IGEM:Stanford/2009/Project Homeostasis/Primer: Difference between revisions

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	Home	Team	Project	Parts	Notebook	Archives

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Project

Research Proposal

Systems Overview

Cloning Plan

Sequences & Primers

Anti-Inflammation

Anti-Immunosuppression

Protocols

Modeling

Overview

Notebook

Results

Future Work

Archived Work

Anti-Immunosuppression Device

Finalized Primers

Primers

[OPTIONAL]: Primers to use GTT TCT TC instead of TATA in front? I.e.

GTTTCTTCgaattcgcggccgcttctagATGGTTTGCTTTTTTTTACCTGCCA

Instead of:

TATA gaattcgcggccgcttctagATGGTTTGCTTTTTTTTACCTGCCA

HlyC (from E. coli str. CFT073)

Forward: TATAgaattcgcggccgcttctagagTTGGTTTGCTTTTTTTTACCTGCC

Reverse: TATA tactagtagcggccgctgcagTTAACCTGTTAATGAAAAATTGAAATC

untire sequence: http://www.ncbi.nlm.nih.gov/nuccore/NC_004431.1/?from=3418210&to=3418884&report=fasta

Added 7/1/09 - Mark

HlyC (from E. coli str. CFT073)

Forward: TATAgaattcgcggccgcttctagagTTGGTTTGCTTTTTTTTACCTGCCACC

Reverse: TATAtactagtagcggccgctgcagCTCTTAACCTGTTAATG

HlyB (from E. coli str. CFT073)

Forward: TATAgaattcgcggccgcttctagagGGAGTCATAATGGATTCTTGTC

Reverse: TATAtactagtagcggccgctgcagGTTAGTCTGACTGTAACTG

HlyD (from E. coli str. CFT073)

Forward: TATAgaattcgcggccgcttctagATGAAAACATGGTTAATGGGGTTCAGCG

Reverse: TATAtactagtagcggccgctgcagCTCTGAGACTTAACGCTCATG

TolC (from E. coli str. CFT073)

Forward: TATAgaattcgcggccgcttctagagGCAAATGAAGAAATTGCTCC

Reverse: TATAtactagtagcggccgctgcagCGTCATCAGTTACGGAAAGGG

Added 7/1/09 - Ming

HlyB (from CFT073)

"Forward": TATAgaattcgcggccgcttctag ATGGATTCTTGTCATAAAATTG

"Reverse": TATAtactagtagcggccgctgcag TTAGTCTGACTGTAACTG

HlyD (from CFT073)

"Forward":TATA gaattcgcggccgcttctag ATGAAAACATGGTTAATG

"Reverse": TATAtactagtagcggccgctgcag TTAACGCTCATGTAAACTTTC

TolC

"Forward": TATAgaattcgcggccgcttctagATGAAGAAATTGCTCCCCATTC

"Reverse": TATAtactagtagcggccgctgcagTCAGTTACGGAAAGGGTTATG'

HlyA

complete genome: [1]

Added 6/28/09 - Robert, Ariana

IL-6 fusion gene

"Forward": TATAGAATTCGCGGCCGCTTCTAGAGGGATGCATCACCGGTTCCGC

"Reverse": ATATTACTAGTAGCGGCCGCTGCAGGGATGCATTCATTTGACGAAGAG

New IL-6 fusion gene (Joe 7-5)

"Forward: TATAgaattcgcggccgcttctagATGCCAACAATAACCACTGC

"Reverse": TATAtactagtagcggccgctgcagTTATGCTGATGCTGTCAAAG

Added 7/1/09 - Anusuya

TrpR (E. coli str. IAI1)

Forward: TATAgaattcgcggccgcttctagATGGCCCAACAATCACCCTATTCAGC

Reverse: 'TATAtactagtagcggccgctgcagTCAATCGCTTTTCAGCAACACCTCTTCCAGCCACTGGC

Trp Promoter and Operator (E. coli: no specified strain )

Forward: TATAgaattcgcggccgcttctagagCTCAAGGCGCACTCCCGTTCTG

Reverse: TATAtactagtagcggccgctgcagTCGATACCCTTTTTACGTGAACTTGCGTACTAG

Protease-binding sequence?

Gene Sequences

trp-operon (promoter-operator) sequence

5' CTCAAGGCGCACTCCCGTTCTGGATAATGTTTTTTGCGCCGACATCATAACGGTTCTGGCAAATATTCTGAAATGACCTGTTGACAATTAATCATCGAACTAGTTAACTAGTACGCAAGTTCACGTAAAAAGGGTATCCA 3'

(Nucleotide Sequence of the Promoter- Operator Region of the Tryptophan Operon of Escherichia coli By G. N. BENNETT, M. E. SCHW~,INGRUB~.R~, K. D. BROW~: C. SQUIRES § AND C. ¥'ANOFSKY )

HylB

ATGATCATCAATAAATTTTCCCTTAAATGGATGTTGGCTATTGCCGTCGCCATCCCTGCGATAGCACTGTTGTTTGTGGCTTTCACCAGTCTAAACACCATGTCAGTGATGCAAGCGCAGTCCAACAGCTTGTATGCCAACACGGCTGCACCAATGCGTGCCATGGCTGAAGCAACCTCACGTATTCCTCGGATGCGTGTCGGTATCGATATGATGCTACTGCAAGAAACGGCGCTCAAAGATGCGAAAGGGGTCCTCAAACGAGTCGAAGAGGCAAGAACCGAAGATATCCCAGAAATGCGTCAAGCAATGCAAGTTGCGGTTGATTCTCAGGTTAATCCGGAACTCAAAGAGCAGGCACGCAAACTTCAAGCTCGTTTTGAACAAATGGTACGTGAAGAGTTAGAGCCTATGCTGCAAGCCTTCGCCAATAACGATATGACCACGGCACAAAACATTTACCGCGATAAATACGCGCCGACCTATGGTGAAATGCGTAAACAAGCCAACCAGATCCTCGATACGCTTTTGCAGCAAGCGGAGCAGCAAAACCATGCCAGTGTGGAAAGCTTCGAAGCAGGACGCACCAAGCAAATGGTGATCATTGCAGCAGGCTTGATCATTTCATTCATCACTTCACTGGTTATCATAACGAACTTACGTAGCCGAGTGGCTTACCTGAAAGATCGTATGAGTTCTGCGGCGGCGAATCTTTCACTGCGTACTCGATTGGAGTTGGATGGTAACGATGAACTGTGTGACATCGGTAAAAGCTTCAATGCGTTCATTGATAAAGTGCATCACTCGATTGAAGAAGTGGCAGAAAACTCAAAAGAGCTGGCGACGATGGCCTCTAGTGTGTCGCAGCGCGCGCACATGACGCAATCTAACTGTGCTTCGCAGCGAGATAGAACAGTGCAAGTTGCGACGGCGATTCATGAGCTTGGTGCCACCGTATCCGAAATCGCTTCCAATGCGGCCATGGCTGCCGATGTCGCGAAGCAAGCGACGCTGCATTCTGGTGAAGGGAAAAAAGTGGTAGGCGAAGTGCAAAATCGGATCCAAACACTGGTCAATGAACTCGATAATGCCACTCAAGTTGTCTCATCACTGGCGACCCAAATTAACGGTATTAGCTCAACACTTGATACCATTCGCAGTATTTCTGAGCAAACGAACCTATTGGCGCTCAACGCTGCGATTGAAGCTGCGCGAGCGGGTGAACAAGGTCGTGGTTTTGCGGTGGTGGCGGATGAAGTTCGCACATTAGCAAGTCGTTCAGCGGCATCGACGGAAGAGATCCAGCAAGTCATTAATCGCCTTCAAACGGAGTCAACTCGCGCAGTAGAAGCAATGGAAAAAGGTCGCTCGCAAAGTGATGTGGTGGTTGAGTTTTCCGCTAAAGCGAACCAATCTCTCACAGAGATCAACAGCCAAATTGATCAGATTAATGATCAAAATATTCAAGTTGCGACCGCGACAGAGGAACAATCAACCGTGGTGGAAGACATTAATCGCAACGTTGAAGACATCAACCAACTGACGACAGAAACCTCGCATGTTGCGGATGAGTTAAGCCGAGCCAGTGCAAGCTTGCAACGTCTCTCTTCGCAACTGGATAAACTGGTGGGCAGTTTTGAACTTTAA http://www.ncbi.nlm.nih.gov/nuccore/NC_012583.1/?from=274978&to=276624&report=fasta NCBI Reference Sequence: NC_012583.1 from Vibrio cholerae O395 chromosome II, complete genome

