Team:NYMU-Taipei/Project/Speedy protein degrader/Sequences
From 2010.igem.org
Contents |
Sequences
SsrA tags
SSrA degradation tags to use:
Tag | Amino Acid | Sequence |
[ LVA]BBa_M0040 | RPAANDENYALVA | gctgcaaacgacgaaaactacgctttagtagct |
Fluorescent proteins
Legend
Effect | Class |
---|---|
Primers: (changes overline/underline and background) | |
Forward Primer | FP |
Overlapping Primer | OP |
Reverse Primer | RP |
Parts: (changes text color) | |
Promoter | P |
Ribosome Binding site | R |
RNA | rna |
Coding region | C |
Terminator | T |
Special | S |
Cutting sites: (changes background) | |
Biobrick cutting site | cut |
Biobrick cutting site mutation | cutmut |
Fluorescent proteins to add degradation tags onto.
- [http://openwetware.org/wiki/IGEM:IMPERIAL/2007/Projects/Reporters#bibkey_Andersen FP comparison]
- Maturation time is the time until the protein starts shining.
GFP
- Used [http://partsregistry.org/wiki/index.php/Part:BBa_E0240 BBa_E0240] as a template:
>[http://partsregistry.org/Part:BBa_E0240 BBa_E0240] | 876bp |
tcacacaggaaagtactagatgcgtaaaggagaagaacttttcactggagttgtcccaattcttgttgaattagatggtgatgttaatgggcacaaattt
tctgtcagtggagagggtgaaggtgatgcaacatacggaaaacttacccttaaatttatttgcactactggaaaactacctgttccatggccaacacttg
tcactactttcggttatggtgttcaatgctttgcgagatacccagatcatatgaaacagcatgactttttcaagagtgccatgcccgaaggttatgtaca
ggaaagaactatatttttcaaagatgacgggaactacaagacacgtgctgaagtcaagtttgaaggtgatacccttgttaatagaatcgagttaaaaggt
attgattttaaagaagatggaaacattcttggacacaaattggaatacaactataactcacacaatgtatacatcatggcagacaaacaaaagaatggaa
tcaaagttaacttcaaaattagacacaacattgaagatggaagcgttcaactagcagaccattatcaacaaaatactccaattggcgatggccctgtcct
tttaccagacaaccattacctgtccacacaatctgccctttcgaaagatcccaacgaaaagagagaccacatggtccttcttgagtttgtaacagctgct
gggattacacatggcatggatgaactatacaaataataatactagagccaggcatcaaataaaacgaaaggctcagtcgaaagactgggcctttcgtttt
atctgttgtttgtcggtgaacgctctctactagagtcacactggctcaccttcgggtgggcctttctgcgtttata
BBa_E0240+LVA (note LVA was added before the taataa double-stop codon)
>BBa_E0240+LVA | 909bp |
tcacacaggaaagtactagatgcgtaaaggagaagaacttttcactggagttgtcccaattcttgttgaattagatggtgatgttaatgggcacaaattt
tctgtcagtggagagggtgaaggtgatgcaacatacggaaaacttacccttaaatttatttgcactactggaaaactacctgttccatggccaacacttg
tcactactttcggttatggtgttcaatgctttgcgagatacccagatcatatgaaacagcatgactttttcaagagtgccatgcccgaaggttatgtaca
ggaaagaactatatttttcaaagatgacgggaactacaagacacgtgctgaagtcaagtttgaaggtgatacccttgttaatagaatcgagttaaaaggt
attgattttaaagaagatggaaacattcttggacacaaattggaatacaactataactcacacaatgtatacatcatggcagacaaacaaaagaatggaa
tcaaagttaacttcaaaattagacacaacattgaagatggaagcgttcaactagcagaccattatcaacaaaatactccaattggcgatggccctgtcct
tttaccagacaaccattacctgtccacacaatctgccctttcgaaagatcccaacgaaaagagagaccacatggtccttcttgagtttgtaacagctgct
gggattacacatggcatggatgaactatacaaaGCTGCAAACGACGAAAACTACGCTTTAGTAGCTtaataatactagagccaggcatcaaataaaacgaaaggctcagtcgaaagactgggcctttcgtttt
atctgttgtttgtcggtgaacgctctctactagagtcacactggctcaccttcgggtgggcctttctgcgtttata
Primers designed by Jesse in iGEM2008.
