BBa_K1833000
1
BBa_K1833000
pT7-eGFP
2015-09-12T11:00:00Z
2015-09-18T11:04:36Z
The RBS-GFP-terminator was obtained from as part BBa_E0840 from the 2015 Distribution (Plate 2, Well 24D). The T7 promoter was obtained as two short oligonucleotides annealed to form sticky ends. The part containing GFP was then cut with EcoRI and XbaI, and ligated with the annealed oligos to form the completed part. Note that due to the choice of technique, a Biobrick scar site remains between the promoter and RBS.
This part produces GFP in the presence of T7 RNA polymerase. It contains a T7 promoter, a strong RBS, the coding region for GFP, and a double terminator. The GFP is a mutant known as GFPmut3b, and the original citation is as follows:
Cormack, B.P., Valdivia, R.H., and S. Falkow. FACS-optimized mutants of green fluorescent protein (GFP). Gene 173: 33-38 (1996). (http://www.sciencedirect.com/science/article/pii/0378111995006850) It has a maximum excitation at 501 nm and maximum emission at 511 nm. For more information, see part BBa_E0040
false
false
_2259_
27388
27388
9
false
The part containing GFP was cut with EcoRI and XbaI, and ligated with the annealed oligos to form the completed part. Note that due to the choice of technique, a Biobrick scar site remains between the promoter and RBS.
false
Konstantin Borisov
component2460119
1
BBa_K1833999
component2460130
1
BBa_E0840
annotation2460119
1
BBa_K1833999
range2460119
1
1
29
annotation2460130
1
BBa_E0840
range2460130
1
30
907
BBa_B0030
1
BBa_B0030
RBS.1 (strong) -- modified from R. Weiss
2003-01-31T12:00:00Z
2015-08-31T04:07:20Z
Released HQ 2013
Strong RBS based on Ron Weiss thesis. Strength is considered relative to <bb_part>BBa_B0031</bb_part>, <bb_part>BBa_B0032</bb_part>, <bb_part>BBa_B0033</bb_part>.
false
true
_44_46_
0
24
7
In stock
false
Varies from -6 to +1 region from original sequence to accomodate BioBricks suffix ("orig" in figure 4-14 of Ron Weiss thesis). <p>No secondary structures are formed in the given RBS region. Users should check for secondary structures induced in the RBS by upstream and downstream elements in the +50 to -50 region, as such structures will greatly affect the strength of the RBS.
Contact info <a href="mailto:(bchow@media.mit.edu)">Brian Chow</a>
true
Vinay S Mahajan, Voichita D. Marinescu, Brian Chow, Alexander D Wissner-Gross and Peter Carr IAP, 2003.
annotation1701
1
RBS-1\Strong
range1701
1
1
15
annotation1702
1
RBS
range1702
1
8
12
annotation7025
1
BBa_B0030
range7025
1
1
15
BBa_B0010
1
BBa_B0010
T1 from E. coli rrnB
2003-11-19T12:00:00Z
2015-08-31T04:07:20Z
Transcriptional terminator consisting of a 64 bp stem-loop.
false
false
_1_
0
24
7
In stock
false
true
Randy Rettberg
annotation4184
1
stem_loop
range4184
1
12
55
annotation7018
1
BBa_B0010
range7018
1
1
80
BBa_K1833999
1
BBa_K1833999
T7 promoter as annealed oligos
2015-09-14T11:00:00Z
2015-09-16T07:58:33Z
Synthesized oligonucleotides as described on main page.
This is a part used as a building block for T7 driven expression. However, this part uses a special method with the following synthesized nucleotides:
pT7sense: aattcgcggccgcttctagataatacgactcactatagggagaa
pT7antisense: ctagttctccctatagtgagtcgtattatctagaagcggccgcg
The following method can be used to insert a T7 promoter before a desired Biobrick part:
1. Cut the desired plasmid (which should contain an RBS and cds at minimum) with EcoRI and XbaI. Purify the plasmid with a method of your choice (both gel purification and PCR purification kits from Qiagen serve to remove the short undesired oligonucleotide from the digestion).