HylD

ATGGCTGCGTTCGGACAAGACGTCTTCAGGCGCTTCGGCACGGTCACCGAACGCTTCTCCGGCACCGGCGGCAAACTCTTCGGCGGCCTGCTCGCGCGCCTGTCGAGCGTGGAGCGCGACGACAGCCTGGACTGGGCGAGCGACGCCGACTGGGCGCGCCTGCAGCAGGAGCCGCTGCGCGCGCGCGCGGTACTCAAGGTCAGCGCGATCGCGCTGCTGGTGCTGCTCGGCTGGGCCGCGGTGGCGCATGTGGACGAGGTGACCAAGGGCGACGGCAAGGTGATCCCGTCCAGCCAGCTGCAGATCATCCAGTCGGTCGATGGCGGCGTGGTGGAGTCGATCGCGGTGCGCGAGGGCGAGGTGGTCGAGGCCGGGCAGCTGCTGCTCAAGGTCGATCCCACCCGTTTCATGTCCTCGCTGCTGGAAAACCGCGCCGAGTTCCTCGCGCTGGAAATCAAGCGCCGGCGGCTCGAGGCGCTGACCCAGGGGACGCCGTTCTCGGTGCCGGCCGAGCTGGAGAAGGAGGCGGCGGACATCGCCGCCCACGAGCGCCGGCTGTATGAATCCAGCCGCGCCGAGATGGACGCGCAGCAGTCGATCGCGCGCGACCAGCTGCGCCAGCGCGAGCAGGAACTCAACGAGGTGCGCGCCCGCAGTGAGAACGCCGCCCGCGCCTACGAACTGGTGCAGCAGGAACTCAAGGTCACCAAGCCGCTCGCGGCCTCCGGCGCGGTATCCGAGGTCGAGCTGCTGCGTCTGGAGCGCGACGTGTCGCGCTTGCGCGGCGAGCGCGAGCAGAGCGCGGCGCAGATTTCGCGCATCCAGTCGGCCATCGCCGAATCCACGCGCAAGATCCAGGAGATCGAGCTGACGGTGCGCAACCAGCTGCGCAACGAGCTTTCGGACACCATGGCGCGGCTGGGCAGCCTGACCCAGGGCAGCCGCATGCTGGCCGACCGCGTCAAGCACGCCGAAATCCGCGCGCCGATGCGCGGCACGGTCAAGCGCATCCTCGTCAACACCGTGGGTGGCGTGGTCCAGCCGGGCAAGGAAGTGGCCGAGATCGTGCCGCTCGACGACGCGCTGATCCTCGAAGCGCAGGTCAAGCCGAGCGACATCGCCTTCCTGCGCCCGGGTCAGTCGGCGCTGGTCAAGTTCTCCGCGTACGACTTCTCGATCTACGGCGGCCTGGAGGCGGTCGTGGACCAGATTTCCGCCGACACCGTGGTCGATGCAAAGGGTTCCGCGTTCTACGTGGTGCGGGTGCGCACGCTGCATTCCTCTCTGGGTGAAAACCTGCCGATCATTCCGGGCATGGTGGCCGAAGTGGACATCCTGACCGGCAAGAAGACCATCCTCTCCTACCTGCTGAAGCCCGTGCTGCGGGCCAAGGCCAACGCACTGTCCGAACGATGA http://www.ncbi.nlm.nih.gov/nuccore/NC_008702.1/?from=674498&to=675913&report=fasta NCBI Reference Sequence: NC_008702.1 from Azoarcus sp. BH72, complete genome

TolC

ATGAAGAAATTGCTCCCCATTCTTATCGGCCTGAGCCTTTCTGGGTTCAGTTCGTTGAGCCAGGCCGAGAACCTGATGCAAGTTTATCAGCAAGCACGCCTTAGTAACCCGGAATTGCGTAAGTCTGCCGCCGATCGTGATGCTGCCTTTGAAAAAATTAATGAAGCGCGCAGTCCATTACTGCCACAGCTAGGTTTAGGTGCAGATTACACCTATAGCAACGGCTACCGCGACGCGAACGGCATCAACTCTAACGCGACCAGTGCGTCCTTGCAGTTAACTCAATCCATTTTTGATATGTCGAAATGGCGTGCGTTAACGCTGCAGGAAAAAGCAGCAGGGATTCAGGACGTCACGTATCAGACCGATCAGCAAACCTTGATCCTCAACACCGCGACCGCTTATTTCAACGTGTTGAATGCTATTGACGTTCTTTCCTATACACAGGCACAAAAAGAAGCGATCTACCGTCAATTAGATCAAACCACCCAACGTTTTAACGTGGGCCTGGTAGCGATCACCGACGTGCAGAACGCCCGCGCACAGTACGATACCGTGCTGGCGAACGAAGTGACCGCACGTAATAACCTTGATAACGCGGTAGAGCAGCTGCGCCAGATCACCGGTAACTACTATCCGGAACTGGCTGCGCTGAATGTCGAAAACTTTAAAACCGACAAACCACAGCCGGTTAACGCGCTGCTGAAAGAAGCCGAAAAACGCAACCTGTCGCTGTTACAGGCACGCTTGAGCCAGGACCTGGCGCGCGAGCAAATTCGCCAGGCGCAGGATGGTCACTTACCGACTCTGGATTTAACGGCTTCTACCGGGATTTCTGACACCTCTTATAGCGGTTCGAAAACCCGTGGTGCCGCTGGTACCCAGTATGACGATAGCAATATGGGCCAGAACAAAGTTGGCCTGAGCTTCTCGCTGCCGATTTATCAGGGCGGAATGGTTAACTCGCAGGTGAAACAGGCACAGTACAACTTTGTCGGTGCCAGCGAGCAACTGGAAAGTGCCCATCGTAGCGTCGTGCAGACCGTGCGTTCCTCCTTCAACAACATTAATGCATCTATCAGTAGCATTAACGCCTACAAACAAGCCGTAGTTTCCGCTCAAAGCTCATTAGACGCGATGGAAGCGGGCTACTCGGTCGGTACGCGTACCATTGTTGATGTGTTGGATGCGACCACCACGTTGTACAACGCCAAGCAAGAGCTGGCGAATGCGCGTTATAACTACCTGATTAATCAGCTGAATATTAAGTCAGCTCTGGGTACGTTGAACGAGCAGGATCTGCTGGCACTGAACAATGCGCTGAGCAAACCGGTTTCCACTAATCCGGAAAACGTTGCACCGCAAACGCCGGAACAGAATGCTATTGCTGATGGTTATGCGCCTGATAGCCCGGCACCAGTCGTTCAGCAAACATCCGCACGCACTACCACCAGTAACGGTCATAACCCTTTCCGTAACTGA http://www.ncbi.nlm.nih.gov/nuccore/NC_000913.2/?from=3176137&to=3177618&report=fasta NCBI Reference Sequence: NC_000913.2 from Escherichia coli str. K-12 substr. MG1655, complete genome

Protease-binding sequence?

