LOCUS       sequence-49419-e        5107 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5107)
  AUTHORS   Rodrigo G, Landrain TE, Jaramillo A
  TITLE     De novo automated design of small RNA circuits for engineering 
            synthetic riboregulation in living cells.
  JOURNAL   Proc Natl Acad Sci U S A. 2012 Sep 18;109(38):15271-6. Epub 2012 Sep
            4.
  PUBMED    22949707
REFERENCE   2  (bases 1 to 5107)
  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: "Proc Natl 
            Acad Sci U S A."; date: "2012-09-18"; volume: "109(38)"; pages: 
            "15271-6. Epub 2012 Sep 4"
COMMENT     Sequence Label: pRAJ12f
FEATURES             Location/Qualifiers
     source          1..5107
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    1..22
                     /label=BioBrick prefix
                     /note="BioBrick prefix for parts that do not start with 
                     ""ATG"""
     CDS             complement(26..703)
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     RBS             710..721
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     protein_bind    822..840
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    847..865
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    904..920
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     RBS             967..978
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             994..1704
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /translation="RKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLKF
                     ICTTGKLPVPWPTLVTTLTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTISFKDDGT
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKAN
                     FKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLEF
                     VTAAGITHGMDELYK"
     misc_feature    1711..1731
                     /label=BioBrick suffix
                     /note="universal suffix for all parts"
     terminator      1767..1798
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
     CDS             complement(1897..2712)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     2620..2639
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2704..2723
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             complement(3070..4020)
                     /codon_start=1
                     /gene="rep101"
                     /product="RepA protein needed for replication with the 
                     pSC101 origin"
                     /label=Rep101
                     /translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
                     RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFS
                     SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
                     EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
                     ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLRNGLRKTLHDALTAKIQLTSFEAKF
                     LSDMQSKHDLNGSFSWLTQKQRTTLENILAKYGRI"
     rep_origin      4068..4290
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     terminator      complement(4782..4876)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     terminator      complement(5032..5061)
                     /label=T3Te terminator
