Vorbereitung im Vorfeld der Mikroinjektion: <ol><li> Blut ernähren Mücken: für die Injektion von Montag bis Mittwoch Feed Weibchen auf der vorigen Freitag. Um Donnerstag und Freitag, Futtermittel Frauen am Montag injizieren der gleichen Woche. </li><li> Bereiten Sie den Quartz Nadeln mit dem Programm 2 und isotonischen Puffer (siehe Materialien). </li><li> &quot;Legen tube&quot; für die Embryonen ist die regelmäßige Drosophila Kultur Durchstechflasche. Wet Watte in den Boden, mit einem nassen Scheibe Filterpapier für sie. </li><li> Bereiten Kunststoff Deckglas durch Kleben beidseitig Klebeband an einem Ende. Trim Band Deckglas, so dass es am Rande des Deckglases endet. </li><li> Bereiten Öl für Austrocknung. </li><li> Bereiten Sie eine Petrischale mit isotonischen Puffer, um Embryonen zu Schlüpfen zu übertragen. </li></ol> Set up der erzwungenen Verlegung: <ol><li> Sammeln 6-10 Blut zugeführt Weibchen mit dem Einsatz von einem Aspirator und übergeben sie an die Drosophila Kultur Fläschchen mit Baumwolle und Filterpapier nass mit isotonischen Puffer. </li><li> Die Mücken werden dann wieder in insectery Bedingungen in der Dunkelheit setzen und dürfen Eier für 1 h und 15 min lag. </li><li> Lassen Erwachsenen in den Käfig zu fliegen, und entfernen Sie das Filterpapier Festplatte mit Embryonen. </li><li> Um line up Eier, tun Sie es unter dem Binokular. Sammeln Trauben von Eiern mit einem feinen Pinsel (Sable, No 0000) und Transfer zum einen vorbereiteten Platz 3MM Whatman-Papier mit isotonischen Puffer getränkt. Schützen Sie das Papier alle Zeit nass. Lassen Sie sich nicht ausgetrocknet Eier zu bekommen. Entfernen Sie die exochorion mit dem Pinsel, hilft es, besser kleben Eier auf das Band. </li><li> Mit feinen ForceP Nr. 5 oder einem feinen Pinsel, abholen dunkleres Grau Embryonen und ordnen Linie auf Platz 3MM Whatman nass mit isotonischen Puffer. Line up von 20 bis 30 Embryonen. Alle Embryonen müssen in der gleichen Ausrichtung wie Injektion auf hinteren Pol sein. Das vordere Ende des Embryos ist etwas breiter als der hintere. Dann, mit Streifen 3MM Whatman-Papier, trockenes Filter in denen die Eier durch starken Druck auf beiden Seiten des Eies Linie nicht berühren, werden die Eier gefüttert. </li><li> Zur Übertragung der Eier, drehen Sie die Folie mit dem doppelseitigen Klebeband (Medizin), und sanft gegen die Eier zu drücken. Das hintere Ende der Eier müssen sehr nah an den Rand des doppelseitigem Klebeband. Die Nässe des Papiers ist entscheidend dafür, ob es zu nass werden sie nicht halten wird. Ich weiß Austrocknung des Embryos von 5-10 Sekunden, aber Austrocknung hängt von feuchten und Temperatur in Mikroinjektion Raum. </li><li> Austrocknung ist entscheidend für Eier: etwas zu viel und Embryonen nicht schlüpfen. Ohne Austrocknung DNA wird nicht in Embryos. </li><li> Abdeckung ausgetrocknet Embryonen mit Halocarbonöl weitere Austrocknung zu verhindern. </li></ol> Mikroinjektion der Embryonen: Der wichtigste Aspekt guter Injektion wird die Qualität der Nadel (siehe Anmerkung). <ol><li> Füllen Sie eine Nadel mit der DNA-Lösung, um mit Hilfe eines Microloader (Eppendorf # 5242 956,003) injiziert werden. Sehr wenig Injektion soliution benötigt wird, von 1 bis 2 ml. </li><li> Schließen Sie die Nadel auf die Eppendorf Transjector, die Einspritzzeit und Druck steuert, sowie der Gegendruck. Mikroinjektion wird mit einem Mikroskop mit einer beweglichen Bühne (Leica) bei 10facher Vergrößerung und Mikromanipulator (Leica). Falls erforderlich, kann eine erhöhte Mikroskoptisch durch Stapeln 05.04 Objektträger vorbereitet werden. Legen Sie das Deckglas tragen die Embryonen auf die erhöhte Bühne. Die Embryonen werden in einem 150 ° Winkel injiziert; Penetration sollte am hinteren Pol sein. Für die Injektion, halte ich die Nadel stationär und bewegen die Bühne des Mikroskops. Wenn die Nadel neu ist, ist die Spitze verschlossen und in der Regel bricht in der ersten Injektion. Ich arbeite immer aus den Druck benötigt wird, um ein kleines Tröpfchen von DNA in Öl zu vertreiben, zwischen jeder Injektion. </li><li> Ich habe die Einspritzzeit um 0,2 bis 0,9 sec und 1000 hPa, wenn die Nadel ist neu. Ich variieren den Druck und die Zeit, bis ein kleiner Tropfen ist zu sehen, die sich aus der Nadel in das Öl. Der Gegendruck muss rund 100 eingestellt werden. Wie der Nadelspitze wird getragen hat den Einspritzdruck reduziert werden, um das Injektionsvolumen niedrig (ca. 300 hPa) zu halten. Danach wird die Nadelspitze zu groß und tötet die meisten der Embryonen. </li><li> Nach der Injektion zu entfernen das Öl mit Seidenpapier und decken die Rutsche mit Embryonen und in der Petrischale mit isotonischen Puffer. Die Schalen werden in der Insektarium und sie dort lassen, bis die Embryonen schlüpfen. Sie fangen an, nach 2-6 Tagen ausbrüten. Übertragen Sie die Larven in destilliertes Wasser mit der Prise gemahlener Fischfutter. </li></ol> Hinweise DNA für die Injektion: Plasmid-DNA-Lösung zur Injektion wurde unter Verwendung Endofree Plasmid-Kit.Construct Plasmid und Helferplasmids vermischen sich in richtigen Konzentration, mit Isopropanol gefällt. Das Pellet wurde mit 70% Ethanol, bevor Resuspendieren in Injektionspuffer gewaschen. Vor der Injektion, saubere DNA mit Millex-GV-Spalte. Izotonic Puffer: <table border="1" cellpadding="8"><tbody><tr><td> 1,68 m NaCl - 88,3 ml 1,68 m NaCl - 2,9 m 1 M Hepes -10,7 ml 1,12 m CaCl 2 bis 2,2 ml dH 2 O - 896 ml </td><td> OR </td><td> 5 M NaCl - 29,67 ml 1 M KCl - 4.87 ml 1 M Hepes - 10,7 ml 1,12 m CaCl 2 bis 2,2 ml dH 2 O - 952,56 ml </td></tr></tbody></table> Passen Sie auf pH 7,2