Anti-Inflammatory Device

Primers:

SoxR (Escherichia coli E24377A)

Forward: TATAgaattcgcggccgcttctagATGGAAAAGAAATTACCCCGC

Reverse: TATAtactagtagcggccgctgcagTTAGTTTTGTTCATCTTCCAGCAAGCGTG

complete gene sequence: [2]

SoxS promoter (Joe 7-5)

Forward: TATAgaattcgcggccgcttctagagAATCGCTTTACCTCAAGTTAACTTG

Reverse: TATAtactagtagcggccgctgcagCAGTTCGTTAATTCATCTGTTGGG

Added 7/1/09 - Anusuya

CrtB (Erwinia Uredovora)

Forward: TATAgaattcgcggccgcttctagATGGCAGTTGGCTCGAAAAG

Reverse: TATAtactagtagcggccgctgcagCTAGAGCGGGCGCTGCCAG

CrtE (Erwinia Uredovora)

Forward: TATAgaattcgcggccgcttctagATGACGGTCTGCGCAAAAAAAC

Reverse: TATAtactagtagcggccgctgcagTTAACTGACGGCAGCGAGTTTTTTG

CrtI (Erwinia Uredovora)

Forward: TATAgaattcgcggccgcttctag ATGAAACCAACTACGGTAATTGGTGC

Reverse: TATAtactagtagcggccgctgcag TGATTGAGTAACGACGG

CrtY (Erwinia Uredovora)

Forward: TATAgaattcgcggccgcttctag ATGCAACCGCATTATGATCTG

Reverse: TATAtactagtagcggccgctgcag TTAACGATGAGTCGTCATAATGGC

Soxs: primer dimers

Blh (uncultured marine bacterium 66A03)

Forward: TATAgaattcgcggccgcttctagATGGGTCTGATGCTGATTG

Reverse: TATAtactagtagcggccgctgcagTCAGTTTTTGATTTTGATACGGG

RalDH(II) (Rattus Norvegicus*)

Forward: TATAgaattcgcggccgcttctagATGCCCGGCGAGGTGAAGGC

Reverse: TATAtactagtagcggccgctgcagTTAGGAGTTCTTCTGGGGGATC

Originally from Rattus Norvegicus but sequence was modified

HlyR: Since HlyR is not a protein. This sequence is sequenced as a chunk with HlyA,HlyB, HlyC. Thus only one primer is needed. primer: TATAtactagtagcggccgctgcagGAATTCCAAGCGAAGTCC

check paper for RalDH(II) http://www.jbc.org/cgi/content/abstract/271/27/16288

Gene sequences: SoxR ATGGAAAAGAAATTACCCCGCATTAAAGCGCTGCTAACCCCCGGCGAAGTGGCGAAACGCAGCGGTGTGGCGGTATCGGCGCTGCATTTCTATGAAAGTAAAGGGTTGATTACCAGTATCCGTAACAGCGGCAATCAGCGGCGATATAAACGTGATGTGTTGCGATATGTTGCAATTATCAAAATTGCTCAGCGTATTGGCATTCCGCTGGCGACCATTGGTGAAGCGTTTGGCGTGTTGCCCGAAGGGCATACGTTAAGTGCGAAAGAGTGGAAACAGCTTTCGTCCCAATGGCGAGAAGAGTTGGATCGGCGCATTCATACCTTAGTGGCGCTGCGTGACGAACTGGACGGATGTATTGGTTGTGGCTGCCTTTCGCGCAGTGATTGCCCGTTGCGTAACCCGGGCGACCGCTTAGGAGAAGAAGGTACCGGCGCACGCTTGCTGGAAGATGAACAAAACTAA http://www.ncbi.nlm.nih.gov/nuccore/NC_009801.1/?from=4616553&to=4617017&report=fasta (NCBI Reference Sequence: NC_009801.1 from Escherichia coli E24377A, complete genome)

SoxS

AATCGCTTTACCTCAAGTTAACTTGAGGAATTATACTCCCCAACAGATGAATTAACGAACTG

Crt genes GenBank accession no. D90087—should we do as the entire cluster (in this case how should we design primers) or in parts?

Blh ATGGGTCTGATGCTGATTGATTGGTGTGCACTGGCTCTGGTTGTTTTCATTGGCCTGCCGCACGGCGCGCTGGATGCTGCCATTTCTTTTTCTATGATCTCTTCTGCAAAACGCATTGCTCGTCTGGCTGGTATTCTGCTGATCTATCTGCTGCTGGCGACCGCGTTCTTCCTGATCTGGTATCAGCTGCCAGCGTTTAGCCTGCTGATCTTCCTGCTGATCTCCATTATCCACTTTGGTATGGCAGACTTCAACGCGTCCCCAAGCAAACTGAAATGGCCGCATATCATCGCCCACGGCGGTGTTGTTACTGTTTGGCTGCCGCTGATCCAGAAAAACGAAGTAACTAAACTGTTTAGCATCCTGACTAACGGTCCGACTCCGATCCTGTGGGACATCCTGCTGATTTTCTTCCTGTGTTGGTCTATTGGCGTGTGTCTGCACACGTACGAAACCCTGCGCTCTAAACATTACAACATCGCCTTTGAACTGATCGGTCTGATTTTCCTGGCGTGGTATGCGCCGCCTCTGGTTACGTTTGCCACTTACTTCTGCTTCATTCATTCCCGTCGCCACTTCTCCTTTGTGTGGAAGCAGCTGCAACACATGTCTTCCAAAAAGATGATGATTGGCAGCGCGATTATCCTGTCCTGTACCTCTTGGCTGATCGGCGGTGGTATCTATTTCTTCCTGAACTCCAAAATGATCGCCTCTGAGGCTGCGCTGCAGACTGTGTTCATCGGTCTGGCGGCACTGACCGTGCCGCACATGATTCTGATCGACTTCATCTTCCGTCCGCACTCTTCCCGTATCAAAATCAAAAACTGA

Kim et al. In vitro characterization of a recombinant Blh protein from an uncultured marine bacterium as a beta-carotene 15,15'-dioxygenase. J Biol Chem. 2009 Jun 5;284(23):15781-93. (supplementary material; 15,15'-Dioxygenase, product of Blh gene, codon-optimized and expressed in E. coli, forms a 64k protein of two 32 kDa dimers)

