Part:BBa_K1884011

RBCS2 Promoter+Paromomycin+RBCS2 Terminator

We provided Paromomycin resistance under the RBCS2 promoter(BBa_K1884010)

Biology

RBCS2 is a promoter of the small subunit of ribulose-1, 5-diphosphate carboxylase, encoded by the nuclear gene and is an inducible promoter for strong light-induced expression.

Paromoycin is a protein synthesis inhibitor in nonresistant cells by binding to 16S ribosomal RNA. This broad-spectrum antibiotic soluble in water, is very similar in action to neomycin. Antimicrobial activity of paromomycin against Escherichia coli and Staphylococcus aureus has been shown.

RBCS2 terminator is Chlamydomonas reinhardtii RuBisCO small subunit 2 terminator which obtained from the pOpt_mVenus_Hyg expression plasmid provided by Chlamydomonas Resource Center.(Fig 1)

Figure 1. The diagram of this composite part in pSB1C3 Backbone.

Usage

For testing the efficiency of paromoycin, we formulated six gradient concentrations of paromoycin solution and add 200ul each into six different beakerflask which contain 200ml LB solid medium, then we put the competent cell into six different LB plate. there is a photograph to show several monoclonal colony growing on a LB plate which has the best selected abilty of Paromoycin.(Fig 2)

Conbined with another resistant device(BBa_K1884012), we are able to screen the green algae which has paromoycin resistance and Hygromycin B resistance. we added 200ul paromoycin solution and hygromycin B solution into 200ml TAP solid medium, then we put the green algae without cytoderm into TAP plate. there is a photograph to show several colony growing on a TAP plate which has the best selected efficiency of paromoycin and Hygromycin B.(Fig 3)

Figure 3. The conlony of green algae selected by paromoycin and Hygromycin B.

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]