Effective expression of the green fluorescent fusion proteins in cultured Aplysia neurons.
The green fluorescent fusion protein and its isoforms are extensively used to monitor gene expression, protein localisation and their dynamics in relations to fundamental cellular processes. However, it has not yet been effectively applied to Aplysia neurons that serve as a powerful model to study the mechanisms underlying neuroplasticity. We report here the development of a procedure combining in vitro transcription of mRNA encoding fluorescent-tagged proteins and its subsequent injection into the cytoplasm to image, in real-time, protein dynamics in cultured Aplysia neurones. To illustrate the efficiency of the procedure we report here the visualisation of actin, microtubules and vesicle trafficking. The results presented here introduce a reliable and effective method to express green fluorescent protein (GFP) fusion proteins in cultured Aplysia neurons.
Sahly
I
I
Department of Neurobiology, Institute of Life Sciences, The Hebrew University of Jerusalem, Jerusalem 91904, Israel.
Erez
H
H
Khoutorsky
A
A
Shapira
E
E
Spira
Micha E
ME
D000818:Animals; D001048:Aplysia; D002478:Cells, Cultured; D015870:Gene Expression; D017930:Genes, Reporter / Q000235:genetics*; D049452:Green Fluorescent Proteins; D066298:In Vitro Techniques; D007202:Indicators and Reagents / Q000378:metabolism; D008164:Luminescent Proteins / Q000235:genetics; D008565:Membrane Proteins / Q000235:genetics; D008845:Microinjections; D008870:Microtubules / Q000502:physiology; D008967:Molecular Biology / Q000379:methods*; D009419:Nerve Tissue Proteins / Q000235:genetics; D009474:Neurons / Q000166:cytology / Q000502:physiology*; D010957:Plasmids / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics / Q000494:pharmacology; D011993:Recombinant Fusion Proteins / Q000235:genetics*; D013572:Synaptic Vesicles / Q000502:physiology; D050825:Synaptosomal-Associated Protein 25; D014158:Transcription, Genetic
D007202:Indicators and Reagents; D008164:Luminescent Proteins; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D050825:Synaptosomal-Associated Protein 25; D049452:Green Fluorescent Proteins