2. Be sure that your oligos have 5' phosphorylation (treat unphosphorylated oligos with T4 Polynucleotide Kinase). Then, anneal the two oligos by slowly cooling from 98C to room temperature.
3. Finally, ligate the cut plasmid with the annealed oligos by using a 1:10 molar ratio of plasmid to insert.
4. Transform the ligation reaction into E. coli, and plate.
5. Pick colonies, and verify the success of the cloning with sequencing or PCR. (Doing a diagnostic digest is not recommended as the size of the original plasmid and the desired plasmid are very similar.)
6. Add a composite part to the iGEM registry by using this part with the original protein coding part.
false
false
_2259_
27388
27388
9
false
When ligating with a cut plasmid, this leaves a Biobrick scar between the T7 promoter and original Biobrick part.
false
Konstantin Borisov
annotation2460116
1
Oligo scar
range2460116
1
24
29
annotation2460074
1
pT7
range2460074
1
1
23
BBa_E0840
1
GFP genera
GFP generator
2004-10-17T11:00:00Z
2015-08-31T04:07:26Z
Released HQ 2013
B0030.E0040.B0015
false
true
_11_1_
0
61
7
In stock
true
true
Jennifer Braff
component1249242
1
BBa_E0040
component1249257
1
BBa_B0012
component1249247
1
BBa_B0010
component1249239
1
BBa_B0030
annotation1249257
1
BBa_B0012
range1249257
1
838
878
annotation1249242
1
BBa_E0040
range1249242
1
22
741
annotation1249247
1
BBa_B0010
range1249247
1
750
829
annotation1249239
1
BBa_B0030
range1249239
1
1
15
BBa_E0040
1
GFP
green fluorescent protein derived from jellyfish Aequeora victoria wild-type GFP (SwissProt: P42212
2004-09-29T11:00:00Z
2016-01-26T02:09:38Z
Released HQ 2013
GFP (mut3b) [note that this part does not have a barcode]
false
true
_11_1_
4206
61
7
In stock
false
true
jcbraff
annotation1934520
1
GFP protein
range1934520
1
1
720
BBa_B0012
1
BBa_B0012
TE from coliphageT7
2003-01-31T12:00:00Z
2015-08-31T04:07:20Z
Derived from the TE terminator of T7 bacteriophage between Genes 1.3 and 1.4 <genbank>V01146</genbank>.
Released HQ 2013
Transcription terminator for the <i>E.coli</i> RNA polymerase.
false
false
_1_
0
24
7
In stock
false
<P> <P>Suggested by Sri Kosuri and Drew Endy as a high efficiency terminator. The 5' end cutoff was placed immediately after the TAA stop codon and the 3' end cutoff was placed just prior to the RBS of Gene 1.4 (before AAGGAG).<P> Use anywhere transcription should be stopped when the gene of interest is upstream of this terminator.