<table><tbody><tr><td>Injection Quartz needles</td><td></td><td>Sutter Instrument Co.</td><td></td><td>O.D.- 1.0 mm, I.D.-0.70, 10 cm length, are prepared by drawing capillary tubing into a fine 3-8 um with a shaft of approximately 100-300 mm micropipette puller. </td></tr><tr><td>Quartz puller</td><td>Tool</td><td>Sutter Instrument Co.</td><td>P-2000</td><td>Set-up: Heat-275, Fil-3, Vel-38, Del-250, Pull-141</td></tr><tr><td>EndoFree plasmid kit</td><td></td><td>Qiagen</td><td></td><td></td></tr><tr><td>Isopropanol</td><td></td><td></td><td></td><td></td></tr><tr><td>Millex-GV column</td><td></td><td></td><td>SLGV RO4 NL</td><td>for DNA cleaning</td></tr><tr><td>Construct DNA</td><td></td><td></td><td></td><td>0.5 mg/ml </td></tr><tr><td>Helper plasmid</td><td></td><td></td><td></td><td>0.3 mg/ml </td></tr><tr><td>Injection solution</td><td>Buffer</td><td></td><td></td><td>1x solution: 5mM KCl, 0.1mM sodium phosphate, ph 6. 8</td></tr><tr><td>Isotonic buffer</td><td></td><td></td><td></td><td>see recipe in the protocol section</td></tr><tr><td>Dessication oil</td><td></td><td></td><td></td><td>Halocarbon oil 700:Halocarbon oil 27(1:1). Mix well. Prepare in advance of microinjection.</td></tr><tr><td>Blood</td><td></td><td></td><td></td><td>to feed mosquitos</td></tr><tr><td>Anopheles stephensi</td><td>Animal</td><td></td><td></td><td>Mosquitos, male and female.</td></tr><tr><td>Drosophila culture vial</td><td></td><td></td><td></td><td>for egg laying</td></tr><tr><td>Petri dishes</td><td></td><td></td><td></td><td>with isotonic buffer, for embryo hatching</td></tr><tr><td>Fine paintbrush</td><td></td><td></td><td></td><td>Sable # 0000</td></tr><tr><td>3MM Whatman paper</td><td></td><td></td><td></td><td></td></tr><tr><td>Forceps #5</td><td></td><td></td><td></td><td></td></tr><tr><td>double sided tape</td><td></td><td></td><td></td><td></td></tr><tr><td>Microloader</td><td></td><td>Eppendorf</td><td>5242 956.003</td><td></td></tr><tr><td>Transjector</td><td></td><td>Eppendorf</td><td></td><td></td></tr><tr><td>Microscope</td><td></td><td>Leica Microsystems</td><td></td><td>with moving stage and micromanipulator, set at 10x magnification </td></tr></tbody></table>

