Part:BBa_K1415105

Pcons+B0034+PBAN(Spodoptera litura)

Fig.3-1 P_cons+RBS+PBAN(Spodoptera litura)

To verify the PBAN of Spodoptera litura can be expressed by the E.coli, we conducted a SDS protein electrophoresis experiment. We first smashed the E.coli containing the PBAN with a sonicator and then took the supernatant divided from the bacterial pellet by centrifugation. Finally, we used the supernatant to run a SDS protein electrophoresis in a 20 % SDS gel.

Fig.3-2 Protein Electrophoresis of P_cons + RBS + 5 different kinds of PBAN (control: plasmid of P_cons+RBS) Each peptide of PBAN is an around 30 amino acids, so we can see the band of PBANs at 2~4 kDa.

Below are biobrick serial numbers of PBAN abbrevation:

BM: BBa_K1415001　　　AA: BBa_K1415009　　　LD: BBa_K1415104

AS: BBa_K1415007　　　SL: BBa_K1415005

Behavior of Target Insects After PBAN Treatment

To investigate what behavior the female moth would show after ingesting PBAN, we put one female moth into a beaker for observation. The beaker is divided into two parts by plastic wrap. The bottom part contains the PBAN solution we prepared, and the upper part is the space for the moth to stay. We soaked cotton that spans the entire length of the beaker with the PBAN solution and sprinkle it with sugar. This way, the moth can suck on the PBAN without drowning in PBAN solution. After all the equipment is set, we put the female moth into the upper part of the beaker. At the time, we started filming as soon as we observed the female moth showing obvious behaviors of sexual stimulation such as flapping their wings. In this observation, the sample moth is Spodoptera litura which we caught in Sunny Morning organic farm.

We observed that the moth could absorb the PBAN in the solution through ingestion, and that the PBAN could stimulate the moth's pheromone gland to produce pheromone. As soon as the moth is sexually excited, it would flap its wings rapidly and move its tail slightly upward .

This video shows the behaviors of female moth after ingesting its separate PBANs. The moths clearly became excited and flapped their wings rapidly.

Fig.3-3 Negative Control: Female moth without eating PBAN (Number = 0). Experiment: Female moth eating our PBAN (Number = 11). In this picture, we can see the PBAN effect that the female moth eating PBAN solution can release much sex pheromone, and attract many male moths.

　

Fig.3-4 Negative Control：sucrose solution, Experiment：Female moth eating PBAN solution. Also, we can see the PBAN effect again from this picture.

Fig.5-2 Shows the entering times of moths(Spodoptera litura) in different conditions. Entering times is defined as the times that moths get into the device. In this experiment, the moth might get out of our device due to it was just a model without trap, such as nets for keeping the moths. However, we can still clearly see the magic power of our pyramidal device by using blue light with PBAN treatment, which attracted more moths to get into the device than the other experiment (only blue light treatment). In addition, we can even observe a unique periodicity of attracting moths only in blue light with PBAN treatment. We consider this phenomenon is related to the physiology of the female moths' rut situation.

Fig.5-2 The entering number (into our pyramidal device) per hour shows that the combination of blue light, PBAN, and our device is indeed magically powerful in insect attraction.

Sequence and Features

Assembly Compatibility:

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

  Illegal NheI site found at 7
  Illegal NheI site found at 30
  
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal NgoMIV site found at 79
  
• 1000
  COMPATIBLE WITH RFC[1000]