Team:Osaka/week1

From 2011.igem.org

(Difference between revisions)
 
(8 intermediate revisions not shown)
Line 1: Line 1:
 +
{{Osaka}}
 +
__NOTOC__
==August 1 (Mon)==
==August 1 (Mon)==
# Preparation of LB agar plates (33 Amp, 18 Kan, 17 Cam).
# Preparation of LB agar plates (33 Amp, 18 Kan, 17 Cam).
Line 9: Line 11:
#* Preparation of glucose solution for making SOC medium.
#* Preparation of glucose solution for making SOC medium.
#* Single-colony streaking of E. coli (strain DH5α) on 2 LB agar plates -> 37˚C incubation o/n
#* Single-colony streaking of E. coli (strain DH5α) on 2 LB agar plates -> 37˚C incubation o/n
-
 
# Transformation of Registry parts (See Table 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.
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.
Line 16: Line 17:
!ID!!Part Name!!Resistance!!Description
!ID!!Part Name!!Resistance!!Description
|-
|-
-
|1-9N||<bbpart>BBa_K118023</bbpart>||A|| RecA promoter
+
|1-9N||<bbpart>BBa_J22106</bbpart>||A|| rec A (SOS) Promoter
|-
|-
|1-2M||<bbpart>BBa_K118022</bbpart>||A||RBS
|1-2M||<bbpart>BBa_K118022</bbpart>||A||RBS
|-
|-
-
|1-14K||<bbpart>BBa_B0034</bbpart>||A||GFP(wild type)
+
|1-14K||<bbpart>BBa_B0040</bbpart>||A||green fluorescent protein derived from jellyfish Aequeora victoria wild-type GFP
|-
|-
-
|1-12M||<bbpart>BBa_B0010</bbpart>||A||RBS+GFP+double terminator
+
|1-12M||<bbpart>BBa_E0240</bbpart>||A||GFP generator
|-
|-
-
|1-1D||<bbpart>BBa_R0010</bbpart>||A||promoter(lacI regulation)
+
|1-1D||<bbpart>BBa_R0010</bbpart>||A||promoter(lacI regulatd)
|-
|-
-
|3-6H||<bbpart>BBa_E1010</bbpart>||A||CrtE I B with RBS
+
|3-6H||<bbpart>BBa_K274100</bbpart>||A||CrtEBI with rbs
|-
|-
-
|2-18H||<bbpart>BBa_E1010</bbpart>||A||RBS+CrtE
+
|2-18H||<bbpart>BBa_K118014</bbpart>||A||rbs+crtE (geranylgeranyl pyrophosphate synthase)
|-
|-
-
|2-16N||<bbpart>BBa_E1010</bbpart>||A||RBS+CrtB
+
|2-16N||<bbpart>BBa_K118006</bbpart>||A||rbs+crtB (phytoene synthase)
|-
|-
-
|2-18H||<bbpart>BBa_E1010</bbpart>||A||RBS+CrtI
+
|2-18H||<bbpart>BBa_K118005</bbpart>||A||rbs+crtI (phytoene dehydrogenase)
|-
|-
-
|1-17P||<bbpart>BBa_E1010</bbpart>||A||RBS+CrtY
+
|1-17P||<bbpart>BBa_I742155</bbpart>||A||crtY (lycopene cyclase) with native rbs
|-
|-
-
|1-1G||<bbpart>BBa_E1010</bbpart>||A||construction plasmid containing mRFP coding device  
+
|1-1G||<bbpart>BBa_J04450</bbpart>||A||construction plasmid containing mRFP coding device  
|-
|-
-
|1-3A||<bbpart>BBa_E1010</bbpart>||C||construction plasmid containing mRFP coding device  
+
|1-3A||<bbpart>BBa_J04450</bbpart>||C||construction plasmid containing mRFP coding device  
|-
|-
-
|1-5A||<bbpart>BBa_E1010</bbpart>||K||construction plasmid containing mRFP coding device  
+
|1-5A||<bbpart>BBa_J04450</bbpart>||K||construction plasmid containing mRFP coding device  
|}
|}
Tomorrow we shall prepare the remainder of the reagents and inoculate SOB with o/n-incubated E. coli.
Tomorrow we shall prepare the remainder of the reagents and inoculate SOB with o/n-incubated E. coli.
Line 69: Line 70:
#* 04(K): 1-1D(upstream), 3-6H(downstream), 1-5A(vector)
#* 04(K): 1-1D(upstream), 3-6H(downstream), 1-5A(vector)
#* 05(K): 1-1D(upstream), 2-18H(downstream), 1-5A(vector)
#* 05(K): 1-1D(upstream), 2-18H(downstream), 1-5A(vector)
-
#* 06(k): 1-9N(upstream), 2-16L(downstream), 1-5A(vector)
+
#* 06(K): 1-9N(upstream), 2-16L(downstream), 1-5A(vector)
# Transformation of Registry parts (See Table 2).
# Transformation of Registry parts (See Table 2).
{|style="border-collapse: separate; border-spacing: 0; border-width: 1px; border-style: solid; border-color: #000; padding:5px;" cellpadding="5"
{|style="border-collapse: separate; border-spacing: 0; border-width: 1px; border-style: solid; border-color: #000; padding:5px;" cellpadding="5"
Line 75: Line 76:
!ID!!Part Name!!Resistance!!Description
!ID!!Part Name!!Resistance!!Description
|-
|-
-
|1-23L||<bbpart>BBa_K118023</bbpart>||A|| double terminator
+
|1-23L||<bbpart>BBa_B0015</bbpart>||A|| double terminator
|}
|}
Line 83: Line 84:
# Transformation of Ligation products
# Transformation of Ligation products
# Preparation of LB plates (33 Amp, 14 Kan)
# Preparation of LB plates (33 Amp, 14 Kan)
 +
[[Team:Osaka/Notebook|Back to Notebook]]
[[Team:Osaka/Notebook|Back to Notebook]]
 +
 +
week '''1'''|[[Team:Osaka/week2|2]]|[[Team:Osaka/week3|3]]|[[Team:Osaka/week4|4]]|[[Team:Osaka/week5|5]]|[[Team:Osaka/week6|6]]|[[Team:Osaka/week7|7]]|[[Team:Osaka/week8|8]]|[[Team:Osaka/week9|9]]|[[Team:Osaka/week10|10]]|[[Team:Osaka/week11|11]]|[[Team:Osaka/week12|12]]|[[Team:Osaka/week13|13]]