RALDH

HlyR:

sequence: GAATTCCAAGCGAAGTCCATCCCCCTCCCTCTTGATTACAAGGGTGATAATTATTATTCGCATTTGTGTG GTAATGGGATAGAAAGGAATGGATAGAAAAAGAACAAAATTAGTATAGCAATAGATATGCCCACTGCATT GAATACTTACAGGGCATTATTTTATTATGTTTAAATTGAAGTGGTCTCTGGTTTGATTTATTTGTTATTC AAGGGGGCTGTTTGGAGATCGGAAAATTCTGTACGTTAAGTGTATTATTTAACCAGTTTCGATGCGTAAC AGATTGATTTTGCTCAGCGGTTATCGCTTTTAAGTTGTTGCTCTTGGCTATCGCGTTTAGGTTATCCGAT TAAAGTCAAATTTCCTGAAAATGCTGTATAGCGCGGGAGTGCACCTTATAGCTGTAGGTAAGTATGTTCA AAAAATAGTCTTGCCGTACAATAATTTTCCATATCCAAACTCACTCCTTCAAGATTCTGGTCCCGGTTTA CGGGTAGTTTCCGGAAGGGCGGTAGCATGCTGATTCAAACTGCAAGATGAAACATTGTCGGAGTTGGATG GAATTAAGTCATGGCTATAGCATTTGGGCGTGCATAACAAAATTGGTCCTCATATTTTAGAGTATGATTG CATATTCACTAATATTTTTACTTTCTGATGCGTGGTGGCATCATGCTTTATGAGATAAACAATCCTGG

blh

ATGGGTCTGATGCTGATTGATTGGTGTGCACTGGCTCTGGTTGTTTTCATTGGCCTGCCGCACGGCGCG CTGGATGCTGCCATTTCTTTTTCTATGATCTCTTCTGCAAAACGCATTGCTCGTCTGGCTGGTATTCTG CTGATCTATCTGCTGCTGGCGACCGCGTTCTTCCTGATCTGGTATCAGCTGCCAGCGTTTAGCCTGCTG ATCTTCCTGCTGATCTCCATTATCCACTTTGGTATGGCAGACTTCAACGCGTCCCCAAGCAAACTGAAA TGGCCGCATATCATCGCCCACGGCGGTGTTGTTACTGTTTGGCTGCCGCTGATCCAGAAAAACGAAGTA ACTAAACTGTTTAGCATCCTGACTAACGGTCCGACTCCGATCCTGTGGGACATCCTGCTGATTTTCTTC CTGTGTTGGTCTATTGGCGTGTGTCTGCACACGTACGAAACCCTGCGCTCTAAACATTACAACATCGCC TTTGAACTGATCGGTCTGATTTTCCTGGCGTGGTATGCGCCGCCTCTGGTTACGTTTGCCACTTACTTC TGCTTCATTCATTCCCGTCGCCACTTCTCCTTTGTGTGGAAGCAGCTGCAACACATGTCTTCCAAAAAG ATGATGATTGGCAGCGCGATTATCCTGTCCTGTACCTCTTGGCTGATCGGCGGTGGTATCTATTTCTTC CTGAACTCCAAAATGATCGCCTCTGAGGCTGCGCTACAAACTGTGTTCATCGGTCTGGCGGCACTGACC GTGCCGCACATGATTCTGATCGACTTCATCTTCCGTCCGCACTCTTCCCGTATCAAAATCAAAAACTGA

RALDH II

ATGCCCGGCGAGGTGAAGGCCGACCCCGCCGCGCTCATGGCCTCGCTGCAGCTCCTGCCG TCACCCACGCCCAACCTCGAGATCAAGTACACCAAGATCTTTATTAACAATGAATGGCAG AACTCAGAGAGTGGGAGAGTGTTCCCTGTCTGTAATCCAGCCACAGGAGAGCAAGTGTGT GAAGTTCAAGAAGCAGACAAGGTGGATATAGACAAGGCAGTGCAGGCGGCCCGCCTGGCC TTCTCTCTTGGTTCTGTGTGGAGAAGGATGGACGCTTCTGAAAGAGGACGTCTGCTGGAC AAGCTTGCAGACTTGGTGGAACGGGACAGGGCAACTCTTGCAACTATGGAATCACTAAAT GGCGGTAAGCCATTCCTGCAAGCTTTTTACATCGATTTGCAGGGAGTCATCAAAACCCTG AGGTATTATGCAGGCTGGGCTGATAAAATTCACGGAATGACCATTCCTGTAGATGGAGAC TATTTTACCTTCACGAGACATGAGCCCATCGGAGTGTGTGGACAGATTATCCCGTGGAAT TTCCCCCTGCTGATGTTCACCTGGAAAATTGCTCCCGCACTGTGCTGTGGTAACACTGTG GTCATCAAGCCTGCAGAGCAGACACCACTCAGTGCACTGTATATGGGAGCCCTCATCAAG GAGGCTGGCTTTCCACCCGGGGTCGTCAACATTCTGCCAGGGTATGGGCCAACAGCAGGG GCAGCAATAGCGTCTCACATTGGCATAGACAAGATTGCATTCACAGGCTCTACTGAGGTT GGAAAGCTTATTCAAGAAGCAGCTGGAAGAAGTAATTTGAAGAGAGTAACTCTGGAACTT GGAGGGAAAAGCCCTAATATTATTTTTGCAGATGCTGACTTGGACTACGCTGTGGAGCAG GCTCACCAGGGTGTGTTCTTCAACCAAGGCCAGTGCTGCACTGCGGGGTCTCGAATCTTT GTGGAGGAGTCCATCTATGAGGAATTTGTGAAAAGAAGTGTGGAAAGAGCCAAGAGGCGC ATTGTGGGGAGCCCCTTTGATCCCACTACTGAGCAGGGTCCTCAGATTGATAAGAAGCAA TACAACAAAATCCTGGAGCTCATCCAGAGCGGTGTGGCCGAGGGCGCCAAGCTCGAGTGT GGAGGCAAAGGACTGGGAAGGAAGGGCTTCTTCATTGAGCCTACCGTGTTCTCCAACGTT ACAGATGACATGCGGATCGCCAAGGAGGAGATCTTTGGTCCTGTCCAGGAAATTTTGAGG TTTAAGACTATGGATGAAGTTATAGAAAGAGCCAATAACTCAGACTTCGGGCTTGTAGCG GCTGTCTTCACTAATGACATCAACAAGGCTCTCATGGTGTCCTCTGCAATGCAAGCTGGG ACAGTTTGGATCAATTGTTACAACGCCTTAAATGCTCAGAGCCCCTTCGGGGGGTTCAAG ATGTCTGGAAATGGGAGAGAAATGGGTGAGTTTGGCTTACGGGAGTACTCAGAAGTAAAG ACCGTGACGGTGAAGATCCCCCAGAAGAACTCCTAA

Comments

For gene sequencing, I would suggest genedesigner
For primer design and selection I would suggest APE and oligo 6.87