                     /note="phage T3 early transcription terminator"
ORIGIN
        1 gaattcgcgg ccgcttctag agttattaag caccggtgga gtgacgacct tcagcacgtt
       61 cgtactgttc aacgatggtg tagtcttcgt tgtgggaggt gatgtccagt ttgatgtcgg
      121 ttttgtaagc acccggcagc tgaaccggtt ttttagccat gtaggtggtt ttaacttcag
      181 cgtcgtagtg accaccgtct ttcagtttca gacgcatttt gatttcacct ttcagagcac
      241 cgtcttccgg gtacatacgt tcggtggaag cttcccaacc catggttttt ttctgcataa
      301 ccggaccgtc ggacgggaag ttggtaccac gcagtttaac tttgtagatg aactcaccgt
      361 cttgcaggga ggagtcctgg gtaacggtaa caacaccacc gtcttcgaag ttcataacac
      421 gttcccattt gaaaccttcc gggaaggaca gtttcaggta gtccgggatg tcagccgggt
      481 gtttaacgta agctttggaa ccgtactgga actgcgggga caggatgtcc caagcgaacg
      541 gcagcggacc acctttggta actttcagtt tagcggtctg ggtaccttcg tacggacgac
      601 cttcaccttc accttcgatt tcgaactcgt gaccgttaac ggaaccttcc atacgaactt
      661 tgaaacgcat gaactctttg ataacgtctt cggaggaagc catctagtat ttctcctctt
      721 tctctagtgg atccgctagc cccagaatcc cttcgagcaa atcctagcga ggtgccgcta
      781 gattcgctca aaggaaccct tcttcctgcc cgtgctcagt atctctatca ctgataggga
      841 tgtcaatctc tatcactgat agggacctag gatgcatata aatgtgagcg gataacattg
      901 acattgtgag cggataacaa gatactgagc acacccagta tcattctctt cttcctgccc
      961 acgcggaaag aggagaaagg tgtaatgact agccgtaaag gcgaagagct gttcactggt
     1021 gtcgtcccta ttctggtgga actggatggt gatgtcaacg gtcataagtt ttccgtgcgt
     1081 ggcgagggtg aaggtgacgc aactaatggt aaactgacgc tgaagttcat ctgtactact
     1141 ggtaaactgc cggtaccttg gccgactctg gtaacgacgc tgacttatgg tgttcagtgc
     1201 tttgctcgtt atccggacca tatgaagcag catgacttct tcaagtccgc catgccggaa
     1261 ggctatgtgc aggaacgcac gatttccttt aaggatgacg gcacgtacaa aacgcgtgcg
     1321 gaagtgaaat ttgaaggcga taccctggta aaccgcattg agctgaaagg cattgacttt
     1381 aaagaagacg gcaatatcct gggccataag ctggaataca attttaacag ccacaatgtt
     1441 tacatcaccg ccgataaaca aaaaaatggc attaaagcga attttaaaat tcgccacaac
     1501 gtggaggatg gcagcgtgca gctggctgat cactaccagc aaaacactcc aatcggtgat
     1561 ggtcctgttc tgctgccaga caatcactat ctgagcacgc aaagcgttct gtctaaagat
     1621 ccgaacgaga aacgcgatca tatggttctg ctggagttcg taaccgcagc gggcatcacg
     1681 catggtatgg atgaactgta caaatgatga tactagtagc ggccgctgca ggagtcacta
     1741 agggttagtt agttagatta gcagaaagtc aaaagcctcc gaccggaggc ttttgactaa
     1801 aacttccctt ggggttatca ttggggctca ctcaaaggcg gtaatcagat aaaaaaaatc
     1861 cttagctttc gctaaggatg atttctgcta gtattattag aaaaactcat cgagcatcaa
     1921 atgaaactgc aatttattca tatcaggatt atcaatacca tatttttgaa aaagccgttt
     1981 ctgtaatgaa ggagaaaact caccgaggca gttccaaaga atggcaaggt cctggtaacg
     2041 gtctgcgatt ccgacccgtc caacatcaat acaacctatt aatttcccct cgtcaaaaat
     2101 aaggttatca agtgagaaat caccatgagt gacgactgaa tccggtgaga atggcaagag
     2161 cttgtgcatt tctttccaga cttgttcaac aggccagcca ttacgctcgt catcaaaatc
     2221 actcgcatca accaaaccgt tattcatgcg tgattgcgcc tgagcaagac gaaatacacg
     2281 atcgctgtta aaaggacaat tacaaacagg aatcgaatgt aaccggcgca ggaacacggc
     2341 cagcgcatca acaatatttt cacctgaatc aggatattct tctaatacct ggaaggctgt
     2401 tttcccagga atcgcggtgg tgagtaacca cgcatcatca ggagtacgga taaaatgctt
     2461 gatggtcggg agaggcataa actccgtcag ccagttgaga cggaccatct catctgtaac