Latest revision as of 05:53, 19 October 2011

August 1 (Mon)

  1. Preparation of LB agar plates (33 Amp, 18 Kan, 17 Cam).


August 2 (Tue)

  1. Culture medium preparation
  2. Competent cells preparation - Nojima Method
    • SOB medium (MgCl2 not yet added) -> stored at 4˚C
    • Preparation of glucose solution for making SOC medium.
    • Single-colony streaking of E. coli (strain DH5α) on 2 LB agar plates -> 37˚C incubation o/n
  3. 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
IDPart NameResistanceDescription
1-9N<bbpart>BBa_J22106</bbpart>A rec A (SOS) Promoter
1-2M<bbpart>BBa_K118022</bbpart>ARBS
1-14K<bbpart>BBa_B0040</bbpart>Agreen fluorescent protein derived from jellyfish Aequeora victoria wild-type GFP
1-12M<bbpart>BBa_E0240</bbpart>AGFP generator
1-1D<bbpart>BBa_R0010</bbpart>Apromoter(lacI regulatd)
3-6H<bbpart>BBa_K274100</bbpart>ACrtEBI with rbs
2-18H<bbpart>BBa_K118014</bbpart>Arbs+crtE (geranylgeranyl pyrophosphate synthase)
2-16N<bbpart>BBa_K118006</bbpart>Arbs+crtB (phytoene synthase)
2-18H<bbpart>BBa_K118005</bbpart>Arbs+crtI (phytoene dehydrogenase)
1-17P<bbpart>BBa_I742155</bbpart>AcrtY (lycopene cyclase) with native rbs
1-1G<bbpart>BBa_J04450</bbpart>Aconstruction plasmid containing mRFP coding device
1-3A<bbpart>BBa_J04450</bbpart>Cconstruction plasmid containing mRFP coding device
1-5A<bbpart>BBa_J04450</bbpart>Kconstruction plasmid containing mRFP coding device

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


August 3 (Wed)

  1. Competent cells preparation (continued)
    • Inoculation of pre-culture SOB from o/n-incubated agar plate colonies.
    • Transfer from pre-culture to growth culture.
  2. Colony check
    • All parts successfully transformed
  3. Transfer to LB culture medium:1-9N,1-2M,1-14K,1-12M,1-1D,1-17P,1-1G,1-5A,2-16,2-18H,2-16N,1-3A,3-6H


August 4 (Thu)

  1. OD measurements throughout the day till required OD (0.3~0.7) was obtained.
  2. Completion of competent cells according to protocol.
  3. Miniprep of 1-9N,1-2M,1-14K,1-12M,1-1D,1-17P,1-1G,1-5A,2-16,2-18H,2-16N,1-3A,3-6H


August 5 (Fri)

  1. Restriction digests of 1-9N,1-2M,1-14K,1-12M,1-1D,1-17P,1-1G,1-5A,2-16,2-18H,2-16N,1-3A,3-6H
  2. Gel electrophoresis
  3. Ligation
    • 01(K): 1-9N(upstream), 1-12M(downstream), 1-5A(vector)
    • 02(K): 1-1D(upstream), 1-12M(downstream), 1-5A(vector)
    • 03(K): 1-9N(upstream), 3-6H(downstream), 1-5A(vector)
    • 04(K): 1-1D(upstream), 3-6H(downstream), 1-5A(vector)
    • 05(K): 1-1D(upstream), 2-18H(downstream), 1-5A(vector)
    • 06(K): 1-9N(upstream), 2-16L(downstream), 1-5A(vector)
  4. Transformation of Registry parts (See Table 2).
Table2
IDPart NameResistanceDescription
1-23L<bbpart>BBa_B0015</bbpart>A double terminator


August 6 (Sat)

  1. Transfer to LB culture medium:1-23L
  2. Transformation of Ligation products
  3. Preparation of LB plates (33 Amp, 14 Kan)


Back to Notebook

week 1|2|3|4|5|6|7|8|9|10|11|12|13

Retrieved from "http://2011.igem.org/Team:Osaka/week1"