Germ-line injection of mRNAs encoding GFP

Hi everyone.

Has anybody tried injecting mRNAs encoding GFP (or other fluorescent proteins) into the hermaphrodite germ-line? We have been injecting lacZ mRNAs as translational reporters, but i need an alternative method that does not involve killing the worm.

Any help or advice would be gratefully received.

Yes we have successfully injected mRNA encoding eGFP made in vitro with a 5’Cap and 3’ poly ‘A’ tail into the gonad. GFP expression in seen in successfully injected worms after about 3-4 hours.

Thanks for the reply. I have a few more questions:

Was fluorescence visible under a dissection microscope?

In what proportion of injected animals could you detect expression?

What concentration of mRNA did you inject?

What reagents did you use for in vitro transcription and tailing?

I really appreciate your help.

Hi there,

I made my mRNA using mSCRIPT mRNA kit from Epicenter. It states a final conc of about 60ug of mRNA can be obtained from their kit. I got around that amount on some samples but others were a little lower ???

The kit comes with 5’Capping and 3’ Poly ‘A’ tailing reagents. I used both linearised plasmids and PCR products with the T7 promoter as templates and both worked well.

I injected my mRNA’s between 500-1000ng/ul. This is high but I didnt see much eGFP under a dissecting scope. We now have a much higher resolution scope but I have stuck with the above concentrations as I know it works.

The number of animals seen with eGFP relates directly with how good your injections are. On a good day I get around 60-70? of my injections to show eGFP. I also noted I got eGFP expression in tissues I mistakenly hit eg the intestinal cells :-\

The only thing of note is we had our eGFP sythesised as a DNA sequence that was codon optimised for C. elegans. I dont know if this is something you would want to consider.

Let me know if you need anything else… Good Luck!


This is great, thank you so much.

Would you consider sharing the eGFP plasmid that you use for in vitro transcription?

If so, contact me on andrewwoodawayatgmaildotcom and i will happily pay for shipping costs.