Part:BBa_K2865015

Left ITR-BNP-eGFP-Poly(A)-Right ITR

This part contains left and right inverted terminal repeats(ITRs) (BBa_K2865002,BBa_K2865003) , BNP promoter（BBa_K2865000）, EGFP reporter, and SV40 polyA (BBa_K2865004).

Usage and Biology

This part is designed to detect AAV9 packaging efficiency if the BNP promoter can be active in heart failure. We all know that the AAV9 genome is built of single strand DNA. ITRs are indispensable in AAV9 genome, regarding as the only cis-acting element in AAV replication and encapsidation. And the ITRs are also needed to get an efficient encapsidation of AAV9 DNA combined with generation of a fully assembled, deoxyribonuclease-resistant AAV particles. And this part is compared to the control group (BBa_K2865014). And we can learn that the Left ITR-CMV-EGFP-Poly(A)-Right ITR part can expressed EGFP in cell and animal tissue which means the constructs of the two parts are all right.

Characterization

We constructed an AAV9 vector containing the part of Left ITR-BNP-EGFP-Poly(A)-Right ITR(BBa_K2865015). And we have done cell experiments and animal experiments to test whether the BNP promoter can be active and make the EGFP gene express. However, beyond our expectations and data research, we found no green fluorescent signal in both two experiments. And the result is shown in the picture.

 

Fig.1 The results of Left ITR-BNP-eGFP-Poly(A)-Right ITR was without green fluorescent signal expression

 

Sequence and Features

 

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BglII site found at 1745
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal NgoMIV site found at 1467
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI site found at 425