IGEM:IMPERIAL/2006/project/Oscillator/project browser/Test Killing Predator Construct/Implementation

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Super Parts Predator Construct
Actual Part AiiA Testing Construct <bbpart>J37022</bbpart>
Sub Parts <bbpart>R0010</bbpart> <bbpart>B0034</bbpart> <bbpart>J37023</bbpart> <bbpart>B0015</bbpart>


Ligation Process

Step 1: Construct <bbpart>J37023</bbpart> from Part <bbpart>C0060</bbpart> and a FLAG immunotag by PCR (sequence from Dr. Mann)

Step 2: Construct <bbpart>J37024</bbpart> by adding a terminator (<bbpart>B0015</bbpart> to J37023

Step 3: Construct <bbpart>J37022</bbpart> by adding the LacI promoter (<bbpart>R0010</bbpart>) and the RBS <bbpart>B0034</bbpart>, which is contained within the part <bbpart>J04500</bbpart>.

People Involved with Ligations:

<bbpart>J37022</bbpart> (Final AiiA test construct)

  • <bbpart>J04500</bbpart> (16P) + <bbpart>J37024</bbpart>
  • J37024 (insert) cut with XbaI & PstI, expected size = 911 bp (on pSB1AK3 = 3189 bp)
  • J04500 (16P) (vector) cut with SpeI & PstI, expected size = 220 bp + 3189 (pSB1AK3) = 3409 bp
  • Miniprep sizes expected: 1131 bp & 3189 bp (pSB1AK3)

<bbpart>J37023</bbpart> (for predator cell/test construct)

  • AiiA with Immunotag
  • Done by PCR
  • Status: Complete (awaiting sequencing results)
  • Expected Part Length = 816 bp
  • Expected Vector Length = Unknown (Hanging T tail vector)

<bbpart>J37024</bbpart> (for AiiA predator cell/test construct)

  • <bbpart>J37023</bbpart> + <bbpart>B0015</bbpart>
    • J37023 (insert) cut with EcoRI & SpeI, expected size = 816 bp
    • B0014 (1G) (vector) cut with EcoRI & XbaI, expected size = 95 bp + 3189 bp (pSB1AK3) = 3284 bp
  • Miniprep sizes expected: 911 bp & 3189 bp (pSB1AK3)
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