injection of an. stephensi embryos to generate malaria-resistant mosquitoes

Die Einführung von exogenen Genen in den Genomen von Moskitos erfordert Mikroinjektion Techniken abgestimmt auf die spezifischen Arten von Interesse. Dieses Video-Protokoll beschreibt eine Methode durch die James-Labor verwendet werden, um DNA-Konstrukte in Anopheles stephensi Embryonen für die Erzeugung von transformierten Mücken microinject. Techniken für die Herstellung Mikroinjektion Nadeln, Sammlung und Aufbereitung von Embryonen und die Durchführung der Mikroinjektion dargestellt.

Watch this Scientific Journal Video about Injection of An. stephensi Embryos to Generate Malaria-resistant Mosquitoes at JoVE.com

Injection of An. stephensi Embryos to Generate Malaria-resistant Mosquitoes

Anopheles stephensi Mücken sind Vektoren für Malaria bewohnen Indien und ganz Asien. Dieses Video zeigt die Technik für die Durchführung microinjections dieser Art mit Transgene, dass Widerstand gegen die Malaria, die Mücke wird zu verleihen. Ein großer Teil der Methodik in diesem Video gezeigt, ist für die Mikroinjektion Techniken anderer Mückenarten.

Injektion eines. stephensi Embryonen zu generieren Malaria-resistenter Moskitos

The introduction of exogenous genes into the genomes of mosquitoes requires microinjection techniques tailored to the specific species of interest. This ...

injection-an-stephensi-embryos-to-generate-malaria-resistant

Research

JoVE Journal

Biology

12.9K Aufrufe.  University of California, Irvine (UCI). Anopheles stephensi Mücken sind Vektoren für Malaria bewohnen Indien und ganz Asien. Dieses Video zeigt die Technik für die Durchführung microinjections dieser Art mit Transgene, dass Widerstand gegen die Malaria, die Mücke wird zu verleihen. Ein großer Teil der Methodik in diesem Video gezeigt, ist für die Mikroinjektion Techniken anderer Mückenarten.

Serie: Injection of An. stephensi Embryos to Generate Malaria-resistant Mosquitoes

Protocol for Production of a Genetic Cross of the Rodent Malaria Parasites

Variation in response to antimalarial drugs and in pathogenicity of malaria parasites is of biologic and medical importance. Linkage mapping has led to successful identification of genes or loci underlying various traits in malaria parasites of rodents1-3 and humans4-6. The malaria parasite Plasmodium yoelii is one of many malaria species isolated from wild African rodents and has been adapted to grow in laboratories. This species reproduces many of the biologic characteristics of the human malaria parasites; genetic markers such as microsatellite and amplified fragment length polymorphism (AFLP) markers have also been developed for the parasite7-9. Thus, genetic studies in rodent malaria parasites can be performed to complement research on Plasmodium falciparum. Here, we demonstrate the techniques for producing a genetic cross in P. yoelii that were first pioneered by Drs. David Walliker, Richard Carter, and colleagues at the University of Edinburgh10.