     2521 atcattggca acgctacctt tgccatgttt cagaaacaac tctggcgcat cgggcttccc
     2581 atacaagcga tagattgtcg cacctgattg cccgacatta tcgcgagccc atttataccc
     2641 atataaatca gcgtccatgt tggagtttaa gcgcggacgg gagcaagacg tttcccgttg
     2701 aatatggctc ataacacccc ttgtattact gtttatgtaa gcagacagtt ttattgttca
     2761 tgatgatata tttttatctt gtgcaatgta acatcagaga ttttgagaca caacgtggct
     2821 ttgttgaata aatcgaactt ttgctgagtt gaaggatcag ctctagtagt tacattgtcg
     2881 atctgttcat ggtgaacagc tttaaatgca ccaaaaactc gtaaaagctc tgatgtatct
     2941 atctttttta caccgttttc atctgtgcat atggacagtt ttccctttga tatctaacgg
     3001 tgaacagttg ttctactttt gtttgttagt cttgatgctt cactgataga tacaagagcc
     3061 ataagaacct cagatccttc cgtatttagc cagtatgttc tctagtgtgg ttcgttgttt
     3121 ttgcgtgagc catgagaacg aaccattgag atcatgctta ctttgcatgt cactcaaaaa
     3181 ttttgcctca aaactggtga gctgaatttt tgcagttaaa gcatcgtgta gtgtttttct
     3241 tagtccgtta cgtaggtagg aatctgatgt aatggttgtt ggtattttgt caccattcat
     3301 ttttatctgg ttgttctcaa gttcggttac gagatccatt tgtctatcta gttcaacttg
     3361 gaaaatcaac gtatcagtcg ggcggcctcg cttatcaacc accaatttca tattgctgta
     3421 agtgtttaaa tctttactta ttggtttcaa aacccattgg ttaagccttt taaactcatg
     3481 gtagttattt tcaagcatta acatgaactt aaattcatca aggctaatct ctatatttgc
     3541 cttgtgagtt ttcttttgtg ttagttcttt taataaccac tcataaatcc tcatagagta
     3601 tttgttttca aaagacttaa catgttccag attatatttt atgaattttt ttaactggaa
     3661 aagataaggc aatatctctt cactaaaaac taattctaat ttttcgcttg agaacttggc
     3721 atagtttgtc cactggaaaa tctcaaagcc tttaaccaaa ggattcctga tttccacagt
     3781 tctcgtcatc agctctctgg ttgctttagc taatacacca taagcatttt ccctactgat
     3841 gttcatcatc tgagcgtatt ggttataagt gaacgatacc gtccgttctt tccttgtagg
     3901 gttttcaatc gtggggttga gtagtgccac acagcataaa attagcttgg tttcatgctc
     3961 cgttaagtca tagcgactaa tcgctagttc atttgctttg aaaacaacta attcagacat
     4021 acatctcaat tggtctaggt gattttaatc actataccaa ttgagatggg ctagtcaatg
     4081 ataattacat gtccttttcc tttgagttgt gggtatctgt aaattctgct agacctttgc
     4141 tggaaaactt gtaaattctg ctagaccctc tgtaaattcc gctagacctt tgtgtgtttt
     4201 ttttgtttat attcaagtgg ttataattta tagaataaag aaagaataaa aaaagataaa
     4261 aagaatagat cccagccctg tgtataactc actactttag tcagttccgc agtattacaa
     4321 aaggatgtcg caaacgctgt ttgctcctct acaaaacaga ccttaaaacc ctaaaggctt
     4381 aagtagcacc ctcgcaagct cgggcaaatc gctgaatatt ccttttgtct ccgaccatca
     4441 ggcacctgag tcgctgtctt tttcgtgaca ttcagttcgc tgcgctcacg gctctggcag
     4501 tgaatggggg taaatggcac tacaggcgcc ttttatggat tcatgcaagg aaactaccca
     4561 taatacaaga aaagcccgtc acgggcttct cagggcgttt tatggcgggt ctgctatgtg
     4621 gtgctatctg actttttgct gttcagcagt tcctgccctc tgattttcca gtctgaccac
     4681 ttcggattat cccgtgacag gtcattcaga ctggctaatg cacccagtaa ggcagcggta
     4741 tcatcaacag gcttacccgt cttactgtcc ctagtgcttg gattctcacc aataaaaaac
     4801 gcccggcggc aaccgagcgt tctgaacaaa tccagatgga gttctgaggt cattactgga
     4861 tctatcaaca ggagtccaag cgagctcgta aacttggtct gacagctcta gctccggcaa
     4921 aaaaacgggc aaggtgtcac caccctgccc tttttcttta aaaccgaaaa gattacttcg
     4981 cgtttgccac ctgacgtcta agaaaaggaa tattcagcaa tttgcccgtg ccgaagaaag
     5041 gcccacccgt gaaggtgagc cagtgagttg attgctacgt aattagttag ttagccctta
     5101 gtgactc
//
