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From 2010.igem.org

August 2 (Mon)

Attendance: Torigata, Takino, Teoh, Tadashi, Yasumoto, Saka

1. Culture medium preparation
   • LB agar plates (49 antibiotic-less plates)
   • LB liquid medium (500 ml)
2. Competent cells preparation - Nojima Method
   • SOB medium (MgCl2 not yet added) -> stored at 4˚C
   • TB buffer -> stored at 4˚C
   • Single-colony streaking of E. coli (strain DH5α) on 2 LB agar plates -> 37˚C incubation o/n

Tomorrow we shall prepare the remainder of the reagents and inoculate SOB with o/n-incubated E. coli.

August 3 (Tue)

Attendance: Nakamura, Kakuda, Saka, Yasumoto, Teoh

1. Competent cells preparation (continued)
   • Preparation of glucose solution for making SOC medium.
   • Inoculation of pre-culture SOB from o/n-incubated agar plate colonies.
   • (Night) Transfer from pre-culture to growth culture.

August 4 (Wed)

Attendance: Nakamura, Saka, Kakuda, Yasumoto, Torigata, Teoh

1. OD measurements throughout the day till required OD (0.3~0.7) was obtained.
2. Completion of competent cells according to protocol.

August 5 (Thu)

Attendance: Nakamura, Yasumoto, Saka, Kakuda, Takino

1. Transformation of Registry parts (See Table 1).

Note: 'ID' is an internal identifier used by wet work members to simplify labeling etc. In the case of iGEM distribution parts it usually refers to the plate location.

Table 1

ID	Part Name	Resistance	Description
2-20J	<bbpart>BBa_K118023</bbpart>	A	C. fermi endocellulase Cen A coding
2-20H	<bbpart>BBa_K118022</bbpart>	A	C. fermi exocellulase Cex coding
1-2M	<bbpart>BBa_B0034</bbpart>	A	RBS
1-13D	<bbpart>BBa_B0010</bbpart>	A	terminator
1-1D	<bbpart>BBa_R0010</bbpart>	A	promoter
1-18F	<bbpart>BBa_E1010</bbpart>	K	RFP coding

August 6 (Fri)

Attendance: Nakamura, Saka, Yasumoto, Takino, Teoh

1. Colony check
   • All transformed cells produced colonies!
   • Non-transformed cells (negative controls) did not grow on Amp, Kan or Cam plates -> confirmed lack of natural antibiotic resistance
2. Colonies transferred to 3ml LB growth medium & incubated o/n at 37˚C

August 7 (Sat)

Attendance: Nakamura, Saka, Yasumoto, Takino

1. Miniprep of o/n-incubated colonies (2-20J, 2-20H, 1-2M, 1-13D, 1-1D, 1-18F) using Sigma-Aldrich Plasmid Miniprep Kit
2. Transformation of construction plasmids (See Table 2)
3. Meeting

Table 2

ID	Part Name	Resistance	Description
1-1C	<bbpart>pSB1A3</bbpart>	A	construction plasmid containing mRFP coding device (<bbpart>BBa_J04450</bbpart>)
1-3A	<bbpart>pSB1C3</bbpart>	C	(" ")
1-5A	<bbpart>pSB1K3</bbpart>	K	(" ")

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