Name: protocols for investigating the host-tissue distribution, transmission-mode, and effect on the host fitness of a densovirus in the cotton bollworm
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Bu çalışma Çin Milli Anahtar Temel Araştırma Programı (No 2013CB127602) ve Çin Ulusal Bilim Vakfı (No 31321004) Yaratıcı Araştırma Grupları için Bilim Fonu tarafından desteklenmiştir.

HaDV2 içermeyen Pamuk Yeşilkurt Colony 1. İnşaat <ol><li> Arka pamuk kurdu larvası 14 saat aydınlık / 10 saat karanlık ve% 60 bağıl nem ile, 25 ± 1 ° C &#39;de kontrollü bir büyüme odası veya bir yapay iklim bölmesine bir suni diyetin 32 (H. armigera). </li><li> İyi havalandırma sağlamak için bir yumurtlama alt-tabaka olarak hizmet etmek üzere gazlı bez ile kaplı çift başına plastik bir kafes (10 cm yükseklik, çapı 5 cm) kullanılarak yeni eclosed kelebekl...

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Son birkaç on yılda, böcek-virüs etkileşimlerine çoğu çalışma bal arısı sağlığı 34, 35, 36, insan hastalığının 37 vektörler üzerine odaklanmıştır, bitki 38 virüsleri, ve biyolojik kontrol ajanları 39 gibi büyük bir potansiyele sahip bazı böcek patojenik virüsleri. Biraz dikkat, özellikle lepidopteran haşerelere de, böcekle...

Son birkaç on yıl içinde, örneğin yeni nesil sıralama olarak dizileme teknolojisinin gelişmesi (NGS), bir çok yeni bir DNA ve RNA virüsleri, özellikle patojenik olmayan virüs değil, aynı zamanda, daha önce bilinen virüsler, 1, 2, 3 yeni izolat keşfini kolaylaştırmıştır 4, 5, 6, 7, 8, 9, 10, 11, 12. Model organizma Drosophila melanogaster olarak, 20&#39;den fazla yeni kısmi virüs genomları metagenomic teknikleri 13 kullanılarak tespit edilmiştir. yeni virüsler de dahil olmak üzere pek çok viral diziler, aynı zamanda, örneğin bal arıları, m gibi başka böceklere tanımlanmıştırosquitoes, Asya narenciye psyllid, böcekler, ve birden fazla lepidopteran türler 14, 15, 16, 17, 18, 19, 20, 21. Gelecekte, daha yeni virüsler bu gelişmiş teknolojileri kullanarak böceklerin keşfedilen olacağını beklenebilir; dolayısıyla, virüs konak etkileşimin anlayışımız, 9 buna göre 6 değişebilir. Birçok yeni virüsler Mutualistik ortakları ziyade sıkı patojenler 22 olarak tanımlanan ediliyor çünkü Örneğin, virüs konak etkileşimleri, önceden düşünüldüğünden daha karmaşık olduğu kabul edilir. Örneğin, Dysaphis plantaginea içinde Mutualistik densovirüs DplDNV kanatlı Morf indükler ve<html> virüs 23 yanı sıra konağın dağılımı kolaylaştırmak, hareketlilik artar. Ayrıca, Mutualistik virüsler memeli sağlık, kuraklık ve bitkilerin soğuk toleransı ve bakteriyel enfeksiyonların 24 etkisine ilişkin olarak tarif edilmiştir. Seneca vadi virüsü-001 nöroendokrin kanseri 25 sahiptir, tümör hücrelerine karşı seçici sitotoksisiteye aracılık gösterilmiştir. Hepatit A virüsü enfeksiyonları hepatit C virüsü replikasyonunu baskılamak ve hepatit C 26 kurtarma yol açabilir. Herpesvirüs gecikme bakteriyel enfeksiyon 27 simbiyotik koruma sağlamaktadır. Insan endojen retrovirüs HERV-W zarf glikoproteini dalak nekroz virüsü 28 hücresel direnç indükler. bir mantar endofit gelen Curvularia termal tolerans virüsü (CThTV) bu mantar ve tropikal panik çim arasındaki Mutualistik etkileşiminde yer alanref &quot;&gt; 29. Dolayısıyla, yeni bulunan virüsler ve onların ana arasındaki etkileşimlerin bilgisi onların biyoloji ve yönetimi konusunda yeni bakış açıları oluşturmak gerekir. Ancak, yeni virüsler, akut enfeksiyon tipik belirgin belirtisi sergileyen özellikle gizli virüsler, nadiren var been araştırdık ve biz olan ev sahipleri yeni bulunan virüslerin etkilerini araştırmak için bir boru hattı ve protokoller gerekir. Daha önce, pamuk kılıfı kurdu yeşil kurt içinde yaygınlığını yeni monosense densovirüs yeşil kurt densovirüs (HaDV2) bildirilmiştir ve HaDV2 ve pamuk kılıfı kurdu 30, 31 arasında bir Mutualistik ilişkinin kanıtları sundular. Bu yazıda ayrıntılı olarak HaDV2 ve pamuk kurdu konak arasındaki etkileşimi incelemek için laboratuvar protokol anlatacağız. Burada sunulan protokol de r inceleyerek araştırmacıların son derece alakalı olabilirÖzellikle lepidopteran haşerelere de, diğer oral bulaşan virüslerin ole.

