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Team:HokkaidoU Japan

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• Top
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  • MULTIMERIZATION
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  • FUTURE WORK
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We conducted SDS-PAGE with (BBa_K2015012+BBa_K2015008/ BBa_K2015012+BBa_K2015009) on （pSB1C3）of E.coli（DH5α）. At first, we made the Gel for SDS-PAGE, with the following the Table1. What was the most important was to add TEMED ffinally because

Table. 1. Gel assay

Materials Volume 30％ Acrilmid 5 mL Tris-Buffer 2.5 mL SDS 100 µL APS 100 µL TEMED 5 µL DW 2.295 ｍL Total 10 ｍL

The following Table. 2 is about preparing the SDS-PAGE.

Table. 2. Experimental condition

Temperature IPTG Concentration (M) Volume (µL) Time to culture 37°C - - - 24 h 37°C + 0.4 6 24 h 37°C + 2 30 24 h 25°C - - - 16 h 25°C + 0.4 6 16 h 25°C + 2 30 16 h After we finished cultivating samples, we took 100 µL out of each samples and made the following operation. Centrifuge with 13000rpm at 24°C for 2min Remove the supernatant Add 50 mL of SDS-Buffer Shake for 1 min Keep at 100°C for 5 min Put on the ice Apply 10 µL to SDS Run electrophoresis for 1.5 h Wash out with MiliQ Shake at 24°C with 34 rpm for 1 h Dye with QuickCBB The fig. 1 shows the result of SDS-PAGE.