>BBa_E0240+LVA | 909bp |
tcacacaggaaagtactagatgcgtaaaggagaagaacttttcactggagttgtcccaattcttgttgaattagatggtgatgttaatgggcacaaattt
tctgtcagtggagagggtgaaggtgatgcaacatacggaaaacttacccttaaatttatttgcactactggaaaactacctgttccatggccaacacttg
tcactactttcggttatggtgttcaatgctttgcgagatacccagatcatatgaaacagcatgactttttcaagagtgccatgcccgaaggttatgtaca
ggaaagaactatatttttcaaagatgacgggaactacaagacacgtgctgaagtcaagtttgaaggtgatacccttgttaatagaatcgagttaaaaggt
attgattttaaagaagatggaaacattcttggacacaaattggaatacaactataactcacacaatgtatacatcatggcagacaaacaaaagaatggaa
tcaaagttaacttcaaaattagacacaacattgaagatggaagcgttcaactagcagaccattatcaacaaaatactccaattggcgatggccctgtcct
tttaccagacaaccattacctgtccacacaatctgccctttcgaaagatcccaacgaaaagagagaccacatggtccttcttgagtttgtaacagctgct
gggattacacatggcatggatgaactatacaaaGCTGCAAACGACGAAAACTACGCTTTAGTAGCTtaataatactagagccaggcatcaaataaaacgaaaggctcagtcgaaagactgggcctttcgtttt
atctgttgtttgtcggtgaacgctctctactagagtcacactggctcaccttcgggtgggcctttctgcgtttata
Primers: (directly copied from iGEM2008 wiki.) Outer primers: gaattcgcggccgcttctagag tcacacaggaaagtactagatgc (prefix + anneal: len:23, tm:56, %gc:43) ctgcagcggccgctactagta ataaacgcagaaaggccca (suffix + anneal: len:19, tm:56, %gc:47) Inner primers: Forward: CGACGAAAACTACGCTTTAGTAGCT taataatactagagccaggcatca (insert + anneal: len:24, tm:55, %gc38) Reverse: AAGCGTAGTTTTCGTCGTTTGCAGC tttgtatagttcatccatgccatg (insert + anneal: len:24, tm:55, %gc:38) Complimented part: CGACGAAAACTACGCTT (len:17, tm:50, %gc:47)
Final product is a Linear PCR DNA containing E0240+LVA.
Redesign Outer FP to include B0034 instead of B0032 (Lets save a cloning cycle)
XbaI+B0034+scar+E0040 start gcttctagag aaagaggagaaa tactag atgcgtaaaggagaagaacttttc (55C,24bp,38%GC) Total:52bp
RFP
PCR of [http://partsregistry.org/Part:BBa_I13507 BBa_I13507]
>[http://partsregistry.org/Part:BBa_I13507 BBa_I13507] | 861bp |
aaagaggagaaatactagatggcttcctccgaagacgttatcaaagagttcatgcgtttcaaagttcgtatggaaggttccgttaacggtcacgagttcg
aaatcgaaggtgaaggtgaaggtcgtccgtacgaaggtacccagaccgctaaactgaaagttaccaaaggtggtccgctgccgttcgcttgggacatcct
gtccccgcagttccagtacggttccaaagcttacgttaaacacccggctgacatcccggactacctgaaactgtccttcccggaaggtttcaaatgggaa
cgtgttatgaacttcgaagacggtggtgttgttaccgttacccaggactcctccctgcaagacggtgagttcatctacaaagttaaactgcgtggtacca
acttcccgtccgacggtccggttatgcagaaaaaaaccatgggttgggaagcttccaccgaacgtatgtacccggaagacggtgctctgaaaggtgaaat
caaaatgcgtctgaaactgaaagacggtggtcactacgacgctgaagttaaaaccacctacatggctaaaaaaccggttcagctgccgggtgcttacaaa
accgacatcaaactggacatcacctcccacaacgaagactacaccatcgttgaacagtacgaacgtgctgaaggtcgtcactccaccggtgcttaataac
gctgatagtgctagtgtagatcgctactagagccaggcatcaaataaaacgaaaggctcagtcgaaagactgggcctttcgttttatctgttgtttgtcg
gtgaacgctctctactagagtcacactggctcaccttcgggtgggcctttctgcgtttata