IGEM:Stanford/2009/Project Homeostasis/Primer: Difference between revisions

Latest revision as of 10:53, 21 August 2009

Anti-Immunosuppression Device

Anti-Inflammatory Device

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@@ Line 1: / Line 1: @@
+<html>
+<style>
+/*----- General */
+body {
+  background: black;
+  color: black;
+}
+</style>
+</html>
+&nbsp; {{Template:Stanford_Main_Menu_0809}}
+{{Template:ProjectMenu}}
 ==Anti-Immunosuppression Device ==
+*[http://openwetware.org/wiki/IGEM:Stanford/2009/Project_Homeostasis/Primer/Primersdoc  Finalized Primers]
 Primers
-'''HlyB (from E. coli str. CFT073)'''
+[OPTIONAL]: Primers to use GTT TCT TC instead of TATA in front?  I.e.
+GTTTCTTCgaattcgcggccgcttctagATGGTTTGCTTTTTTTTACCTGCCA
+Instead of:
+TATA    gaattcgcggccgcttctagATGGTTTGCTTTTTTTTACCTGCCA
+*'''HlyC (from E. coli str. CFT073)'''
+Forward: TATAgaattcgcggccgcttctagagTTGGTTTGCTTTTTTTTACCTGCC
+Reverse: '''''TATA tactagtagcggccgctgcagTTAACCTGTTAATGAAAAATTGAAATC'''''
+untire sequence: http://www.ncbi.nlm.nih.gov/nuccore/NC_004431.1/?from=3418210&to=3418884&report=fasta
+Added 7/1/09 - ''Mark''
+*'''HlyC (from E. coli str. CFT073)'''
+''Forward'': TATAgaattcgcggccgcttctagagTTGGTTTGCTTTTTTTTACCTGCCACC
+''Reverse'': '''''TATAtactagtagcggccgctgcagCTCTTAACCTGTTAATG'''''
+*'''HlyB (from E. coli str. CFT073)'''
 ''Forward'': TATAgaattcgcggccgcttctagagGGAGTCATAATGGATTCTTGTC
-''Reverse'': TATAtactagtagcggccgctgcagGTTAGTCTGACTGTAACTG
+''Reverse'': '''''TATAtactagtagcggccgctgcagGTTAGTCTGACTGTAACTG'''''
+*'''HlyD (from E. coli str. CFT073)'''
+''Forward'': TATAgaattcgcggccgcttctagATGAAAACATGGTTAATGGGGTTCAGCG
+''Reverse'': '''''TATAtactagtagcggccgctgcagCTCTGAGACTTAACGCTCATG'''''
+*'''TolC (from E. coli str. CFT073)'''
+''Forward'': TATAgaattcgcggccgcttctagagGCAAATGAAGAAATTGCTCC
+''Reverse'': '''''TATAtactagtagcggccgctgcagCGTCATCAGTTACGGAAAGGG'''''
+Added 7/1/09 - ''Ming''
-'''HlyD (from E. coli str. CFT073)'''
+*'''HlyB (from CFT073)'''
-''Forward'': TATAgaattcgcggccgcttctagATGAAAACATGGTTAATGGGGTTCAGCG
-''Reverse'': TATAtactagtagcggccgctgcagCTCTGAGACTTAACGCTCATG
+"Forward": TATAgaattcgcggccgcttctag ATGGATTCTTGTCATAAAATTG
-'''TolC (from E. coli str. CFT073)'''
+"Reverse": '''''TATAtactagtagcggccgctgcag TTAGTCTGACTGTAACTG'''''
-''Forward'': TATAgaattcgcggccgcttctagagGCAAATGAAGAAATTGCTCC
-''Reverse'': TATAtactagtagcggccgctgcagCGTCATCAGTTACGGAAAGGG
+*'''HlyD (from CFT073)'''
+"Forward":TATA gaattcgcggccgcttctag ATGAAAACATGGTTAATG
+"Reverse": '''''TATAtactagtagcggccgctgcag TTAACGCTCATGTAAACTTTC'''''
-'''HlyB (from CFT073)'''
+*'''TolC'''
-TATAgaattcgcggccgcttctag ATGGATTCTTGTCATAAAATTG
+"Forward": TATAgaattcgcggccgcttctagATGAAGAAATTGCTCCCCATTC
-TATAtactagtagcggccgctgcag TTAGTCTGACTGTAACTG
+"Reverse": '''''TATAtactagtagcggccgctgcagTCAGTTACGGAAAGGGTTATG''''''
-'''HlyD (from CFT073)'''
+*'''HlyA'''
+complete genome: [http://www.ncbi.nlm.nih.gov/nuccore/NC_004431.1/?from=3418896&to=3421970&report=fasta]
-TATA gaattcgcggccgcttctag ATGAAAACATGGTTAATG
-TATAtactagtagcggccgctgcag TTAACGCTCATGTAAACTTTC
+Added 6/28/09 - ''Robert, Ariana''
-'''TolC'''
+*'''IL-6 fusion gene'''
+"Forward": TATAGAATTCGCGGCCGCTTCTAGAGGGATGCATCACCGGTTCCGC
-TATAgaattcgcggccgcttctagATGAAGAAATTGCTCCCCATTC
+"Reverse": '''''ATATTACTAGTAGCGGCCGCTGCAGGGATGCATTCATTTGACGAAGAG'''''
-TATAtactagtagcggccgctgcagTCAGTTACGGAAAGGGTTATG
+*'''New IL-6 fusion gene (Joe 7-5)'''
+"Forward: TATAgaattcgcggccgcttctagATGCCAACAATAACCACTGC
+"Reverse": TATAtactagtagcggccgctgcagTTATGCTGATGCTGTCAAAG
-'''IL-6 fusion gene'''
-TATAGAATTCGCGGCCGCTTCTAGAGGGATGCATCACCGGTTCCGC
-ATATTACTAGTAGCGGCCGCTGCAGGGATGCATTCATTTGACGAAGAG
-Protease-binding sequence?
+Added 7/1/09 - ''Anusuya''
+*'''TrpR''' ''(E. coli str. IAI1)''
+''Forward'': TATAgaattcgcggccgcttctagATGGCCCAACAATCACCCTATTCAGC
+''Reverse'': ''''''TATAtactagtagcggccgctgcagTCAATCGCTTTTCAGCAACACCTCTTCCAGCCACTGGC'''''
+*'''Trp Promoter and Operator''' ''(E. coli: no specified strain )''
+''Forward:'' TATAgaattcgcggccgcttctagagCTCAAGGCGCACTCCCGTTCTG
+''Reverse:'' TATAtactagtagcggccgctgcagTCGATACCCTTTTTACGTGAACTTGCGTACTAG
+*Protease-binding sequence?
 Gene Sequences
-'''HylB'''
+*'''trp-operon (promoter-operator) sequence'''
