LOCUS       sequence-199474-        2480 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  In vitro expression of CyPet from the T7 promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2480)
  AUTHORS   Stark JC, Huang A, Nguyen PQ, Dubner RS, Hsu KJ, Ferrante TC, 
            Anderson M, Kanapskyte A, Mucha Q, Packett JS, Patel P, Patel R, Qaq
            D, Zondor T, Burke J, Martinez T, Miller-Berry A, Puppala A, 
            Reichert K, Schmid M, Brand L, Hill LR, Chellaswamy JF, Faheem N, 
            Fetherling S, Gong E, Gonzalzles EM, Granito T, Koritsaris J, Nguyen
            B, Ottman S, Palffy C, Patel A, Skweres S, Slaton A, Woods T, 
            Donghia N, Pardee K, Collins JJ, Jewett MC
  TITLE     BioBits Bright: A fluorescent synthetic biology education kit.
  JOURNAL   Sci Adv. 2018 Aug 1;4(8):eaat5107. doi: 10.1126/sciadv.aat5107. 
            eCollection 2018 Aug.
  PUBMED    30083609
REFERENCE   2  (bases 1 to 2480)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Sci Adv."; 
            date: "2018-08-1"; volume: "4(8):eaat5107. doi"; pages: " 
            10.1126/sciadv.aat5107. eCollection 2018 Aug"
COMMENT     Sequence Label: pJL1-CyPet
FEATURES             Location/Qualifiers
     source          1..2480
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     32..51
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     159..178
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        159..177
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             209..231
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             239..952
                     /codon_start=1
                     /product="cyan fluorescent protein optimized for FRET with 
                     YPet (Nguyen and Daugherty, 2005)"
                     /label=CyPet
                     /translation="MSKGEELFGGIVPILVELEGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSVMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIKA
                     NFKARHNITDGSVQLADHYQQNTPIGDGPVILPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(1007..1025)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1021..1068
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(1086..1901)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSKPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     1809..1828
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     1893..1912
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      join(2023..2480,1..131)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
       61 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
      121 gcctatggaa aaacgccagc aacgcgatcc cgcgaaatta atacgactca ctatagggag
      181 accacaacgg tttccctcta gaaataattt tgtttaactt taagaaggag atatacatat
      241 gtctaaaggt gaagaattat tcggcggtat cgtcccaatt ttagttgaat tagagggtga
      301 tgttaatggt cacaaatttt ctgtctccgg tgaaggtgaa ggtgatgcta cgtacggtaa
      361 attgacctta aaatttattt gtactactgg taaattgcca gttccatggc caaccttagt
      421 cactactctg acttggggtg ttcaatgttt ttctagatac ccagatcata tgaaacaaca
      481 tgactttttc aagtctgtca tgccagaagg ttatgttcaa gaaagaacta tttttttcaa
      541 agatgacggt aactacaaga ccagagctga agtcaagttt gaaggtgata ccttagttaa
      601 tagaatcgaa ttaaaaggta ttgattttaa agaagatggt aacattttag gtcacaaatt
      661 ggaatacaac tatatctctc acaatgttta catcaccgct gacaaacaaa agaatggtat
      721 caaagctaac ttcaaagcca gacacaacat taccgatggt tctgttcaat tagctgacca
      781 ttatcaacaa aatactccaa ttggtgatgg tccagtcatc ttgccagaca accattactt
      841 atccactcaa tctgccttat ctaaagatcc aaacgaaaag agagaccaca tggtcttgct
      901 cgaatttgtt actgctgctg gtattaccca tggtatggat gaattgtaca aataagtcga
      961 ccggctgcta acaaagcccg aaaggaagct gagttggctg ctgccaccgc tgagcaataa
     1021 ctagcataac cccttggggc ctctaaacgg gtcttgaggg gttttttgct gaaagccaat
     1081 tctgattaga aaaactcatc gagcatcaaa tgaaactgca atttattcat atcaggatta
     1141 tcaataccat atttttgaaa aagccgtttc tgtaatgaag gagaaaactc accgaggcag
     1201 ttccatagga tggcaagatc ctggtatcgg tctgcgattc cgactcgtcc aacatcaata
     1261 caacctatta atttcccctc gtcaaaaata aggttatcaa gtgagaaatc accatgagtg
     1321 acgactgaat ccggtgagaa tggcaaaagc ttatgcattt ctttccagac ttgttcaaca
     1381 ggccagccat tacgctcgtc atcaaaatca ctcgcatcaa ccaaaccgtt attcattcgt
     1441 gattgcgcct gagcgagacg aaatacgcga tcgctgttaa aaggacaatt acaaacagga
     1501 atcgaatgca accggcgcag gaacactgcc agcgcatcaa caatattttc acctgaatca
     1561 ggatattctt ctaatacctg gaatgctgtt ttcccgggga tcgcagtggt gagtaaccat
     1621 gcatcatcag gagtacggat aaaatgcttg atggtcggaa gaggcataaa ttccgtcagc
     1681 cagtttagtc tgaccatctc atctgtaaca tcattggcaa cgctaccttt gccatgtttc
     1741 agaaacaact ctggcgcatc gggcttccca tacaatcgat agattgtcgc acctgattgc
     1801 ccgacattat cgcgagccca tttataccca tataaatcag catccatgtt ggaatttaat
     1861 cgcggcttcg agcaagacgt ttcccgttga atatggctca taacacccct tgtattactg
     1921 tttatgtaag cagacagttt tattgttcat gatgatatat ttttatcttg tgcaatgtaa
     1981 catcagagat tttgagacac aacgtgagat caaaggatct tcttgagatc ctttttttct
     2041 gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc
     2101 ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc
     2161 aaatactgtt cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc
     2221 gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc
     2281 gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg
     2341 aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata
     2401 cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta
     2461 tccggtaagc ggcagggtcg
//
