Part:BBa_K3278006

pDawn with YF2

pDawn is a blue light control system which can start the expression of target gene under the inducement of blue light. More information in BBa_K1075044 added by iGEM13_Bonn.

However, expression leakage and delay are the two main problems of pdawn system. Leakage refers to the expression of target protein without light. According to our measurement, samples strictly kept away from light still had significant expression of GFP after being cultured for one night. Therefore, we carefully studied the gene circuits of pdawn system [2] [3] and hypothesized that enhancing the strength of the promoter of YF1 may enhance the repression of protein expression in the dark, thus solving the leakage problem.

In this part, we tried to change YF1 into YF2. YF2 is reported in Andreas Möglich’s paper[2]. By changing 24 bps in the original sequence, YF2 was reported to have higher promotion effect on FixJ expression, which means stronger inhibitory effect on target gene’s expression.

pDawn with T7 promoter is abbreviated as T7c below. pDawn with tac promoter is abbreviated as tac below. pDawn with modified tac promoter is abbreviated as mtac below. pDawn with YF2 is abbreviated as YF2 below.

[Go for the protocol of characterization:https://2019.igem.org/Team:ShanghaiTech_China/LightControl]

Fig1. pDawn-YF2-0 hour

Fig2. pDawn-YF2-6 hour

Fig3. pDawn-YF2-12 hour

Fig4. Dark to light-12 hour

Fig5. pDawn-YF2-24 hour

Fig6. Dark to light-24 hour

Fig7. pDawn-mEGFP-30 hour

Fig8. Dark to light-30 hour

Fig9. pDawn-YF2-36 hour

Fig10. Dark to light-36 hour

To see more changes on pDawn, go to the websites below.

pDawn-mEGFP: <a href="https://parts.igem.org/Part:BBa_K3278000"> BBa_3278000</a>

pDawn with T7 promoter: <a href="https://parts.igem.org/Part:BBa_K3278001"> BBa_3278001</a>

pDawn with mtac promoter: <a href="https://parts.igem.org/Part:BBa_K3278002"> BBa_3278002</a> (good)

pdawn with tac promoter: <a href="https://parts.igem.org/Part:BBa_K3278005"> BBa_3278005</a>

pDawn with YF2: <a href="https://parts.igem.org/Part:BBa_K3278006"> BBa_3278006</a> (good)

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 2148
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal NgoMIV site found at 46
  Illegal NgoMIV site found at 178
  Illegal NgoMIV site found at 272
  Illegal NgoMIV site found at 565
  Illegal NgoMIV site found at 1059
  Illegal NgoMIV site found at 1077
  Illegal NgoMIV site found at 1167
  Illegal AgeI site found at 397
  Illegal AgeI site found at 1525
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI site found at 1620
  Illegal BsaI.rc site found at 508