Part:BBa_K2751008

YPet skeleton

Usage and Biology

YPet is a type of YFP that is improved specifically for exhibition of Forster resonance energy transfer(FRET). It is a constitutively fluorescent YFP derived from Aequorea victoria. CyPet is developed along with YPet so that the pair of Fluorescent proteins(YPet and CyPet) can produce drastic signal changes in FRET observations, making them ideal for measurements requiring the use of FRET.

In this part, YPet is flanked with NheI, KpnI, BamHI at 5’ end and SalI and XhoI at 3’ end. Therefore, other sequences can be easily cloned onto this part and so can YPet be easily inserted to other sequences. Also, polyhistidine tag(his tag) is added to the 3’ end of YPet, enabling the YPet produced to be purified through Ni+ chromotography.[3]

In short, this is a highly modifiable and convenient part with YPF optimized for FRET reactions.

We have successfully put this part under the control of a T7 promoter and produced functional YPet with his tag. The results can be seen below:

reference: 1.PS, N. (2018). Evolutionary optimization of fluorescent proteins for intracellular FRET. - PubMed - NCBI. [online] Ncbi.nlm.nih.gov. Available at: https://www.ncbi.nlm.nih.gov/pubmed/15696158 [Accessed 13 Oct. 2018]. 2.https://www.fpbase.org/protein/ypet/ 3. Spriestersbach A, e. (2018). Purification of His-Tagged Proteins. - PubMed - NCBI. [online] Ncbi.nlm.nih.gov. Available at: https://www.ncbi.nlm.nih.gov/pubmed/26096499 [Accessed 13 Oct. 2018].

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal NheI site found at 40
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BamHI site found at 61
  Illegal XhoI site found at 790
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI.rc site found at 713