Team:UT-Tokyo/LabNote

From 2011.igem.org

(Difference between revisions)
Line 190: Line 190:
*Transformation
*Transformation
*Overnight culture on LB plate (with 100ug/ml ampicilline)
*Overnight culture on LB plate (with 100ug/ml ampicilline)
 +
 +
=='11/06/01(Wed)==
 +
*Picking up colony and transfer to LB medium
 +
*Making LB medium
 +
 +
=='11/06/02(Thu)==
 +
*Miniprep
 +
*Making Glycerol(50%)(for cryopreservation)
 +
 +
=='11/06/07(Tue)==
 +
*Nanodrop
 +
*Transformation
 +
*Culture from frozen stock
 +
 +
=='11/06/08(Wed)==
 +
*Picking up colony
 +
 +
=='11/06/09(Thu)==
 +
*Miniprep
 +
 +
=='11/06/13(Mon)==
 +
*Planting Negative control
 +
*Making frozen stock
 +
*Transformation(BBa_E0240,B0032,B0015,B0014,E0040,J31000,J44000)
 +
 +
=='11/06/14(Tue)==
 +
*Picking up colony
 +
*Making Mg reagent
 +
 +
=='11/06/15(Wed)==
 +
*Miniprep
 +
 +
=='11/06/17(Fri)==
 +
*Picking up colony(B0032,B0015,B0040,E0240,J31000,J44000)
 +
*Miniprep(B0014)
 +
*Dissolution(J23119,J23118,K16500,J22106)
 +
 +
=='11/06/21(Tue)==
 +
*Making frozen stock(B0032,B0015,B0040,E0240,J31000,J44000)
 +
*Passafe culture(E0240,J31000)
 +
*Nanodrop again
 +
 +
=='11/06/22(Wed)==
 +
*Miniprep(E0240,J31000)
 +
*Making Competent cell
 +
 +
=='11/06/24(Fri)==
 +
*Digest (product of Miniprep 06/09)(EScut)
 +
*Making agarose gel
 +
*Electrophoresis
 +
 +
=='11/06/28(Tue)==
 +
*Digest(product of Miniprep 06/09)(EPcut)
 +
*Electrophoresis
 +
*Defrost and transformation(BBa_E0030,E0020,I712019,I712052,J52008）
 +
*Making 1×TAE, agalose gel, LB medium, LB plate(amp),
 +
 +
=='11/06/29(Wed)==
 +
*Picking up colony
 +
*Making LB broth
 +
 +
=='11/9/12 (Mon)==
=='11/9/12 (Mon)==

Revision as of 23:26, 14 September 2011

Parts List

Table 1. List of iGEM parts
Number Part Content Plasmid Length (bp)
#1 J23119 c.Promoter (Strong) pSB1A2 35
#2 J23118 c.Promoter (Medium) BBa_J61002 35
#3 B0032 RBS pSB1A2 13
#5 B0014 d.Ter pSB1AK3 95
#9 E0240 RBS-GFP-d.Ter pSB1A2 876
#10 E0040 GFP pSB1A2 681
#11 E1010 RFP pSB2K3 723
#14 I712019 fLuc pSB1AK8 1653
#17 J52008 rLuc pSB1AK3 936
#20 R0011 lacP pSB1A2 55
#21 C0012 LacI pSB1A2 1128
#22 I712074 pT7 pSB1AK8 46
#23 K145001 T7 RNA Pol. pSB1A2 2655
#24 J22106 recAp pSB1A2 192
#27 C0083 AspA pSB2K3 1518
#28 K112808 T4 phage lysis device (no promoter) pSB1A2 1785
#29 - CheZ pSB1AK3 728
#30 K117000 Lysis gene pSB1A2 144
#31 - LexA pSB1AK3 750
#33 - sulAp pSB1AK3 67
#34 - uvrAp pSB1AK3 96
#35 - recNp
#36 R0051 cI-repressed promoter pSB1A2 49
#37 C0051 cI repressor (LVA tagged) pSB1A2 \
#38 - RecA

Lab Diary

For convenience sake, each part (genes, promotors, etc) is represented by consecutive numbers (e.g. #20 for lac promotor). By pointing the mouse on the part number in the construct, you can find out the details of the part.


  • May
  • June
  • July
  • August
  • September
  • October

Please enable Javascript to view this calendar.

'11/5/18 (Wed)

  • Making LB medium, 50×TAE, Tris-HCl (pH8.0)

'11/5/24 (Tue)

  • Making SOB medium, 0.5M EDTA (pH8.0)

'11/5/25(Wed)

  • Making TB(pH=6.7), LB plate

'11/5/26(Thu)

  • Making Mgaq(for SOB medium), Competent cell
  • TB filtration

'11/5/31(Tue)

  • iGEM parts resuspension + frozen stock (at -20℃)
  • Transformation
  • Overnight culture on LB plate (with 100ug/ml ampicilline)

'11/06/01(Wed)

  • Picking up colony and transfer to LB medium
  • Making LB medium

'11/06/02(Thu)

  • Miniprep
  • Making Glycerol(50%)(for cryopreservation)

'11/06/07(Tue)

  • Nanodrop
  • Transformation
  • Culture from frozen stock

'11/06/08(Wed)

  • Picking up colony

'11/06/09(Thu)

  • Miniprep

'11/06/13(Mon)

  • Planting Negative control
  • Making frozen stock
  • Transformation(BBa_E0240,B0032,B0015,B0014,E0040,J31000,J44000)

'11/06/14(Tue)

  • Picking up colony
  • Making Mg reagent

'11/06/15(Wed)

