Part:BBa_K4770033

GS_Cp_GS_Level_M

Description and Biology

Level Ms are the assembly of different transcriptional units into the same vector.

In AlgaGenix we have designed what we call “gene-pairs Level Ms”. All of them follow the same structure: they contain the HygroR cassette, level 1 for two different genes (pCM1:NR + pCM1 NiR, pCM1 GS+ pCM1 Cp-GS or pCM1 IPT + pCM1 LOG), mVenus cassette, and mCherry cassette.

In this case, we are using the combination pCM1: GS + pCM1 Cp-GS

With this vector, we aim to see the effects of overexpressing both forms (cytosolic and in the chloroplast) of Glutamate Synthetase, the enzyme responsible for the last step on the assimilation pathway simultaneously. Additionally, we are also able to normalize the expression level depending on the insertion site of the vector (by measuring the signal of the mCherry cassette), at the same time, we will be able to see the decrease of nitrates concentration in the media due to an enhancement of the nitrates assimilation pathway thanks to the mVenus cassette signal.

How can this part be used

This Level M is aimed to assembled with HygroR cassette in position 3, pCM1 GS in position 4, pCM1Cp-GS in position 5, mCherry cassette in position 6 and mVenus cassette in position 7.

Should you use this part and need advice on how to use it, please do not hesitate to contact us at algagenix@gmail.com

Source of this part

Position 3: Hygro R cassette BBa_K4770016

Position 4: pCM1 GS: BBa_K4770020

Position 5: pCM1 Cp-GS BBa_K4770021

Position 6: mCherry cassette BBa_K4770017

Position 7: mVenus cassette BBa_K4770025

Design considerations

AlgaGenix’s Level Ms are designed to have the same structure and not having to build different pieces with different positions for each transcriptional unit. Each piece has it’s own design considerations that can be looked up on the individual part’s pages.

Sequence and Features

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Illegal EcoRI site found at 4725
  Illegal EcoRI site found at 8383
  Illegal PstI site found at 10029
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal EcoRI site found at 4725
  Illegal EcoRI site found at 8383
  Illegal PstI site found at 10029
  Illegal NotI site found at 9765
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal EcoRI site found at 4725
  Illegal EcoRI site found at 8383
  Illegal BglII site found at 5063
  Illegal BglII site found at 8721
  Illegal BamHI site found at 4369
  Illegal BamHI site found at 8027
  Illegal XhoI site found at 4581
  Illegal XhoI site found at 8239
  
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal EcoRI site found at 4725
  Illegal EcoRI site found at 8383
  Illegal PstI site found at 10029
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal EcoRI site found at 4725
  Illegal EcoRI site found at 8383
  Illegal PstI site found at 10029
  Illegal NgoMIV site found at 1265
  Illegal NgoMIV site found at 1447
  Illegal NgoMIV site found at 3927
  Illegal NgoMIV site found at 4879
  Illegal NgoMIV site found at 7585
  Illegal NgoMIV site found at 8537
  Illegal NgoMIV site found at 10506
  
• 1000
  COMPATIBLE WITH RFC[1000]