IGEM:MIT/2005/Inventory: Difference between revisions

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===Primers===
===Primers===
====Specified:====
Please click link in order to see [[../Sequences of primers specified/]]. They consist of the following:
Please click link in order to see [[../Sequences of primers specified/]]. They consist of the following:


Line 125: Line 126:
#<b>FecR</b> fwd, rev, mid-fwd, mid-rev
#<b>FecR</b> fwd, rev, mid-fwd, mid-rev
#<b>ToxR ATCC</b> fwd, rev
#<b>ToxR ATCC</b> fwd, rev
#<b>''Linker''</b> fwd1
#<b>''Linker''</b> fwd1, rev1, fwd2, rev2, rev3,
#<b>''FecA Promoter''</b>  
#<b>scFv fwd flag test</b> (for introducing a start codon)
#<b>FecA knockout with Tetracycline marker</b>
#<b>Fur knockout with Chloramphenicol marker</b>
#<b>FecA promoter</b> Reverse, version2
#<B>''Fusion''</b>  
#<B>''Fusion''</b>  
  <u>Arrived: 8/4/2005</u>
  <u>Arrived: 8/4/2005</u>
Line 150: Line 154:
  Rev571-574
  Rev571-574


====Specified:====
 
#<b>ToxR</b> (ATCC)
#<b>scFv 3' linker</b>
#<b>scFv fwd flag test</b> (for introducing a start codon)
#<b>FecA knockout with Tetracycline marker</b>
#<b>Fur knockout with Chloramphenicol marker</b>
#<b>FecA'-scFv fusion</b>
#<b>FecA promoter</b>
(n.b: "mid" refers to primers intended to introduce point mutations to take out restriction sites in our parts)
(n.b: "mid" refers to primers intended to introduce point mutations to take out restriction sites in our parts)

Latest revision as of 12:06, 5 August 2005

File:PlasmidInventory IGEM2005.xls

Strains

E.coli Strain Genotypes

  1. NB115 (pCT-4-4-20) -- 7.02 -80°C
  2. NB116 (pCT302 S101A) -- 7.02 -80°C
  3. NB118 (pCT-4M5.3) -- 7.02 -80°C
  4. MC4100
    • Genotype: F- araD139 delta(argF-lac)U169 prsL150 relA1 deoC1 rbsR fthD5301 fruA25 lambda- [s]

Plasmids

[[../Plasmid Info/]]

  1. pSB1AK3-1 -- as plate, to be made into glycerol stock thursday
  2. pSB1AC3-1 -- ditto
  3. pSB1AT3-1 -- ditto
  4. pSB2K3-1 -- ditto
  5. File:PCT302.pdf
  6. [[../PCT302 (aka pCT-4-4-20)/]]-- 7.02 4°C
  7. [[../PCT302 S101A/]] -- 7.02 4°C
  8. [[../PCT-4M5.3/]] -- 7.02 4°C

PCR Amplification

  1. Spin PCR purification kit ()
  2. Platinum, TAQ polymerase (DNA polymerase for PCR)
  3. DNTP mix (25mm ea. dntp) (nucleotides for PCR)

Agarose gels

  1. Agarose 100g (pouring gels)
  2. 10X TAE buffer (pouring/running gels)
  3. 1 kb DNA ladder 250µg (molecular weight standard)
  4. Ethidium Bromide Solution (10mg/mL) (staining DNA)

Cutting/joining DNA fragments

  1. EcoR1 10,000 u (Restriction Enzyme)
  2. PstI 10,000 u (Restriction Enzyme)
  3. SpeI 2500 u (Restriction Enzyme)
  4. Xba1 3,000 u (Restriction Enzyme)
  5. T4 DNA Ligase 100,000un (Ligating DNA fragments)

Purifying DNA

  1. Spin Plasmid Kit, Mini(for minipreps)
  2. Gel Extraction Kit 50 (purifying from gel slices)
  3. Spin PCR Purification Kit (purifying PCR products)