+'   CTCAAGGCGCACTCCCGTTCTGGATAATGTTTTTTGCGCCGACATCATAACGGTTCTGGCAAATATTCTGAAATGACCTGTTGACAATTAATCATCGAACTAGTTAACTAGTACGCAAGTTCACGTAAAAAGGGTATCCA  3'
+(Nucleotide Sequence of the Promoter- Operator Region of the Tryptophan Operon of Escherichia coli  By G. N. BENNETT, M. E. SCHW~,INGRUB~.R~, K. D. BROW~:
+C. SQUIRES § AND C. ¥'ANOFSKY )
+*'''HylB'''
 ATGATCATCAATAAATTTTCCCTTAAATGGATGTTGGCTATTGCCGTCGCCATCCCTGCGATAGCACTGTTGTTTGTGGCTTTCACCAGTCTAAACACCATGTCAGTGATGCAAGCGCAGTCCAACAGCTTGTATGCCAACACGGCTGCACCAATGCGTGCCATGGCTGAAGCAACCTCACGTATTCCTCGGATGCGTGTCGGTATCGATATGATGCTACTGCAAGAAACGGCGCTCAAAGATGCGAAAGGGGTCCTCAAACGAGTCGAAGAGGCAAGAACCGAAGATATCCCAGAAATGCGTCAAGCAATGCAAGTTGCGGTTGATTCTCAGGTTAATCCGGAACTCAAAGAGCAGGCACGCAAACTTCAAGCTCGTTTTGAACAAATGGTACGTGAAGAGTTAGAGCCTATGCTGCAAGCCTTCGCCAATAACGATATGACCACGGCACAAAACATTTACCGCGATAAATACGCGCCGACCTATGGTGAAATGCGTAAACAAGCCAACCAGATCCTCGATACGCTTTTGCAGCAAGCGGAGCAGCAAAACCATGCCAGTGTGGAAAGCTTCGAAGCAGGACGCACCAAGCAAATGGTGATCATTGCAGCAGGCTTGATCATTTCATTCATCACTTCACTGGTTATCATAACGAACTTACGTAGCCGAGTGGCTTACCTGAAAGATCGTATGAGTTCTGCGGCGGCGAATCTTTCACTGCGTACTCGATTGGAGTTGGATGGTAACGATGAACTGTGTGACATCGGTAAAAGCTTCAATGCGTTCATTGATAAAGTGCATCACTCGATTGAAGAAGTGGCAGAAAACTCAAAAGAGCTGGCGACGATGGCCTCTAGTGTGTCGCAGCGCGCGCACATGACGCAATCTAACTGTGCTTCGCAGCGAGATAGAACAGTGCAAGTTGCGACGGCGATTCATGAGCTTGGTGCCACCGTATCCGAAATCGCTTCCAATGCGGCCATGGCTGCCGATGTCGCGAAGCAAGCGACGCTGCATTCTGGTGAAGGGAAAAAAGTGGTAGGCGAAGTGCAAAATCGGATCCAAACACTGGTCAATGAACTCGATAATGCCACTCAAGTTGTCTCATCACTGGCGACCCAAATTAACGGTATTAGCTCAACACTTGATACCATTCGCAGTATTTCTGAGCAAACGAACCTATTGGCGCTCAACGCTGCGATTGAAGCTGCGCGAGCGGGTGAACAAGGTCGTGGTTTTGCGGTGGTGGCGGATGAAGTTCGCACATTAGCAAGTCGTTCAGCGGCATCGACGGAAGAGATCCAGCAAGTCATTAATCGCCTTCAAACGGAGTCAACTCGCGCAGTAGAAGCAATGGAAAAAGGTCGCTCGCAAAGTGATGTGGTGGTTGAGTTTTCCGCTAAAGCGAACCAATCTCTCACAGAGATCAACAGCCAAATTGATCAGATTAATGATCAAAATATTCAAGTTGCGACCGCGACAGAGGAACAATCAACCGTGGTGGAAGACATTAATCGCAACGTTGAAGACATCAACCAACTGACGACAGAAACCTCGCATGTTGCGGATGAGTTAAGCCGAGCCAGTGCAAGCTTGCAACGTCTCTCTTCGCAACTGGATAAACTGGTGGGCAGTTTTGAACTTTAA
 http://www.ncbi.nlm.nih.gov/nuccore/NC_012583.1/?from=274978&to=276624&report=fasta NCBI Reference Sequence: NC_012583.1 from Vibrio cholerae O395 chromosome II, complete genome
 '''
-HylD'''
+*HylD'''
 ATGGCTGCGTTCGGACAAGACGTCTTCAGGCGCTTCGGCACGGTCACCGAACGCTTCTCCGGCACCGGCGGCAAACTCTTCGGCGGCCTGCTCGCGCGCCTGTCGAGCGTGGAGCGCGACGACAGCCTGGACTGGGCGAGCGACGCCGACTGGGCGCGCCTGCAGCAGGAGCCGCTGCGCGCGCGCGCGGTACTCAAGGTCAGCGCGATCGCGCTGCTGGTGCTGCTCGGCTGGGCCGCGGTGGCGCATGTGGACGAGGTGACCAAGGGCGACGGCAAGGTGATCCCGTCCAGCCAGCTGCAGATCATCCAGTCGGTCGATGGCGGCGTGGTGGAGTCGATCGCGGTGCGCGAGGGCGAGGTGGTCGAGGCCGGGCAGCTGCTGCTCAAGGTCGATCCCACCCGTTTCATGTCCTCGCTGCTGGAAAACCGCGCCGAGTTCCTCGCGCTGGAAATCAAGCGCCGGCGGCTCGAGGCGCTGACCCAGGGGACGCCGTTCTCGGTGCCGGCCGAGCTGGAGAAGGAGGCGGCGGACATCGCCGCCCACGAGCGCCGGCTGTATGAATCCAGCCGCGCCGAGATGGACGCGCAGCAGTCGATCGCGCGCGACCAGCTGCGCCAGCGCGAGCAGGAACTCAACGAGGTGCGCGCCCGCAGTGAGAACGCCGCCCGCGCCTACGAACTGGTGCAGCAGGAACTCAAGGTCACCAAGCCGCTCGCGGCCTCCGGCGCGGTATCCGAGGTCGAGCTGCTGCGTCTGGAGCGCGACGTGTCGCGCTTGCGCGGCGAGCGCGAGCAGAGCGCGGCGCAGATTTCGCGCATCCAGTCGGCCATCGCCGAATCCACGCGCAAGATCCAGGAGATCGAGCTGACGGTGCGCAACCAGCTGCGCAACGAGCTTTCGGACACCATGGCGCGGCTGGGCAGCCTGACCCAGGGCAGCCGCATGCTGGCCGACCGCGTCAAGCACGCCGAAATCCGCGCGCCGATGCGCGGCACGGTCAAGCGCATCCTCGTCAACACCGTGGGTGGCGTGGTCCAGCCGGGCAAGGAAGTGGCCGAGATCGTGCCGCTCGACGACGCGCTGATCCTCGAAGCGCAGGTCAAGCCGAGCGACATCGCCTTCCTGCGCCCGGGTCAGTCGGCGCTGGTCAAGTTCTCCGCGTACGACTTCTCGATCTACGGCGGCCTGGAGGCGGTCGTGGACCAGATTTCCGCCGACACCGTGGTCGATGCAAAGGGTTCCGCGTTCTACGTGGTGCGGGTGCGCACGCTGCATTCCTCTCTGGGTGAAAACCTGCCGATCATTCCGGGCATGGTGGCCGAAGTGGACATCCTGACCGGCAAGAAGACCATCCTCTCCTACCTGCTGAAGCCCGTGCTGCGGGCCAAGGCCAACGCACTGTCCGAACGATGA
 http://www.ncbi.nlm.nih.gov/nuccore/NC_008702.1/?from=674498&to=675913&report=fasta NCBI Reference Sequence: NC_008702.1 from Azoarcus sp. BH72, complete genome
-'''TolC'''
+*'''TolC'''
