Team:Stockholm/22 September 2010

From 2010.igem.org

(Difference between revisions)
(Andreas)
m
 
(3 intermediate revisions not shown)
Line 1: Line 1:
{{Stockholm/Top2}}
{{Stockholm/Top2}}
 +
==Andreas==
==Andreas==
===Gel verification===
===Gel verification===
Line 22: Line 23:
===ON cultures===
===ON cultures===
#5 ml LB + appr. antibiotic (100 Amp, 25 Cm or 50 Km); 37 °C, 225 rpm
#5 ml LB + appr. antibiotic (100 Amp, 25 Cm or 50 Km); 37 °C, 225 rpm
-
#*pSB1A2.RBS⋅yCCS: clones 2, 3 and 4
+
#*pSB1A2.RBS⋅yCCS: clones 3 and 4
#*pEX.N-TAT⋅SOD⋅His: clones 3 and 4
#*pEX.N-TAT⋅SOD⋅His: clones 3 and 4
#*pSB1C3.N-LMWP⋅SOD⋅His: clones 1 and 4
#*pSB1C3.N-LMWP⋅SOD⋅His: clones 1 and 4
#*pSB1K3.N-LMWP⋅SOD⋅His: clone 1
#*pSB1K3.N-LMWP⋅SOD⋅His: clone 1
 +
 +
::'''***Note to self***'''<br />
 +
::''It might be possible that pSB1A2.RBS&sdot;yCCS clones have been mixed up with pEX.N-TAT&sdot;SOD&sdot;His. After plasmid prep, this should be tested by digesting the samples and analyze digestion sizes.
 +
 +
 +
 +
 +
 +
 +
 +
== Mimmi ==
 +
 +
=== SOD / SOD.his / his.SOD / yCCS ===
 +
 +
{|
 +
| colspan="3" | ''colony PCR''
 +
| rowspan="8" width="100" |
 +
! samples
 +
| rowspan="8" width="100" |
 +
! colspan="2" | Conditions
 +
| rowspan="3" |
 +
|-
 +
| colspan="3" |
 +
| pEX.SOD x3
 +
! time
 +
! &deg;C
 +
|-
 +
! mix
 +
| (µl)
 +
| x13
 +
| pEX.SOD.his x3
 +
| 5m
 +
| 95
 +
|-
 +
| mastermix
 +
| 24.5
 +
| rowspan="5" |
 +
| pEX.his.SOD x3
 +
| 30s
 +
| 95
 +
| )
 +
|-
 +
| DNA
 +
| 0.5
 +
| pEX.yCCS x3
 +
| 30s
 +
| 55
 +
| > 30 cycles
 +
|-
 +
| align="right" | tot
 +
| 25µl
 +
| rowspan="3" |
 +
| 1m20s
 +
| 72
 +
| )
 +
|-
 +
| colspan="2" rowspan="2" |
 +
| 5m
 +
| 72
 +
|-
 +
| oo
 +
| 25
 +
|}
 +
 +
 +
 +
==== Gel ====
 +
 +
[[Image:2010-09-22_pEX.SOD_+_yCCS.jpg|200px|thumb|left|]]
 +
{|
 +
! well
 +
! width="120" | sample
 +
! well
 +
! sample
 +
|-
 +
| 1
 +
| ladder
 +
| 9
 +
| ladder
 +
|-
 +
| 2
 +
| pEX.SOD.his 1
 +
| 10
 +
| pEX.SOD 1
 +
|-
 +
| 3
 +
| pEX.SOD.his 2
 +
| 11
 +
| pEX.SOD 2
 +
|-
 +
| 4
 +
| pEX.SOD.his 3
 +
| 12
 +
| pEX.SOD 3
 +
|-
 +
| 5
 +
| pEX.his.SOD 1
 +
| 13
 +
| pEX.yCCS 1
 +
|-
 +
| 6
 +
| pEX.his.SOD 1
 +
| 14
 +
| pEX.yCCS 2
 +
|-
 +
| 7
 +
| pEX.his.SOD 1
 +
| 15
 +
| pEX.yCCS 2
 +
|-
 +
| 8
 +
| pEX.SOD.his
 +
| 16
 +
| -
 +
|}
 +
 +
{{Stockholm/Footer}}

Latest revision as of 11:05, 26 October 2010


Contents

Andreas

Gel verification

Colony PCR gel verification of new assemblies: pSB1A2.RBS.yCCS, pEX.N-TAT*SOD*His, pSB1C3.N-LMWP*SOD*His & pSB1K3.N-LMWP*SOD*His.
4 μl λ; 4 μl sample.
λ = O'GeneRuler 1 kb DNA ladder.

Of 21/9 colony PCR samples

1 % agarose, 120 V

Expected bands

  • pSB1A2.RBS⋅yCCS (pA.RBSy): 1018 bp
  • pEX.N-TAT⋅SOD⋅His (pEX.nTSH): 735 bp
  • pSB1C3.N-LMWP⋅SOD⋅His (pC.nLSH): 857 bp
  • pSB1K3.N-LMWP⋅SOD⋅His (pK.nLSH): 857 bp
  • pSB1C3.SOD⋅His (PC): 815 bp

Results

  • pSB1A2.RBS⋅yCCS: Seemingly correct-sized bands for clones 2, 3 and 4.
  • pEX.N-TAT⋅SOD⋅His: Seemingly correct-sized bands for clones 3 & 4.
  • pSB1C3.N-LMWP⋅SOD⋅His: Seemingly correct-sized bands for clones 1 & 4.
  • pSB1K3.N-LMWP⋅SOD⋅His: Seemingly correct-sized band for clone 1.

ON cultures

  1. 5 ml LB + appr. antibiotic (100 Amp, 25 Cm or 50 Km); 37 °C, 225 rpm
    • pSB1A2.RBS⋅yCCS: clones 3 and 4
    • pEX.N-TAT⋅SOD⋅His: clones 3 and 4
    • pSB1C3.N-LMWP⋅SOD⋅His: clones 1 and 4
    • pSB1K3.N-LMWP⋅SOD⋅His: clone 1
***Note to self***
It might be possible that pSB1A2.RBS⋅yCCS clones have been mixed up with pEX.N-TAT⋅SOD⋅His. After plasmid prep, this should be tested by digesting the samples and analyze digestion sizes.




Mimmi

SOD / SOD.his / his.SOD / yCCS

colony PCR samples Conditions
pEX.SOD x3 time °C
mix (µl) x13 pEX.SOD.his x3 5m 95
mastermix 24.5 pEX.his.SOD x3 30s 95 )
DNA 0.5 pEX.yCCS x3 30s 55 > 30 cycles
tot 25µl 1m20s 72 )
5m 72
oo 25


Gel

2010-09-22 pEX.SOD + yCCS.jpg
well sample well sample
1 ladder 9 ladder
2 pEX.SOD.his 1 10 pEX.SOD 1
3 pEX.SOD.his 2 11 pEX.SOD 2
4 pEX.SOD.his 3 12 pEX.SOD 3
5 pEX.his.SOD 1 13 pEX.yCCS 1
6 pEX.his.SOD 1 14 pEX.yCCS 2
7 pEX.his.SOD 1 15 pEX.yCCS 2
8 pEX.SOD.his 16 -





The Faculty of Science at Stockholm University Swedish Vitiligo association (Svenska Vitiligoförbundet) Geneious Fermentas/ Sigma-Aldrich/