Eppendorf 012: Microinjection of plasmid DNA or double stran Manual do Utilizador descarregar pdf

Microinjection of plasmid DNA or double

stranded RNA into the gonads of

C. elegans

FemtoJet express | No 012/06

Userguide

Author: Ryuji Minasaki, Max Planck Institute for Developmental Biology,

Abteilung IV (Evolutionsbiologie), Tübingen, Germany

FIG. 1: Intragonadal injection in C. elegans

Introduction

C. elegans was established as a model organism three decades ago by Sydney Brenner (Brenner 1974).

Two major applications for microinjection in C. elegans are the production of transgenic worms (Mello et al.

1991) and the knockdown of genes by RNA interference (Fire et al. 1998).

Here, we describe a microinjection technique that has been established in our laboratory for these purposes.

Applications for microinjection in C. elegans

•

Injection of DNA to generate transgenic lines

•

Injection of dsRNA to induce RNAi

•

Injection of proteins

Research interests

Transgenic animals can be generated for a wide range of purposes. Some of the more popular uses are outlined below:

•

Direct transformation of mutant strains using cloned DNA by germline microinjection

•

Expression pattern analysis (e.g., GFP fusion constructs)

•

Structure/function analysis

•

Promoter Bashing

•

Identification of gain-of-function phenotype through overexpression / ectopic expression of an endogenous

or foreign gene

Table 1: Applications for microinjection in worm

1 2 3

Userguide 1
References 3

Resumo do Conteúdo

Página 1 - Userguide

Microinjection of plasmid DNA or double stranded RNA into the gonads of C. elegansFemtoJet express | No 012/06UserguideAuthor: Ryuji Minasaki, Max Pla

Página 2

Equipment•Inverted microscope (Axiovert®135, Carl Zeiss AG, Germany)•Micromanipulator TransferMan NK2 (Eppendorf AG, Germany)•Microinjector FemtoJet e

Página 3 - References

DescriptionProgrammable microinjector with externalpressure supplyProportional micromanipulator for microinjectionDynamic micromanipulator for microin