IGEM:MIT/2005/Inventory

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 in Lab

Vector in pSB1AT3-1 plasmid

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)

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)

Streaked out on a plate

1. I0500 (pBad/AraC promoter)

Bio Bricks ordered as Stabs

1. R0010
2. R0051
3. I1030
4. I1031
5. I1032
6. I1033
7. J13002

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

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

Ordered/Received:

• Non-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

Specified:

1. ToxR (ATCC)
2. scFv 5' linker (for FecA fusions)
3. scFv fwd flag test (for introducing a start codon)

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