Genetic crosses in P. yoelii and other rodent malaria parasites are conducted by infecting mice Mus musculus with an inoculum containing gametocytes of two genetically distinct clones that differ in phenotypes of interest and by allowing mosquitoes to feed on the infected mice 4 days after infection. The presence of male and female gametocytes in the mouse blood is microscopically confirmed before feeding. Within 48 hrs after feeding, in the midgut of the mosquito, the haploid gametocytes differentiate into male and female gametes, fertilize, and form a diploid zygote (Fig. 1). During development of a zygote into an ookinete, meiosis appears to occur11. If the zygote is derived through cross-fertilization between gametes of the two genetically distinct parasites, genetic exchanges (chromosomal reassortment and cross-overs between the non-sister chromatids of a pair of homologous chromosomes; Fig. 2) may occur, resulting in recombination of genetic material at homologous loci. Each zygote undergoes two successive nuclear divisions, leading to four haploid nuclei. An ookinete further develops into an oocyst. Once the oocyst matures, thousands of sporozoites (the progeny of the cross) are formed and released into mosquito hemoceal. Sporozoites are harvested from the salivary glands and injected into a new murine host, where pre-erythrocytic and erythrocytic stage development takes place. Erythrocytic forms are cloned and classified with regard to the characters distinguishing the parental lines prior to genetic linkage mapping. Control infections of individual parental clones are performed in the same way as the production of a genetic cross.

Genetic crosses of rodent malaria parasites are performed by feeding two genetically distinct parasites to mosquitoes. Recombinant progeny are cloned from mouse blood after allowing mosquitoes to bite infected mice. This video shows how to produce genetic crosses of Plasmodium yoelii and is applicable to other rodent malaria parasites.

Protokoll für die Herstellung eines genetischen Kreuz der Nager Malaria-Erregern

Variation in response to antimalarial drugs and in pathogenicity of malaria parasites is of biologic and medical importance. Linkage mapping has led to ...

Forschung

Immunologie und Infektion

Microinjection of A. aegypti Embryos to Obtain Transgenic Mosquitoes

In this video, Nijole Jasinskiene demonstrates the methodology employed to generate transgenic Aedes aegypti mosquitoes, which are vectors for dengue fever.  The techniques for correctly preparing microinjection needles, dessicating embryos, and performing microinjection are demonstrated.

In this video, Nijole Jasinskiene demonstrates the methodology employed to generate transgenic Aedes aegypti mosquitoes, which are vectors for dengue fever. The techniques for correctly preparing microinjection needles, dessicating embryos, and performing microinjection are demonstrated.

Mikroinjektion von A. aegypti Embryonen, um transgene Moskitos erhalten

In this video, Nijole Jasinskiene demonstrates the methodology employed to generate transgenic Aedes aegypti mosquitoes, which are vectors for dengue ...

Biologie

Site-Directed &#966;C31-Mediated Integration and Cassette Exchange in Anopheles Vectors of Malaria

Functional genomic analysis and related strategies for genetic control of malaria rely on validated and reproducible methods to accurately modify the genome of Anopheles mosquitoes. Amongst these methods, the &#966;C31 system allows precise and stable site-directed integration of transgenes, or the substitution of integrated transgenic cassettes via recombinase-mediated cassette exchange (RMCE). This method relies on the action of the Streptomyces &#966;C31 bacteriophage integrase to catalyze recombination between two specific attachment sites designated attP (derived from the phage) and attB (derived from the host bacterium). The system uses one or two attP sites that have been integrated previously into the mosquito genome and attB site(s) in the donor template DNA. Here we illustrate how to stably modify the genome of attP-bearing Anopheles docking lines using two plasmids: an attB-tagged donor carrying the integration or exchange template and a helper plasmid encoding the &#966;C31 integrase. We report two representative results of &#966;C31-mediated site-directed modification: the single integration of a transgenic cassette in An. stephensi and RMCE in An. gambiae mosquitoes. &#966;C31-mediated genome manipulation offers the advantage of reproducible transgene expression from validated, fitness neutral genomic sites, allowing comparative qualitative and quantitative analyses of phenotypes. The site-directed nature of the integration also substantially simplifies the validation of the single insertion site and the mating scheme to obtain a stable transgenic line. These and other characteristics make the &#966;C31 system an essential component of the genetic toolkit for the transgenic manipulation of malaria mosquitoes and other insect vectors.

The protocol describes how to achieve site-directed modifications in the genome of Anopheles malaria mosquitoes using the &#966;C31 system. Modifications described include both the integration and the exchange of transgenic cassettes in the genome of attP-bearing docking lines.

Ortsgerichtete φC31-vermittelte Integration und Kassettenaustausch in Anopheles-Vektoren der Malaria

Functional genomic analysis and related strategies for genetic control of malaria rely on validated and reproducible methods to accurately modify the ...