false
Reshma Shetty
annotation1690
1
polya
range1690
1
28
41
annotation1686
1
T7 TE
range1686
1
8
27
annotation7020
1
BBa_B0012
range7020
1
1
41
annotation1687
1
stop
range1687
1
34
34
BBa_B0010_sequence
1
ccaggcatcaaataaaacgaaaggctcagtcgaaagactgggcctttcgttttatctgttgtttgtcggtgaacgctctc
BBa_B0030_sequence
1
attaaagaggagaaa
BBa_K1833000_sequence
1
taatacgactcactatagggagaactagaattaaagaggagaaatactagatgcgtaaaggagaagaacttttcactggagttgtcccaattcttgttgaattagatggtgatgttaatgggcacaaattttctgtcagtggagagggtgaaggtgatgcaacatacggaaaacttacccttaaatttatttgcactactggaaaactacctgttccatggccaacacttgtcactactttcggttatggtgttcaatgctttgcgagatacccagatcatatgaaacagcatgactttttcaagagtgccatgcccgaaggttatgtacaggaaagaactatatttttcaaagatgacgggaactacaagacacgtgctgaagtcaagtttgaaggtgatacccttgttaatagaatcgagttaaaaggtattgattttaaagaagatggaaacattcttggacacaaattggaatacaactataactcacacaatgtatacatcatggcagacaaacaaaagaatggaatcaaagttaacttcaaaattagacacaacattgaagatggaagcgttcaactagcagaccattatcaacaaaatactccaattggcgatggccctgtccttttaccagacaaccattacctgtccacacaatctgccctttcgaaagatcccaacgaaaagagagaccacatggtccttcttgagtttgtaacagctgctgggattacacatggcatggatgaactatacaaataataatactagagccaggcatcaaataaaacgaaaggctcagtcgaaagactgggcctttcgttttatctgttgtttgtcggtgaacgctctctactagagtcacactggctcaccttcgggtgggcctttctgcgtttata
BBa_E0040_sequence
1
atgcgtaaaggagaagaacttttcactggagttgtcccaattcttgttgaattagatggtgatgttaatgggcacaaattttctgtcagtggagagggtgaaggtgatgcaacatacggaaaacttacccttaaatttatttgcactactggaaaactacctgttccatggccaacacttgtcactactttcggttatggtgttcaatgctttgcgagatacccagatcatatgaaacagcatgactttttcaagagtgccatgcccgaaggttatgtacaggaaagaactatatttttcaaagatgacgggaactacaagacacgtgctgaagtcaagtttgaaggtgatacccttgttaatagaatcgagttaaaaggtattgattttaaagaagatggaaacattcttggacacaaattggaatacaactataactcacacaatgtatacatcatggcagacaaacaaaagaatggaatcaaagttaacttcaaaattagacacaacattgaagatggaagcgttcaactagcagaccattatcaacaaaatactccaattggcgatggccctgtccttttaccagacaaccattacctgtccacacaatctgccctttcgaaagatcccaacgaaaagagagaccacatggtccttcttgagtttgtaacagctgctgggattacacatggcatggatgaactatacaaataataa
BBa_E0840_sequence
1
attaaagaggagaaatactagatgcgtaaaggagaagaacttttcactggagttgtcccaattcttgttgaattagatggtgatgttaatgggcacaaattttctgtcagtggagagggtgaaggtgatgcaacatacggaaaacttacccttaaatttatttgcactactggaaaactacctgttccatggccaacacttgtcactactttcggttatggtgttcaatgctttgcgagatacccagatcatatgaaacagcatgactttttcaagagtgccatgcccgaaggttatgtacaggaaagaactatatttttcaaagatgacgggaactacaagacacgtgctgaagtcaagtttgaaggtgatacccttgttaatagaatcgagttaaaaggtattgattttaaagaagatggaaacattcttggacacaaattggaatacaactataactcacacaatgtatacatcatggcagacaaacaaaagaatggaatcaaagttaacttcaaaattagacacaacattgaagatggaagcgttcaactagcagaccattatcaacaaaatactccaattggcgatggccctgtccttttaccagacaaccattacctgtccacacaatctgccctttcgaaagatcccaacgaaaagagagaccacatggtccttcttgagtttgtaacagctgctgggattacacatggcatggatgaactatacaaataataatactagagccaggcatcaaataaaacgaaaggctcagtcgaaagactgggcctttcgttttatctgttgtttgtcggtgaacgctctctactagagtcacactggctcaccttcgggtgggcctttctgcgtttata
BBa_B0012_sequence
1
tcacactggctcaccttcgggtgggcctttctgcgtttata
BBa_K1833999_sequence
1
taatacgactcactatagggagaactaga
igem2sbol
1
iGEM to SBOL conversion
Conversion of the iGEM parts registry to SBOL2.1
Chris J. Myers
James Alastair McLaughlin
2017-03-06T15:00:00.000Z