实验动物模型

Injection of C. elegansPrepare injection padsline up three 22x50 mm coverslips place drop of molten 2% agarose on each quickly place slide on top of the agarose and wait 20 seconds (see figure) carefu ...

Worm Injectionspads:melt 2% agarose in water keep molten at 42oCuse 25mm x 25mm or larger microscope slidesspread out microscope slides onto a heat resistant glass or metal trayusing a pasteur pipet p ...

oocyte isolation Anethesize female using benzocaine - put ~5ml 6% benzocaine (stock solution in 100% ethanol) into 1L water.- leave animal until it is limp and completely unresponsive.- place animal o ...

Chi's method Pick your embryos (~5 per tube) and place in an autoclaved 1.5 ml capped centrifuge tube Wash embryos 1 ml of autoclaved DEPCed-PBS (containing 1x PBS; pH 7.4) -- take care not to lyze or ...

Oocyte transfer: IntroductionProtocol submitted by: This chapter describes a method that has been used successfully to study the roles of a number of maternal mRNAs in Xenopus embryos (Kofron et al. 1 ...

1-2 g fresh material freezer-dried ground with 0.2g sand (if necessary) and then homogenized with 10ml RNA extraction buffer (see below). Spin at 8000rpm 4oC for 10 minutes Remove the supernatants to ...

Overview Using this procedure it is possible to follow the development of pollen mother cells through to mature pollen. Sterile males were made using irradiation mutagenesis of Landsberg erecta seed ...

Competent agro prep for electroporationday 1 1. Start 75 mL overnight cultures of agro (strain GV3101 C58C1 Rifr pMP90 Gmr Koncz & Schell) in YEP in 250 mL baffle flasks. 2. Grow at 28 °C shaking. 3. ...

Culturing Wormsby Michael Koelle1. Bacterial strain for feeding worms: OP50 a uracil auxotroph. Streak OP50 out on a 9 cm NGM agar plate grow overnight at 37°. Can then parafilm the plate and keep it ...

Development of techniques for primary culture of C. elegans embryonic neurons Laird Bloom MIT from Ph.D. thesis Massachusetts Institute of Technology 1993 Introduction One of the major limitations of ...

F2 Mutant Screen - non "clonal"by Michael Koelle adapted from Erik Jorgenson4/6/941. Mutagenize mixed population worms as per normal. Best to do this first thing in the morning in order to get several ...

Fermentor worm chowPrep Have autoclaved: 250 ml straight neck flask 100 mmfunnel Fermentor vessel with all openings covered top loosened on its side in tub with wedges after a/c sit upright to seal a ...

FREEZING WORM STOCKS1. Grow 3-5 6cm. plates of worms on NGM with OP50 until the plate is starved with lots of young larvae. The L1s are the ones that survive freezing the best and are the most likely ...

Freezing Worms by Michael Koelle 4/6/94 I. The preferred method 1. Wash worms off of 1 large plate or 3 small plates in 3 mls S Basal into a sterile 15 ml disposable centrifuge tube. Worms should be h ...

Growing worms in liquidFirst make worm food -- grow liters of bacteria Inoculate each of several liters (4 is a good number) of LB in 2 liter flasks with 1 ml of an overnight culture of AMA1004. Grow ...

Lineaging of C. elegans Embryosby Pressure-free MountingMaterials:Glass Slides 18mm x 18mm Coverslips Petroleum Jelly #15 Disposable Scalpel Embryonic Blastomere Medium ...

Integrating extrachromasomal arrays into the C. elegans chromosomes Why and how to do it by Michael Koelle 4/6/94 What is the benefit of integrating an extrachromosomal array? Extrachromosomal arrays ...

Lethal phase determination1. Pick a bunch of heterozygote L4 hermaphrodites to a new plate.2. The next day there will be eggs on the plate; pick exactly 20 of these to each of a few plates. Make sure ...

Integrating extrachromasomal arrays into the C. elegans chromosomesWhy and how to do itby Michael Koelle12/20/94What is the benefit of integrating an extrachromosomal array? Extrachromosomal arrays su ...

Microinjecting worms by Michael Koelle8/23/94You will have a couple frustrating sessions when you first attempt this technique but everyone seems to master injection after a few days and it works very ...