction instrument can be prohibitive for many laboratories. Here, the study provides a budget-friendly magnetic-bead-based DNA extraction protocol, which is suitable for low to medium throughput. The protocol described here was successfully tested using individual Aedes aegypti mosquito samples. The reduced costs associated with high quality DNA extraction will increase the application of high throughput sequencing to resource limited labs and studies.

Described here is a DNA extraction protocol using magnetic beads to produce high quality DNA extractions from mosquitoes. These extractions are suitable for a downstream next-generation sequencing approach.

用于下一代测序的基于磁珠的蚊子DNA提取协议

A recently published DNA extraction protocol using magnetic beads and an automated DNA extraction instrument suggested that it is possible to extract high ...