Genetik

Small-Cage Laboratory Trials of Genetically-Engineered Anopheline Mosquitoes

Control of mosquito-borne pathogens using genetically-modified vectors has been proposed as a promising tool to complement conventional control strategies. CRISPR-based homing gene drive systems have made transgenic technologies more accessible within the scientific community. Evaluation of transgenic mosquito performance and comparisons with wild-type counterparts in small laboratory cage trials provide valuable data for the design of subsequent field cage experiments and experimental assessments to refine the strategies for disease prevention. Here, we present three different protocols used in laboratory settings to evaluate transgene spread in anopheline mosquito vectors of malaria. These include inundative releases (no gene-drive system), and gene-drive overlapping and non-overlapping generation trials. The three trials vary in a number of parameters and can be adapted to desired experimental settings. Moreover, insectary studies in small cages are part of the progressive transition of engineered insects from the laboratory to open field releases. Therefore, the protocols described here represent invaluable tools to provide empirical values that will ultimately aid field implementation of new technologies for malaria elimination.

The protocols reported here illustrate three alternative ways to assess the performance of genetically-engineered mosquitoes destined for vector control in laboratory-contained small cage trials. Each protocol is tailored to the specific modification the mosquito strain bears (gene drive or non-gene drive) and the types of parameters measured.

Kleinkäfig-Laborversuche mit gentechnisch veränderten Anopheline-Mücken

Control of mosquito-borne pathogens using genetically-modified vectors has been proposed as a promising tool to complement conventional control ...

Microinjection Method for Anopheles gambiae Embryos

Embryo microinjection techniques are essential for many molecular and genetic studies of insect species. They provide a means to introduce exogenous DNA fragments encoding genes of interest as well as favorable traits into the insect germline in a stable and heritable manner. The resulting transgenic strains can be studied for phenotypic changes resulting from the expression of the integrated DNA to answer basic questions or used in practical applications. Although the technology is straightforward, it requires of the investigator patience and practice to achieve a level of skill that maximizes efficiency. Shown here is a method for microinjection of embryos of the African malaria mosquito, Anopheles gambiae. The objective is to deliver by microinjection exogenous DNA to the embryo so that it can be taken up in the developing germline (pole) cells. Expression from the injected DNA of transposases, integrases, recombinases, or other nucleases (for example CRISPR-associated proteins, Cas) can trigger events that lead to its covalent insertion into chromosomes. Transgenic An. gambiae generated from these technologies have been used for basic studies of immune system components, genes involved in blood-feeding, and elements of the olfactory system. In addition, these techniques have been used to produce An. gambiae strains with traits that may help control the transmission of malaria parasites.

Microinjection Method for Anopheles gambiae Embryos

Microinjection techniques are essential to introduce exogenous genes into the genomes of mosquitoes. This protocol explains a method used by the James laboratory to microinject DNA constructs into Anopheles gambiae embryos to generate transformed mosquitoes.

Mikroinjektionsmethode für Anopheles gambiae Embryonen

Embryo microinjection techniques are essential for many molecular and genetic studies of insect species. They provide a means to introduce exogenous DNA ...

Inoculating Anopheles gambiae Mosquitoes with Beads to Induce and Measure the Melanization Immune Response

The stimulation of immune responses is a common tool in invertebrate studies to examine the efficacy and the mechanisms of immunity. This stimulation is based on the injection of non-pathogenic particles into insects, as the particles will be detected by the immune system and will induce the production of immune effectors. We focus here on the stimulation of the melanization response in the mosquito Anopheles gambiae. The melanization response results in the encapsulation of foreign particles and parasites with a dark layer of melanin. To stimulate this response, mosquitoes are inoculated with beads in the thoracic cavity using microcapillary glass tubes. Then, after 24 hr, the mosquitoes are dissected to retrieve the beads. The degree of melanization of the bead is measured using image analysis software. Beads do not have the pathogenic effects of parasites, or their capacity to evade or suppress the immune response. These injections are a way to measure immune efficacy and the impact of immune stimulations on other life history traits, such as fecundity or longevity. It is not exactly the same as directly studying host-parasite interactions, but it is an interesting tool to study immunity and its evolutionary ecology.

Inoculating Anopheles gambiae Mosquitoes with Beads to Induce and Measure the Melanization Immune Response

Through inoculation with beads, the described technique enables the stimulation of the mosquito melanization response in the hemolymph circulating system. The amount of melanin covering the beads can be measured after dissection as a measure of the immune response.

Impfen<em&gt; Anopheles gambiae</em&gt; Mosquitoes mit Perlen der Melanisierung Immunantwort zu induzieren und Messen

The stimulation of immune responses is a common tool in invertebrate studies to examine the efficacy and the mechanisms of immunity. This stimulation is ...

