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方法文章

Drosophila 卵收集与去壳:一种去除卵最外层的方法

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2023年4月30日

本文内容

摘要

来源:Koyle, M. L. 等。无菌和已知菌群条件下培养果蝇Drosophila melanogaster。J. Vis. Exp. (2016)。

Drosophila 胚胎被保护性的外膜所包围。为了接触胚胎,通常需要去除这些膜中最外层的卵壳膜(chorion)。本视频介绍了一种常用的使用次氯酸钠进行脱卵壳(dechorionation)的方法。示例实验方案展示了该操作流程,并采用了无菌操作技术,以满足无菌或已知菌群动物培养的需要。

方案

本实验方案 摘自  Koyle 等人饲养果蝇 黑腹果蝇 在无菌和已知菌群条件下 J. Vis. Exp. (2016).

1. 准备产卵笼

  1. 通过微波加热100 ml水、10 g酿酒酵母、10 g葡萄糖和1 g琼脂来制备葡萄汁琼脂平板。在步骤1.1中煮沸3次,并加入10 g冷冻葡萄汁浓缩液,以提高琼脂平板上卵的可见度。
  2. 当琼脂冷却至55 °C时,将20 ml倒入100 mm培养皿中,静置使其凝固。
  3. 通过将1 g酿酒酵母与15 g水混合制成酵母糊,覆盖琼脂平板表面。将酵母糊倒在琼脂平板上,确保表面完全覆盖,然后倒去多余部分,留下一层薄薄的酵母残留物。若使用多个平板,可依次将酵母糊倒在多个平板上。
  4. 将琼脂平板转移至笼子底部。
    注意:32盎司的熟食容器是丙烯酸果蝇笼的良好替代品。
    1. 通过在容器顶部切一个孔,并用无毒胶水将透气网膜粘在孔上,为32盎司熟食容器制作盖子。如果胶水可溶于水,则应避免盖子接触水分。
  5. 将200–300只果蝇转移至容器中,并盖上盖子。用空的培养皿盖覆盖有网膜保护的孔,以防止琼脂表面水分蒸发;如需要,可在盖子内部放置一张....

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材料

本文使用的材料清单
姓名公司目录编号评论
酿酒酵母MP Biomedicals, LLC.903312http://www.mpbio.com/product.php?pid=02903312
葡萄糖Sigma Aldrich158968-3KGhttp://www.sigmaaldrich.com/catalog/product/aldrich/158968?lang=en®ion=US
琼脂Fisher--Lab Scientificfly802010https://www.fishersci.com/shop/products/drosophila-agar-8-100mesh-10kg/nc9349177
Welch's 100% 葡萄汁浓缩液沃尔玛或其他杂货店9116196http://www.walmart.com/ip/Welch-s-Frozen-100-Grape-Juice-Concentrate-11.5-oz/10804406
饲养笼:32 盎司半透明圆形熟食容器Webstaurant Store999L5032Yhttp://www.webstaurantstore.com/newspring-delitainer-sd5032y-32-oz-translucent-round-deli-container-24-pack/999L5032Y.html
半透明圆形熟食容器盖Webstaurant Store999YNL500http://www.webstaurantstore.com/newspring-delitainer-ynl500-translucent-round-deli-container-lid-60-pack/999YNL500.html
培养瓶Genesee Scientific32-130https://geneseesci.com/shop-online/product-details/?product=32-130
Droso-PlugsGenesee Scientific49-101https://geneseesci.com/shop-online/product-details/?product=49-101
尼龙网Genesee Scientific57-102https://geneseesci.com/shop-online/product-details/715/?product=57-102
塑料衬套Home Depot100404002http://www.homedepot.com/p/Carlon-1-in-Non-Metallic-Terminal-Adapter-E943FR-CTN/100404002
塑料衬套盖Home Depot100153897http://www.homedepot.com/p/1-1-4- in-Rigid-Plastic-Insulated-Bushing-2-Pack-27529/100153897
样本杯MedSupply PartnersK01-207067http://www.medsupplypartners.com/covidien-specimen-containers.html
Repeater M4Eppendorf4982000322https://online-shop.eppendorf.us/US-en/Manual-Liquid-Handling-44563/Dispensers--Burettes-44566/Repeater-M4-PF-44619.html
50 ml 离心管TrueLine Centrifuge TubesTR2003https://www.lpsinc.com/Catalog4.asp?catalog_nu=TR2003
食物盒USA Scientific2316-5001http://www.usascientific.com/search.aspx?find=2316-5001
Lysing Matrix D BulkMP Biomedicals, LLC.116540434http://www.mpbio.com/search.php?q=6540-434&s=Search
滤芯吸头,300 μlUSA Scientific1120-9810http://www.usascientific.com/search.aspx?find=1120-9810
培养皿Laboratory Product SalesM089303https://www.lpsinc.com/Catalog4.asp?catalog_nu=M089303
乙醇Decon Laboratories, INC.2701http://www.deconlabs.com/products.php?ID=88
画笔沃尔玛5133http://www.walmart.com/ip/Chenille-Kraft-5133-Acrylic-Handled-Brush-Set-Assorted-Sizes-colors-8-Brushes-set/41446005
镊子Fisher08-882https://www.fishersci.com/shop/products/fisherbrand-medium-pointed-forceps-3/p-128693
家用漂白剂(6-8% 次氯酸盐)沃尔玛550646751http://www.walmart.com/ip/Clorox-Concentrated-Regular-Bleach-121-fl-oz/21618295
通用蛋白胨Genesee Scientific20-260https://geneseesci.com/shop-online/product-details/?product=20-260
酵母提取物 Fisher ScientificBP1422-500https://www.fishersci.com/shop/products/fisher-bioreagents-microbiology-media-additives-yeast-extract-3/bp1422500?matchedCatNo=BP1422500
磷酸氢二钾Sigma AldrichP3786-1KGhttp://www.sigmaaldrich.com/catalog/
柠檬酸铵Sigma Aldrich25102-500ghttp://www.sigmaaldrich.com/catalog/
乙酸钠VWR97061-994https://us.vwr.com/store/catalog/product.jsp?catalog_number=97061-994
硫酸镁Fisher ScientificM63-500https://www.fishersci.com/shop/products/magnesium-sulfate-heptahydrate-crystalline-certified-acs-fisher-chemical-3/m63500?matchedCatNo=M63500
硫酸锰Sigma Aldrich10034-96-5http://www.sigmaaldrich.com/catalog/
MRS 粉末Sigma Aldrich69966-500Ghttp://www.sigmaaldrich.com/catalog/product/sial/69966?lang=en®ion=US
96 孔板读数仪BioTek (Epoch)NAhttp://www.biotek.com/products/microplate_detection/
1.7 ml 离心管USA Scientific1615-5500http://www.usascientific.com/search.aspx?find=1615-5500
滤芯吸头,1,000 μlUSA Scientific1122-1830http://www.usascientific.com/search.aspx?find=1122-1830
96 孔板Greiner Bio-One655101https://shop.gbo.com/en/usa/articles/
陶瓷珠MP Biomedicals, LLC.6540-434http://www.mpbio.com/product.php?pid=116540434
组织匀浆器MP Biomedicals, LLC.116004500http://www.mpbio.com/product.php?pid=116004500
一级生物安全柜Thermo ScientificModel 1395http://www.thermoscientific.com/en/product/1300-series-class-ii-type-a2-biological-safety-cabinet-packages.html

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