Difference between revisions of "Team:SZU China/LZU-China"

LZU_China

The in-depth Communication of LZU&SZU iGEM

Due to Lanzhou University(LZU) has a very good experience in cancer research, and LZU iGEM debuted in Asia Division in 2013, with the《Twinkle Medicine Hunter of Cancer 》 obtained a good achievement,and because the project of SZU iGEM is on cancer research this year,we reached cooperation agreement. Part of the experimental verification results of our project was LZU iGEM helping us design and verify.
The entries of LZU iGEM This year is about environmental protection,and just last year,Exactly right the project we did last year related environmental protection,called 《Alkaline Cellulase Dream Factory》.
Last year is SZU first time to participate in iGEM,and we won the gold medal.Therefore,LZU iGEM is very interested in and confident of us,and we communicated online.For their projects,we provided constructive suggestions on the model and helped them to design a part of the model.
Therefore,in order to further in-depth exchanges with us and thank-us-for-help,LZU iGEM traveled long distances from Lanzhou to Shenzhen on August 19, 2015.In the course of communication,we have reached a consensus,and hope that our two school teams can work together for a long time to achieve win-win.

We help LZU_China to build some part of the model:

During the period of LZU-China conduct this project, a tough problem occurred that the mu-circuit needs both a huge voltage difference and a stable voltage so that it needs to eliminate interference from supply voltage. Only with a steady reliable device can they work out a stable millivolt level voltage. Therefore, this question confused them for a long time but eventually accomplished their voltage stabilizing circuit with the help of us. We provided this voltage stabilizing circuit.

TPS5430 characteristic
TPS5430 owns good characteristics. Its capability can be seen as follows.
High current output: 3A (peak value 4A)
Wide voltage input range: 5.5V~36V
High conversion efficiency: in best condition can be 95%
Wide voltage input range: the lowest can be 1.221 V
Internal compensation minimizes the number of external devices
Regular 500kHz slew rate
Have over current protection and overheated protecting switch
Have ENA and when it is in the off mode, its current is only 17uA.
Internal Slow Start
  Compared with some other similar type DC switching power supply converting chip, TPS5430 has a better performance in high conversion efficiency.
Fig 1: In the condition of 12V input voltage and 5V output voltage, the relational graph between conversion efficiency and output current can be seen as follow.

(2) Inner structure and function
① Oscillate or frequency
Regular 500kHz slew rate makes it produce a more accurate in the same requirement of output ripple output inductance.
② Reference voltage
Through scaling temperature to stabilize the energy band gap, the reference voltage system can produce an accurate reference signal. After testing, in the proper temperature, when the 1.221V voltage output, it can keep the balance of energy band gap and mu-circuit.
③ ENA and Slow Start
When ENA voltage exceeds the peak value, the converter and the slow start inside will begin working. And when it is lower than the peak value, the converter and the slow start inside will stop working. When the ENA foot landing or the voltage is lower than 0.5V, the converter will stop working. ENA foot can hang in the air.
④UVLO
TPS5430 with UVLO circuit, no matter it is powering on or powering off, if the VIN lower than the peak voltage, the converting chip will not work. The typical hysteresis of UVLO comparator is 330mV.
⑤Boost Capacit or
Between the foot of BOOT and PH, link a 0.01μF ceramic capacitor to provide a threshold voltage for the high side.
⑥VSENSE and Internal Compensati on
The output voltage can be feedback to the VSENSE foot through external resistor partial pressure. In the steady state, the voltage on the VSENSE foot equals 1.221V. TPS5430 has compensating circuit inside which helps to simplify the chip design.
⑦Voltage Feed-Forward
Voltage Feed-Forward inside ensures that there is always a constant gain of power chip, no matter how input voltage changes. So voltage feed-forward can simplify the analysis of stability and improve the instant response. The typical value of TPS5430 voltage feed-forward is 25.
⑧ Pulse-Width-Modulation Control
Converter adopts fixed-frequency to modulate.
⑨Overcurrent Protection
The overcurrent protection circuit is provided to trigger the overcurrent protection on the condition that the inner overcurrent indicator is set to true when current overloads.
⑩ Thermal Shutdown
When the junction temperature is higher than the shutdown temperature, voltage parameter will be set to ground and “high-side” MOSFET will be shut down. Controlled by soft-starting circuit, chip will restart when the temperature is 14℃ lower than the shutdown point. They are able to gain the stable voltage within the gap of 5V and input the voltage into the amplifying circuit of themselves through this circuit. So this circuit secures the stability of the amplifying circuit and makes exploitation easier for them.

Verifying the accessibility and the portability of the device

Besides the voltage stabilizing module we optimized for them, LZU-China also sent us the major components of their “Micro Holmes” device (including the pcDuino board, the voltage magnification circuit board, the analogue-to-digital Conversion chip, and the touch screen). So we can help them verify the accessibility and the portability of their device. According to their specification, we successfully connected these electronic components, start the device and tested the software pre-installed in their pcDuino board. Although most functions of their device can work normally, there were also some small bugs. For example, in the “initialization” UI, they put the “concentration” title in front of the “Output Voltage”. We reported these bugs to them, and they correct them.

LZU_China helps us:

Based on the above experiments, the system was improved.The improving content is: put effector gene(Renilla Luciferase) in a third plasmid,which has strong promoter SV40.Firefly luciferase was replaced with Renilla Luciferase.The expectation to do this is that:In the same time to increase the security and the expression of the system,and increasing the influence of the orthogonal system to the project system. Through many times transfection experiment of using three plasmids system to transfect T24 cells, the result shows, the safety of the system was verified is good.Compared the expression of the two plasmid system in Hela cervical cancer cells,the expression of three plasmid system in the controll group without Ack has the same level, and the expression of three plasmid system is based on the system expectation to make that improvement,and this further shows that our improvement has no effect on the security of the system. The expression of the two groups of without Ack and adding ACK has reached more than 7 times. This shows that the effect of orthogonal system on the whole system has been improved. In addition, the average expression of adding Ack experimental group has reached 790.5, and in others successful transfection experiments, the highest number of test results have reached 1000.All the data indicated that the expectation of improving the system expression amount has been reached a certain level.

Rlu:Relative light unit
No ACK in T24:Representing plasmids were transfected into T24,without adding ACK
Add ACK in T24:Representing plasmids were transfected into T24,with adding ACK

The verification of two plasmids system and three plasmids system in different cells: Replacing Rlu with GFP as an effector gene,transfected the plasmids into different cells,and the results were verified by using the computer connected microscope. After replacing Rlu with GFP,we transfected T24 bladder cancer cells using two plasmids system and three plasmids system respectively.The positive controll group is transfecting T24 using a plasmid which carried normal GFP behind CMV promoter.The result we got using Leica fluorescence microscope photographs as follows:

Fig.2 The result in T24 of positive controll group

Fig.3 Experimental group without ACK(two plasmids system)

Fig.4 Experimental group adding ACK(two plasmids system)

Fig.5 Experimental group without ACK(three plasmids system)

Fig.6 Experimental group adding ACK(three plasmids system)