Generating Genetically Modified Plasmodium berghei Sporozoites

Malaria is a deadly disease caused by the parasite Plasmodium and is transmitted through the bite of female Anopheles mosquitoes. The sporozoite stage of Plasmodium deposited by mosquitoes in the skin of vertebrate hosts undergoes a phase of mandatory development in the liver before initiating clinical malaria. We know little about the biology of Plasmodium development in the liver; access to the sporozoite stage and the ability to genetically modify such sporozoites are critical tools for studying the nature of Plasmodium infection and the resulting immune response in the liver. Here, we present a comprehensive protocol for the generation of transgenic Plasmodium berghei sporozoites. We genetically modify blood-stage P. berghei and use this form to infect Anopheles mosquitoes when they take a blood meal. After the transgenic parasites undergo development in the mosquitoes, we isolate the sporozoite stage of the parasite from the mosquito salivary glands for in vivo and in vitro experimentation. We demonstrate the validity of the protocol by generating sporozoites of a novel strain of P. berghei expressing the green fluorescent protein (GFP) subunit 11 (GFP11), and show how it could be used to investigate the biology of liver-stage malaria.

Generating Genetically Modified Plasmodium berghei Sporozoites

Malaria is transmitted through inoculation of the sporozoite stage of Plasmodium by infected mosquitoes. Transgenic Plasmodium has allowed us to understand the biology of malaria better and has contributed directly to malaria vaccine development efforts. Here, we describe a streamlined methodology to generate transgenic Plasmodium berghei sporozoites.

Gentechnisch veränderte Plasmodium berghei Sporozoiten

Malaria is a deadly disease caused by the parasite Plasmodium and is transmitted through the bite of female Anopheles mosquitoes. The sporozoite stage of ...

Fluoreszenz-In-situ-Hybridisierung zur Untersuchung der Spermatogenese von Mücken

Whole-Mount Fluorescence In Situ Hybridization to Study Spermatogenesis in the Anopheles Mosquito

Spermatogenesis is a complex biological process during which diploid cells undergo successive mitotic and meiotic division followed by large structural changes to form haploid spermatozoa. Besides the biological aspect, studying spermatogenesis is of paramount importance for understanding and developing genetic technologies such as gene drive and synthetic sex ratio distorters, which, by altering Mendelian inheritance and the sperm sex ratio, respectively, could be used to control pest insect populations. These technologies have proven to be very promising in lab settings and could potentially be used to control wild populations of Anopheles mosquitoes, which are vectors of malaria. Due to the simplicity of the testis anatomy and their medical importance, Anopheles gambiae, a major malaria vector in sub-Saharan Africa, represents a good cytological model for studying spermatogenesis. This protocol describes how whole-mount fluorescence in situ hybridization (WFISH) can be used to study the dramatic changes in cell nuclear structure through spermatogenesis using fluorescent probes that specifically stain the X and Y chromosomes. FISH usually requires the disruption of the reproductive organs to expose mitotic or meiotic chromosomes and allow the staining of specific genomic regions with fluorescent probes. WFISH enables the preservation of the native cytological structure of the testis, coupled with a good level of signal detection from fluorescent probes targeting repetitive DNA sequences. This allows researchers to follow changes in the chromosomal behavior of cells undergoing meiosis along the structure of the organ, where each phase of the process can clearly be distinguished. This technique could be particularly useful for studying chromosome meiotic pairing and investigating the cytological phenotypes associated with, for example, synthetic sex ratio distorters, hybrid male sterility, and the knock-out of genes involved in spermatogenesis.

Autor im Rampenlicht: Whole-Mount Fluoreszenz In situ Hybridisierung zur Untersuchung der Spermatogenese in der Anopheles-Mücke  

Given their simple anatomy, Anopheles testes offer a good cytological model for studying spermatogenesis. This protocol describes whole-mount fluorescence in situ hybridization, a technique used to investigate this biological process, as well as the phenotype of transgenic strains harboring mutations in the genes involved in sperm production.

Ganzkörper-Fluoreszenz-in-situ-Hybridisierung zur Untersuchung der Spermatogenese in der Anopheles-Mücke

Spermatogenesis is a complex biological process during which diploid cells undergo successive mitotic and meiotic division followed by large structural ...

Injektion eines. stephensi Embryonen zu generieren Malaria-resistenter Moskitos

Zusammenfassung

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