  • Miniprep

'11/06/17(Fri)

  • Picking up colony(B0032,B0015,B0040,E0240,J31000,J44000)
  • Miniprep(B0014)
  • Dissolution(J23119,J23118,K16500,J22106)

'11/06/21(Tue)

  • Making frozen stock(B0032,B0015,B0040,E0240,J31000,J44000)
  • Passafe culture(E0240,J31000)
  • Nanodrop again

'11/06/22(Wed)

  • Miniprep(E0240,J31000)
  • Making Competent cell

'11/06/24(Fri)

  • Digest (product of Miniprep 06/09)(EScut)
  • Making agarose gel
  • Electrophoresis

'11/06/28(Tue)

  • Digest(product of Miniprep 06/09)(EPcut)
  • Electrophoresis
  • Defrost and transformation(BBa_E0030,E0020,I712019,I712052,J52008）
  • Making 1×TAE, agalose gel, LB medium, LB plate(amp),

'11/06/29(Wed)

  • Picking up colony
  • Making LB broth


'11/9/12 (Mon)

  • Colony PCR
    • #20-#3-14-5-2-3-17-5
    • #20-3-14-5-#2-3-17-5
    • #1-#3-14-5-2-3-17-5
    • #36-3-14-5-2-3-17-5-#20-3-37-5 (no colony)
    • #36-3-14-5-2-3-17-5-#20-3-37-5-pSB1A3 (PCR failed?)

Note: #20-3-37-5 colonies on the master-plate are RED. Maybe #2-3-11-5...
Note: #36-3-14-5-2-3-17-5-#20-3-37-5-pSB1A3 PCR amplification doesn't work. Seems failed extension...

  • PCR amplification
    • #20-3-37-5 from Master plate
    • #36-3-14-5-2-3-17-5 from 0911 MP product

Note: Only one #20-3-37-5 colony is NOT RED.

  • Miniprep.
    • #20
    • #29 (pSB1AK3)
    • #2-3-17-5
    • #2-3-37-5
    • BBa_J04450 (pSB1C3)
  • Digest
    • #36-3-14-5-2-3-17-5_ES (from PCR)
    • #20-3-37-5_XP (from PCR)
    • pSB1A3_EP
    • pSB1C3_EP
    • #20-3-37-5_XP (from MP) -> O/N
    • J04450_EP -> O/N
    • #2-9_XP -> O/N
    • #20-3-29-5_ES -> O/N
  • Gel extraction
    • #20_SP
    • #36-3-14-5_SP
    • #2-3-17-5_EX (from O/N digests)
    • #36-3-14-5-2-3-17-5_ES
    • #20-3-37-5_XP (from today's PCR digests)
  • Culture for Miniprep.
    • #20-3-37-5 from 0911 master plate -> O/N
  • Dual Luciferase Assay
    • #20-3-14-5-2-3-17-5
    • #1-3-14-5-2-3-17-5
    • #36-3-14-5-2-3-17-5-20-3-37-5-pSB1A3
  • Ligation -> O/N
    • #36-3-14-5-2-3-17-5_ES-#20-3-37-5_XP-pSB1A3_EP
    • #36-3-14-5-2-3-17-5_ES-#20-3-37-5_XP-pSB1C3_EP
    • #3-27-5_XP-#20_SP
    • #36-3-29-5_ES-#20-3-37-5_XP-pSB1AK3_EP

'11/9/13 (Tue)

  • Colony PCR
    • #36-3-14-5-2-3-17-5_ES-#20-3-37-5_XP-pSB1A3_EP
    • #36-3-14-5-2-3-17-5_ES-#20-3-37-5_XP-pSB1C3_EP (failed)
    • #3-27-5_XP-#20_SP (failed)
    • #36-3-29-5_ES-#20-3-37-5_XP-pSB1AK3_EP
  • Culture for Luciferase Assay
    • #36-3-14-5-2-3-17-5-20-3-37-5(pSB1A3, pSB1C3) (IPTG 0, 1, 10, 38, 100uM)
    • #20-3-14-5-2-3-17-5 (IPTG 0, 1, 10, 38, 100uM)
    • #1-3-14-5-2-3-17-5 (IPTG 0uM)

Note: Seemes OD600 should NOT exceed 0.4~0.5, or intracellular luciferase will be saturated.

  • Miniprep.
    • #20-3-14-5-2-3-17-5
    • #1-3-14-5-2-3-17-5
    • #20-3-37-5 (from a non-red colony on the master plate)
  • Gel ext.
    • #20-3-37-5_XP (from non-red colony)
    • #20-3-29-5_ES(failed)
    • #2-9_XP, pSB1C3_EP

Note: #20-3-29-5 was not cut!

  • Dual Luciferase Assay
    • #1-3-14-5-2-3-17-5 (IPTG 0uM)
    • #20-3-14-5-2-3-17-5 (IPTG 0, 1, 10, 38, 100uM)
    • #36-3-14-5-2-3-17-5-20-3-37-5 (#2-3-11-5?) (IPTG 0, 1, 10, 38, 100uM)
  • Sequencing preparation
    • #20-3-37-5 (from a non-red colony on the master plate)
    • #20-3-29-5
    • #3-27-5
    • #31
  • Digest
    • #20_E
    • #3-29-5_S (from MP products) (cut check)
  • Ligation
    • #20_SP-#3-27-5_XP
  • Transformation -> O/N
    • #3-14-5-2-3-17-5
    • #1-3-14-5-2-3-17-5
    • #20-3-14-5-2-3-17-5 (for plate stock)
    • #20-3-37-5 (from a non-red colony on the master plate) (for plasmid check)

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