>[http://partsregistry.org/Part:BBa_I13507 BBa_I13507]+LVA | 869bp |
aaagaggagaaatactagatggcttcctccgaagacgttatcaaagagttcatgcgtttcaaagttcgtatggaaggttccgttaacggtcacgagttcg
aaatcgaaggtgaaggtgaaggtcgtccgtacgaaggtacccagaccgctaaactgaaagttaccaaaggtggtccgctgccgttcgcttgggacatcct
gtccccgcagttccagtacggttccaaagcttacgttaaacacccggctgacatcccggactacctgaaactgtccttcccggaaggtttcaaatgggaa
cgtgttatgaacttcgaagacggtggtgttgttaccgttacccaggactcctccctgcaagacggtgagttcatctacaaagttaaactgcgtggtacca
acttcccgtccgacggtccggttatgcagaaaaaaaccatgggttgggaagcttccaccgaacgtatgtacccggaagacggtgctctgaaaggtgaaat
caaaatgcgtctgaaactgaaagacggtggtcactacgacgctgaagttaaaaccacctacatggctaaaaaaccggttcagctgccgggtgcttacaaa
accgacatcaaactggacatcacctcccacaacgaagactacaccatcgttgaacagtacgaacgtgctgaaggtcgtcactccaccggtgctGCTGCAA
ACGACGAAAACTACGCTTTAGTAGCTtaataatactagagccaggcatcaaataaaacgaaaggctcagtcgaaagactgggcctttcgttttatctgtt
gtttgtcggtgaacgctctctactagagtcacactggctcaccttcgggtgggcctttctgcgtttata
>[http://partsregistry.org/Part:BBa_I13507 BBa_I13507]+LVA | 869bp |
aaagaggagaaatactagatggcttcctccgaagacgttatcaaagagttcatgcgtttcaaagttcgtatggaaggttccgttaacggtcacgagttcg
aaatcgaaggtgaaggtgaaggtcgtccgtacgaaggtacccagaccgctaaactgaaagttaccaaaggtggtccgctgccgttcgcttgggacatcct
gtccccgcagttccagtacggttccaaagcttacgttaaacacccggctgacatcccggactacctgaaactgtccttcccggaaggtttcaaatgggaa
cgtgttatgaacttcgaagacggtggtgttgttaccgttacccaggactcctccctgcaagacggtgagttcatctacaaagttaaactgcgtggtacca
acttcccgtccgacggtccggttatgcagaaaaaaaccatgggttgggaagcttccaccgaacgtatgtacccggaagacggtgctctgaaaggtgaaat
caaaatgcgtctgaaactgaaagacggtggtcactacgacgctgaagttaaaaccacctacatggctaaaaaaccggttcagctgccgggtgcttacaaa
accgacatcaaactggacatcacctcccacaacgaagactacaccatcgttgaacagtacgaacgtgctgaaggtcgtcactccaccggtgctGCTGCAA
ACGACGAAAACTACGCTTTAGTAGCTtaataatactagagccaggcatcaaataaaacgaaaggctcagtcgaaagactgggcctttcgttttatctgtt
gtttgtcggtgaacgctctctactagagtcacactggctcaccttcgggtgggcctttctgcgtttata
Primers to order: Outer FP: gaattcgcggccgcttctagag aaagaggagaaatactagatggct (55C,24bp,38%GC) Total: Inner RP: CTACTAAAGCGTAGTTTTCGTCGTTTGCAGCagcaccggtggagtga (56C,16bp,63%GC) Total:47bp Primers that are the same as GFP: Inner FP: CGACGAAAACTACGCTTTAGTAGCTtaataatactagagccaggcatca (55C,24bp,38%GC) Total:49bp Outer RP: ctgcagcggccgctactagta tataaacgcagaaaggccca (56C,20bp,45%GC) Total:41bp Overlapping region: 55C,23bp,43%GC
YFP
- [http://partsregistry.org/wiki/index.php/Part:BBa_E0032 YFP with LVA tag]
CFP
- [http://partsregistry.org/wiki/index.php/Part:BBa_E0022 CFP with LVA tag]
Measurements
- [http://www.ncbi.nlm.nih.gov/pmc/articles/PMC106306/?tool=pubmed one to follow] unless there's a better one KULeuven08 followed it.
- Paper found by LiFeng.