 ATGAAGAAATTGCTCCCCATTCTTATCGGCCTGAGCCTTTCTGGGTTCAGTTCGTTGAGCCAGGCCGAGAACCTGATGCAAGTTTATCAGCAAGCACGCCTTAGTAACCCGGAATTGCGTAAGTCTGCCGCCGATCGTGATGCTGCCTTTGAAAAAATTAATGAAGCGCGCAGTCCATTACTGCCACAGCTAGGTTTAGGTGCAGATTACACCTATAGCAACGGCTACCGCGACGCGAACGGCATCAACTCTAACGCGACCAGTGCGTCCTTGCAGTTAACTCAATCCATTTTTGATATGTCGAAATGGCGTGCGTTAACGCTGCAGGAAAAAGCAGCAGGGATTCAGGACGTCACGTATCAGACCGATCAGCAAACCTTGATCCTCAACACCGCGACCGCTTATTTCAACGTGTTGAATGCTATTGACGTTCTTTCCTATACACAGGCACAAAAAGAAGCGATCTACCGTCAATTAGATCAAACCACCCAACGTTTTAACGTGGGCCTGGTAGCGATCACCGACGTGCAGAACGCCCGCGCACAGTACGATACCGTGCTGGCGAACGAAGTGACCGCACGTAATAACCTTGATAACGCGGTAGAGCAGCTGCGCCAGATCACCGGTAACTACTATCCGGAACTGGCTGCGCTGAATGTCGAAAACTTTAAAACCGACAAACCACAGCCGGTTAACGCGCTGCTGAAAGAAGCCGAAAAACGCAACCTGTCGCTGTTACAGGCACGCTTGAGCCAGGACCTGGCGCGCGAGCAAATTCGCCAGGCGCAGGATGGTCACTTACCGACTCTGGATTTAACGGCTTCTACCGGGATTTCTGACACCTCTTATAGCGGTTCGAAAACCCGTGGTGCCGCTGGTACCCAGTATGACGATAGCAATATGGGCCAGAACAAAGTTGGCCTGAGCTTCTCGCTGCCGATTTATCAGGGCGGAATGGTTAACTCGCAGGTGAAACAGGCACAGTACAACTTTGTCGGTGCCAGCGAGCAACTGGAAAGTGCCCATCGTAGCGTCGTGCAGACCGTGCGTTCCTCCTTCAACAACATTAATGCATCTATCAGTAGCATTAACGCCTACAAACAAGCCGTAGTTTCCGCTCAAAGCTCATTAGACGCGATGGAAGCGGGCTACTCGGTCGGTACGCGTACCATTGTTGATGTGTTGGATGCGACCACCACGTTGTACAACGCCAAGCAAGAGCTGGCGAATGCGCGTTATAACTACCTGATTAATCAGCTGAATATTAAGTCAGCTCTGGGTACGTTGAACGAGCAGGATCTGCTGGCACTGAACAATGCGCTGAGCAAACCGGTTTCCACTAATCCGGAAAACGTTGCACCGCAAACGCCGGAACAGAATGCTATTGCTGATGGTTATGCGCCTGATAGCCCGGCACCAGTCGTTCAGCAAACATCCGCACGCACTACCACCAGTAACGGTCATAACCCTTTCCGTAACTGA
 http://www.ncbi.nlm.nih.gov/nuccore/NC_000913.2/?from=3176137&to=3177618&report=fasta NCBI Reference Sequence: NC_000913.2 from Escherichia coli str. K-12 substr. MG1655, complete genome
-IL-6 fusion gene
-Protease-binding sequence?
+*Protease-binding sequence?
 ==Anti-Inflammatory Device==
@@ Line 76: / Line 174: @@
 '''SoxR''' (Escherichia coli E24377A)
-TAT AGA ATT CGC GGC CGC TTC TAG ATG GAA AAG AAA TTA CCC CGC
+Forward: TATAgaattcgcggccgcttctagATGGAAAAGAAATTACCCCGC
-'''''TAT ATA CTA GTA GCG GCC GCT GCA GTT AGT TTT GTT CAT CTT CCA GCA AGC GTG'''''
+Reverse: '''''TATAtactagtagcggccgctgcagTTAGTTTTGTTCATCTTCCAGCAAGCGTG'''''
+complete gene sequence: [http://www.ncbi.nlm.nih.gov/nuccore/NC_009801.1/?from=4616553&to=4617017&report=fasta]
+'''SoxS promoter''' (Joe 7-5)
+Forward: TATAgaattcgcggccgcttctagagAATCGCTTTACCTCAAGTTAACTTG
+Reverse: TATAtactagtagcggccgctgcagCAGTTCGTTAATTCATCTGTTGGG
+Added 7/1/09 - ''Anusuya''
 '''CrtB''' ''(Erwinia Uredovora)''
-TATAgaattcgcggccgcttctagATGGCAGTTGGCTCGAAAAG
+''Forward'': TATAgaattcgcggccgcttctagATGGCAGTTGGCTCGAAAAG
-'''''TATAtactagtagcggccgctgcagCTAGAGCGGGCGCTGCCAG'''''
+''Reverse'': '''''TATAtactagtagcggccgctgcagCTAGAGCGGGCGCTGCCAG'''''
 '''CrtE''' ''(Erwinia Uredovora)''
-TATAgaattcgcggccgcttctagATGACGGTCTGCGCAAAAAAAC
+''Forward'': TATAgaattcgcggccgcttctagATGACGGTCTGCGCAAAAAAAC
-'''''TATAtactagtagcggccgctgcagTTAACTGACGGCAGCGAGTTTTTTG'''''
+''Reverse'': '''''TATAtactagtagcggccgctgcagTTAACTGACGGCAGCGAGTTTTTTG'''''
 '''CrtI'''  ''(Erwinia Uredovora)''
-TATAgaattcgcggccgcttctag ATGAAACCAACTACGGTAATTGGTGC
+''Forward'': TATAgaattcgcggccgcttctag ATGAAACCAACTACGGTAATTGGTGC
-TATAtactagtagcggccgctgcag TGATTGAGTAACGACGG
+''Reverse'': '''''TATAtactagtagcggccgctgcag TGATTGAGTAACGACGG'''''
 '''CrtY''' ''(Erwinia Uredovora)''
-TATAgaattcgcggccgcttctag ATGCAACCGCATTATGATCTG
+''Forward'': TATAgaattcgcggccgcttctag ATGCAACCGCATTATGATCTG
-TATAtactagtagcggccgctgcag TTAACGATGAGTCGTCATAATGGC
+''Reverse'': '''''TATAtactagtagcggccgctgcag TTAACGATGAGTCGTCATAATGGC'''''
@@ Line 112: / Line 220: @@
-'''Blh''' ''(Halobacterium sp. NRC1)''
+'''Blh''' ''(uncultured marine bacterium 66A03)''
+''Forward'': TATAgaattcgcggccgcttctagATGGGTCTGATGCTGATTG
+''Reverse'': TATAtactagtagcggccgctgcagTCAGTTTTTGATTTTGATACGGG
-TATAgaattcgcggccgcttctagATGCCACACGGCGCGATCG
-'''''TATAtactagtagcggccgctgcagTCAGAGGACGCCCTGCAC'''''
+'''RalDH(II)''' ''(Rattus Norvegicus*)''
-'''RalDH(II)''' ''(Rattus Norvegicus)''
+''Forward'': TATAgaattcgcggccgcttctagATGCCCGGCGAGGTGAAGGC
-TATAgaattcgcggccgcttctagATGCCCGGCGAGGTGAAGGC
+''Reverse'': '''''TATAtactagtagcggccgctgcagTTAGGAGTTCTTCTGGGGGATC'''''
-TATAtactagtagcggccgctgcagTTAGGAGTTCTTCTGGGGGATC
+*Originally from Rattus Norvegicus but sequence was modified
+'''HlyR''': Since HlyR is not a protein. This sequence is sequenced as a chunk with HlyA,HlyB, HlyC. Thus only one primer is needed.
+primer: TATAtactagtagcggccgctgcagGAATTCCAAGCGAAGTCC
+check paper for RalDH(II) http://www.jbc.org/cgi/content/abstract/271/27/16288
 Gene sequences:
-SoxR
+'''SoxR'''