Microbiology supplies

  1. LB amp -- Cold Room
  2. Amp stock -- -20 Fridge
  3. VWR Petri Dish (100x15mm, case of 500) (for pouring plates)
  4. Test Tube, Culture (Polystyrene, 12x75, case of 1000) (special tubes for FACS)


Bio Bricks

pSB1AT3-1

  1. BBa_J07012 scFv anti-fluorescein [4-4-20]
  2. BBa_J07013 scFv anti-fluorescein [S101A]
  3. BBa_J07014 scFv anti-fluorescein [4M5.3]
  4. BBa_J07017 FecA protein (full length)

pSB1AC3

  1. BBa_J07021 FecR
  2. BBa_J07022 FecI

pSB1AK3

  1. BBa_J07043 J07012.B0015 scFv4-4-20.TT
  2. BBa_J07044 J07013.B0015 scFvS101A.TT
  3. BBa_J07045 J07014.B0015 scFv4M5.3.TT

Miniprepped DNA

  1. R0053 -- Amp -- KBlab -- 4°C
  2. R0040 -- Amp -- KBlab -- 4°C
  3. R0011 -- Amp -- KBlab -- 4°C
  4. [[../Quad part inverters/]]
  5. E0420 -- Amp -- KBlab -- 4°C
  6. E0430 -- Amp -- KBlab -- 4°C
  7. E0840 (rbs:gfp:term)
  8. B0015 (term)
  9. R0040 (tetR promoter)
  10. R0011 (lacI regulated promoter)
  11. B0030 (Strong RBS)
  12. J13002 (R0040.B0034)

Streaked out on a plate

  1. I0500 (pBad/AraC promoter)

Ordered as Stabs

  1. R0010
  2. R0051
  3. J04500
  4. I1030
  5. I1031
  6. I1032
  7. I1033

Assemblies Ordered from Registry

note: the registry probably will not make these in any reasonable time frame. We need to make them ourselves.

  1. J07036
  2. J07037
  3. J07038
  4. J07039
  5. J07040
  6. J07041
  7. J07042


Planning/Building Bio Bricks

  1. BBa_J07005 [msr] E.coli JM83 alkaline phosphatase gene (PhoA) 1416 bp
  2. BBa_J07006 [tmp] malE 1191 bp
  3. BBa_J07007 [reg] ctx promoter 145 bp
  4. BBa_J07008 [tmp] ToxR w.t. (from Vibrio Cholarae, aa's 1-294) 1160 bp
  5. BBa_J07009 [reg] ToxR' (aa's 1-210) 630 bp
  6. BBa_J07010 [reg] ToxR inner (aa's 1-198; cytoplasm + TM) 594 bp
  7. BBa_J07011 [rpt] ctx promoter :: gfp reporter 871 bp

Primers

Specified:

Please click link in order to see [[../Sequences of primers specified/]]. They consist of the following:

Ordered/Received:

  • Not completely or not received primers are listed in italics
  1. scFv fwd, rev
  2. FecA fwd, rev, mid-fwd, mid-rev
  3. FecA promoter fwd, rev
  4. MalE fwd, rev
  5. PhoA fwd, rev, mid1-fwd, mid1-rev, mid2-fwd, mid2-rev
  6. FecI fwd, rev
  7. FecR fwd, rev, mid-fwd, mid-rev
  8. ToxR ATCC fwd, rev
  9. Linker fwd1, rev1, fwd2, rev2, rev3,
  10. scFv fwd flag test (for introducing a start codon)
  11. FecA knockout with Tetracycline marker
  12. Fur knockout with Chloramphenicol marker
  13. FecA promoter Reverse, version2
  14. Fusion
Arrived: 8/4/2005
fwd & rev138
fwd & rev155
fwd & rev176
fwd & rev178
fwd180
fwd & rev527
fwd & rev571
fwd & rev138-141
fwd & rev155-158
fwd & rev176-180
fwd & rev178-182 
fwd & rev180-184
fwd & rev521-529
fwd & rev523-531
fwd & rev527-531
fwd & rev567-576 
fwd571-574
Arrived: 8/5/2005
Rev180
Rev571-574


(n.b: "mid" refers to primers intended to introduce point mutations to take out restriction sites in our parts)

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