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	<tbody>
		<tr>
			<td>24-well plate</td>
			<td>Corning</td>
			<td>07-200-740</td>
			<td>Multiple suppliers available.</td>
		</tr>
		<tr>
			<td>DNA extraction kit</td>
			<td>TIANGEN</td>
			<td>DP304-03</td>
			<td>Multiple suppliers available.</td>
		</tr>
		<tr>
			<td>thermal cycler</td>
			<td>Veriti; Applied Biosystems</td>
			<td>4375786</td>
			<td></td>
		</tr>
		<tr>
			<td>PBS</td>
			<td>Corning</td>
			<td>21-040-CV</td>
			<td></td>
		</tr>
		<tr>
			<td>0.22 &#181;m membrane filter</td>
			<td>Millipore</td>
			<td>SLGS025NB</td>
			<td></td>
		</tr>
		<tr>
			<td>pEASY-T Cloning Vector</td>
			<td>TransGen, Beijing, China</td>
			<td>CT301-02</td>
			<td></td>
		</tr>
		<tr>
			<td>Tweezers</td>
			<td>IDEAL-TEK</td>
			<td>2.SA</td>
			<td></td>
		</tr>
		<tr>
			<td>Premix Ex Taq (Probe qPCR)</td>
			<td>Takara</td>
			<td>RR390A</td>
			<td></td>
		</tr>
		<tr>
			<td>Probes</td>
			<td>Invitrogen</td>
			<td>Custom order</td>
			<td></td>
		</tr>
		<tr>
			<td>Primers</td>
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</table>

protocols for investigating the host-tissue distribution, transmission-mode, and effect on the host fitness of a densovirus in the cotton bollworm

Birçok yeni virüsler yeni nesil dizileme teknolojileri kullanarak hayvan konakçılarda keşfedildi. Daha önce, bir lepidopteran türlerde, bir Mutualistik virüsü, yeşil kurt densovirüs (HaDV2) bildirilen, pamuk kurdu, yeşil kurt (Hubner). Burada, şu anda kendi ana bilgisayarda HaDV2 etkisini incelemek için kullanılan protokolleri açıklar. İlk olarak, tek bir üreme çifti bir HaDV2 içermeyen pamuk kurdu kolonisi kurmak. Bir HaDV2 enfekte diğer enfekte olmamış: Daha sonra, ağız yoluyla, aynı genetik arkaplana sahip iki koloniler elde etmek için filtre edilmiş sıvının HaDV2 içeren bazı yeni doğmuş larva yavruları inoküle. HaDV2 konak doku dağılımı ve transmisyon verimi belirlemek için protokoller gibi bir protokolü, sunulan HaDV2 enfekte olmuş ve -uninfected bireyler arasında hayat tablosu parametreleri (örneğin, larva, pupa ve yetişkin dönemleri ve doğurganlığı) karşılaştırmak. Bu protokoller, woULD ayrıca haşere, özellikle lepidopteran ana diğer oral yolla bulaşan virüsler etkilerinin araştırılması için uygun olmaktadır.

HaDV2 içermeyen (Şekil 3A) olan ebeveynlerden dölleri NONINF-suşu olarak yetiştirilir. Başarıyla DNA şablonları kaliteli (Şekil 3B) olduğunu öne süren, aynı DNA şablonları kullanılarak, aktin geni amplifiye edilmiştir. Buna ek olarak, rasgele seçilen sekiz dölleri de HaDV2 (Şekil 3C), HaNPV (Şekil 3D), ve Wolbachia (Şekil 3E) serbest edildi. Yine, aktin g...

Watch this Scientific Journal Video about Protocols for Investigating the Host-tissue Distribution, Transmission-mode, and Effect on the Host Fitness of a Densovirus in the Cotton Bollworm at JoVE.com

Protocols for Investigating the Host-tissue Distribution, Transmission-mode, and Effect on the Host Fitness of a Densovirus in the Cotton Bollworm (Video) | JoVE

Burada, bir lepidopteran tür içindeki bir densovirüs konakçı fitness konak doku dağılımı, iletim modunu ve etkisi, pamuk kurtları araştırmak için bir protokol mevcut. Bu protokol aynı zamanda diğer oral bulaşan virüsler ve bunların haşere arasındaki etkileşimi incelemek için kullanılabilir.

Pamuk Yeşilkurt bir densovirüs Host Fitness Host doku Dağılımı, İletim-modunu ve Etkisi incelenmesi için Protokoller

Many novel viruses have been discovered in animal hosts using next-generation sequencing technologies. Previously, we reported a mutualistic virus, ...

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