 ATGGAAAAGAAATTACCCCGCATTAAAGCGCTGCTAACCCCCGGCGAAGTGGCGAAACGCAGCGGTGTGGCGGTATCGGCGCTGCATTTCTATGAAAGTAAAGGGTTGATTACCAGTATCCGTAACAGCGGCAATCAGCGGCGATATAAACGTGATGTGTTGCGATATGTTGCAATTATCAAAATTGCTCAGCGTATTGGCATTCCGCTGGCGACCATTGGTGAAGCGTTTGGCGTGTTGCCCGAAGGGCATACGTTAAGTGCGAAAGAGTGGAAACAGCTTTCGTCCCAATGGCGAGAAGAGTTGGATCGGCGCATTCATACCTTAGTGGCGCTGCGTGACGAACTGGACGGATGTATTGGTTGTGGCTGCCTTTCGCGCAGTGATTGCCCGTTGCGTAACCCGGGCGACCGCTTAGGAGAAGAAGGTACCGGCGCACGCTTGCTGGAAGATGAACAAAACTAA
 	http://www.ncbi.nlm.nih.gov/nuccore/NC_009801.1/?from=4616553&to=4617017&report=fasta
 	(NCBI Reference Sequence: NC_009801.1 from Escherichia coli E24377A, complete genome)
+'''SoxS'''
+AATCGCTTTACCTCAAGTTAACTTGAGGAATTATACTCCCCAACAGATGAATTAACGAACTG
 Crt genes
@@ Line 140: / Line 260: @@
 Kim et al. In vitro characterization of a recombinant Blh protein from an uncultured marine bacterium as a beta-carotene 15,15'-dioxygenase. J Biol Chem. 2009 Jun 5;284(23):15781-93. (supplementary material; 15,15'-Dioxygenase, product of Blh gene, codon-optimized and expressed in E. coli, forms a 64k protein of two 32 kDa dimers)
-RALDH?
+RALDH
+HlyR:
+sequence: GAATTCCAAGCGAAGTCCATCCCCCTCCCTCTTGATTACAAGGGTGATAATTATTATTCGCATTTGTGTG
+GTAATGGGATAGAAAGGAATGGATAGAAAAAGAACAAAATTAGTATAGCAATAGATATGCCCACTGCATT
+GAATACTTACAGGGCATTATTTTATTATGTTTAAATTGAAGTGGTCTCTGGTTTGATTTATTTGTTATTC
+AAGGGGGCTGTTTGGAGATCGGAAAATTCTGTACGTTAAGTGTATTATTTAACCAGTTTCGATGCGTAAC
+AGATTGATTTTGCTCAGCGGTTATCGCTTTTAAGTTGTTGCTCTTGGCTATCGCGTTTAGGTTATCCGAT
+TAAAGTCAAATTTCCTGAAAATGCTGTATAGCGCGGGAGTGCACCTTATAGCTGTAGGTAAGTATGTTCA
+AAAAATAGTCTTGCCGTACAATAATTTTCCATATCCAAACTCACTCCTTCAAGATTCTGGTCCCGGTTTA
+CGGGTAGTTTCCGGAAGGGCGGTAGCATGCTGATTCAAACTGCAAGATGAAACATTGTCGGAGTTGGATG
+GAATTAAGTCATGGCTATAGCATTTGGGCGTGCATAACAAAATTGGTCCTCATATTTTAGAGTATGATTG
+CATATTCACTAATATTTTTACTTTCTGATGCGTGGTGGCATCATGCTTTATGAGATAAACAATCCTGG
+blh
+ATGGGTCTGATGCTGATTGATTGGTGTGCACTGGCTCTGGTTGTTTTCATTGGCCTGCCGCACGGCGCG
+CTGGATGCTGCCATTTCTTTTTCTATGATCTCTTCTGCAAAACGCATTGCTCGTCTGGCTGGTATTCTG
+CTGATCTATCTGCTGCTGGCGACCGCGTTCTTCCTGATCTGGTATCAGCTGCCAGCGTTTAGCCTGCTG
+ATCTTCCTGCTGATCTCCATTATCCACTTTGGTATGGCAGACTTCAACGCGTCCCCAAGCAAACTGAAA
+TGGCCGCATATCATCGCCCACGGCGGTGTTGTTACTGTTTGGCTGCCGCTGATCCAGAAAAACGAAGTA
+ACTAAACTGTTTAGCATCCTGACTAACGGTCCGACTCCGATCCTGTGGGACATCCTGCTGATTTTCTTC
+CTGTGTTGGTCTATTGGCGTGTGTCTGCACACGTACGAAACCCTGCGCTCTAAACATTACAACATCGCC
+TTTGAACTGATCGGTCTGATTTTCCTGGCGTGGTATGCGCCGCCTCTGGTTACGTTTGCCACTTACTTC
+TGCTTCATTCATTCCCGTCGCCACTTCTCCTTTGTGTGGAAGCAGCTGCAACACATGTCTTCCAAAAAG
+ATGATGATTGGCAGCGCGATTATCCTGTCCTGTACCTCTTGGCTGATCGGCGGTGGTATCTATTTCTTC
+CTGAACTCCAAAATGATCGCCTCTGAGGCTGCGCTACAAACTGTGTTCATCGGTCTGGCGGCACTGACC
+GTGCCGCACATGATTCTGATCGACTTCATCTTCCGTCCGCACTCTTCCCGTATCAAAATCAAAAACTGA
+RALDH II
+ATGCCCGGCGAGGTGAAGGCCGACCCCGCCGCGCTCATGGCCTCGCTGCAGCTCCTGCCG
+TCACCCACGCCCAACCTCGAGATCAAGTACACCAAGATCTTTATTAACAATGAATGGCAG
+AACTCAGAGAGTGGGAGAGTGTTCCCTGTCTGTAATCCAGCCACAGGAGAGCAAGTGTGT
+GAAGTTCAAGAAGCAGACAAGGTGGATATAGACAAGGCAGTGCAGGCGGCCCGCCTGGCC
+TTCTCTCTTGGTTCTGTGTGGAGAAGGATGGACGCTTCTGAAAGAGGACGTCTGCTGGAC
+AAGCTTGCAGACTTGGTGGAACGGGACAGGGCAACTCTTGCAACTATGGAATCACTAAAT
+GGCGGTAAGCCATTCCTGCAAGCTTTTTACATCGATTTGCAGGGAGTCATCAAAACCCTG
+AGGTATTATGCAGGCTGGGCTGATAAAATTCACGGAATGACCATTCCTGTAGATGGAGAC
+TATTTTACCTTCACGAGACATGAGCCCATCGGAGTGTGTGGACAGATTATCCCGTGGAAT
+TTCCCCCTGCTGATGTTCACCTGGAAAATTGCTCCCGCACTGTGCTGTGGTAACACTGTG
+GTCATCAAGCCTGCAGAGCAGACACCACTCAGTGCACTGTATATGGGAGCCCTCATCAAG
+GAGGCTGGCTTTCCACCCGGGGTCGTCAACATTCTGCCAGGGTATGGGCCAACAGCAGGG
+GCAGCAATAGCGTCTCACATTGGCATAGACAAGATTGCATTCACAGGCTCTACTGAGGTT
+GGAAAGCTTATTCAAGAAGCAGCTGGAAGAAGTAATTTGAAGAGAGTAACTCTGGAACTT
+GGAGGGAAAAGCCCTAATATTATTTTTGCAGATGCTGACTTGGACTACGCTGTGGAGCAG
+GCTCACCAGGGTGTGTTCTTCAACCAAGGCCAGTGCTGCACTGCGGGGTCTCGAATCTTT
+GTGGAGGAGTCCATCTATGAGGAATTTGTGAAAAGAAGTGTGGAAAGAGCCAAGAGGCGC
+ATTGTGGGGAGCCCCTTTGATCCCACTACTGAGCAGGGTCCTCAGATTGATAAGAAGCAA
+TACAACAAAATCCTGGAGCTCATCCAGAGCGGTGTGGCCGAGGGCGCCAAGCTCGAGTGT
+GGAGGCAAAGGACTGGGAAGGAAGGGCTTCTTCATTGAGCCTACCGTGTTCTCCAACGTT
+ACAGATGACATGCGGATCGCCAAGGAGGAGATCTTTGGTCCTGTCCAGGAAATTTTGAGG
+TTTAAGACTATGGATGAAGTTATAGAAAGAGCCAATAACTCAGACTTCGGGCTTGTAGCG
+GCTGTCTTCACTAATGACATCAACAAGGCTCTCATGGTGTCCTCTGCAATGCAAGCTGGG
+ACAGTTTGGATCAATTGTTACAACGCCTTAAATGCTCAGAGCCCCTTCGGGGGGTTCAAG
+ATGTCTGGAAATGGGAGAGAAATGGGTGAGTTTGGCTTACGGGAGTACTCAGAAGTAAAG
+ACCGTGACGGTGAAGATCCCCCAGAAGAACTCCTAA
+==Comments==
+*For gene sequencing, I would suggest genedesigner
+*For primer design and selection I would suggest APE and oligo 6.87