Part:BBa_K4335023

Rbcs2 Promotor+mCherry+Rbcs2 Term

BBa_K4335022 is a composite part consisting of a promoter, a fluorescent protein gene and a terminator. mCherry [BBa_K4335004] is a red fluorescent protein.[1] Fluorescent reporter genes for biological applications. We optimized its codon to express in Chlamydomonas reinhardtii more efficiently.

Usage

We introduced the mCherry gene into plasmid pTX2038 as its reporter gene. Using PCRRBCS2 as promoter and TCRRBCS2 as terminator, mCherry gene expression was realized.

Result

Plasmid construction

To validate the mCherry sequence in our vector, we designed mCherry-f and mCherry-R primers to verify our successful assembly.

mCherry and the position of primer targeting

Electrophoregram of amplification products,M is DNA Marker.

Functional Identification

We introduced the mCherry-containing plasmid pTX2038 into Chlamydomonas reinhardtii by electroporation and observed it using Fluorescence microscope.

Fluorescence excitation of positive clones transferred into pTX2038 vector compared with wild type mCherry, together with DIC field and chlorophyll excitation as fluorescence control.

Reference

[1] Codon usage and tRNA content in unicellular and multicellular organisms. T Ikemura. Mol Biol Evol (1985) 2 (1): 13-34.

[2]Parenteau, J. et al. Introns are mediators of cell response to starvation. Nature 565, 612–617 (2019).

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]