NYMU Taipei/Construct

From 2007.igem.org

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             <ul>
             <ul>
                 <li>our first contruct for promoter activity assay under glucose stimulus</li>
                 <li>our first contruct for promoter activity assay under glucose stimulus</li>
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                 <li>Insert size: 866bp</li>
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                 <li>Insert size: 866bp (XbaI to SpeI)</li>
                 <li>Total vector size: 3,019bp</li>
                 <li>Total vector size: 3,019bp</li>
                 <li>V-I: 2,153bp</li>
                 <li>V-I: 2,153bp</li>

Revision as of 18:24, 26 October 2007

Contents

Assembly Ordering and Progress

system 1

System 1: glucose-sensing insulin generator (5 task left: 2 PCR and 3 biobrick standard assembly)
pOmpC RBS + TAT
 INSA RBS + TAT
 INSB RBS CinR RBS HSL D-term duration members assembly status
          RBS + CinR RBS + HSL   8/30-8/31 翔 1st assembly 9/01, 07
          CinR + HSL (RBS+)   9/1-9/4 宥,君,威 2nd assembly 9/02, 07
          CinR+HSL(RBS+) + D-term  9/16-9/27 宥,君,翔,pett 3rd assembly 9/27 07
  RBS + TATA+ INSA (PCR) RBS + TATB+ INSB (PCR)            9/28-9/30

銘, pett

 basic part (PCR) 9/30, 07
pOmpC + TATA_INSA (failed) TATB_INSB + CinR+HSL+D-term (wrong assembly)
 -10/5 pett 4th assembly 10/5,7,8 07
pOmpC + TATA_INSA               -10/11 pett 4th assembly
pOmpC + TATA_INS_A + TATB_INS_B
           -10/23   4th assembly
pOmpC + TATA_INS_A + TATB_INS_B + CinR+HSL+D-term     Final

system 2

System 2: insulin-sensing insulin destoryer (5 tasks left: 2 PCR and 3 biobrick standard assembly)
pCinRHSL OmpRBS RBS + TATD
 IDE D-term duration members assembly status
  OmpRBS (PCR) RBS + TATD+ IDE (PCR)
   -102

銘, 茹, pett

 basic part (PCR) 10/2 07
pCinRHSL + OmpRBS       -10/5 pett 1st assembly 10/5 07
pCinRHSL + OmpRBS + TATD_IDE (RBS+)   -10/11  pett 2nd assembly
pCinRHSL+OmpRBS+TATD_IDE + D-term     Final

 

Promoter assay construct: pOmpC+(RBS+EYFP)

POmpC-EYFP (plasmid).jpg
  • our first contruct for promoter activity assay under glucose stimulus
  • Insert size: 866bp (XbaI to SpeI)
  • Total vector size: 3,019bp
  • V-I: 2,153bp
  • May be checked with: BtgZI
  • 8/27, 2007

System 1: pOmpC + (TATA+INSA) + (TATB+INSB) + (RBS+CinR) + (RBS+HSL) + D-term

Phase 1 of system 1: RBS + CinR

RBS-CinR (plasmid).jpg
  • Insert size: 790bp
  • Total vector size: 2943bp
  • V-I: 2153bp
  • May be checked with: BsaBI

Phase 1 of system 1: RBS + HSL

RBS-HSL (plasmid).jpg
  • Insert size: 740bp
  • Total vector size: 2801bp
  • V-I: 2053bp
  • May be checked with: AgeI

Phase 1 of system 1: TATA + INSA = TATA_INSA

TATA INSA (insert).jpg
  • Insert size: 106bp
  • PCR product size: 132bp
  • May be checked with: RsaI

Phase 1 of system 1: TATB + INSB = TATB_INSB

TATB INSB (insert).jpg
  • Insert size: 133bp
  • PCR product size: 159bp
  • May be checked with AluI

Phase 2 of system 1: pOmpC + TATA_INSA

POmpC-TATA INSA (plasmid).jpg
  • Insert size: 230bp
  • Total vector size: 2291bp
  • V-I: 2053bp

Phase 2 of system 1: (RBS+CinR) + (RBS+HSL) = CinR+HSL

<tbody> </tbody>
300px
  • Insert size: 1530bp
  • Total vector size: 3591bp
  • V-I: 2053bp

Phase 3 of system 1: pOmpC + TATA_INSA + TATB_INSB

POmpC-TATA INSA-TATB INSB (plasmid).jpg
  • Insert size: 502bp
  • Total vector size: 2,655bp
  • V-I: 2,153bp

Phase 3 of system 1: CinR+HSL + D-term = CinR+HSL+D-term

CinR-HSL-D-term (plasmid).jpg
  •  
  • item 2
  • item 3
  • item 4
  • item 5

Phase 3 of system 1: pOmpC + TATA_INSA + TATB_INSB + CinR+HSL+D-term

System 2: pCinRHSL+ OmpRBS + (TATD+IDE) + D-term

Phase 1 of system 2: TATD + IDE = TATD_IDE

Phase 1 of system 2: pCinRHSL + OmpRBS

Phase 2 of system 2: pCinRHSL + OmpRBS + TATD_IDE

Phase 3 of system 2: pCinRHSL + OmpRBS +  TATD_IDE + D-term

Basic Constructs

pOmpC in pSB1A2

<tbody> </tbody>
300px
  • BsmFI may be used to confirm the presence of pOmpC.
  • item 2
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  • item 5

pCinRHSL in pSB1A2

<tbody> </tbody>
300px
  • item 1
  • item 2
  • item 3
  • item 4
  • item 5

RBS in pSB1A2

<tbody> </tbody>
300px
  • BseR1 can be applied to check the insertion of RBS
  • item 2
  • item 3
  • item 4
  • item 5

D-term in pSB1AK3

<tbody> </tbody>
300px
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  • item 5

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