LOCUS       sequence-49417-e        5153 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 5153)
  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 5153)
  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: pRAJ12
FEATURES             Location/Qualifiers
     source          1..5153
                     /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    868..886
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    893..911
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    950..966
                     /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             1013..1024
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             1040..1750
                     /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    1757..1777
                     /label=BioBrick suffix
                     /note="universal suffix for all parts"
     terminator      1813..1844
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
     CDS             complement(1943..2758)
                     /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     2666..2685
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2750..2769
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             complement(3116..4066)
                     /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      4114..4336
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     terminator      complement(4828..4922)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     terminator      complement(5078..5107)
                     /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 atcgatgcga aaaaaccccg ccgaagcggg gttttttgcg
      781 gctagcccca gaatcccttc gagcaaatcc tagcgaggtg ccgctagatt cgctcaaagg
      841 aacccttctt cctgcccgtg ctcagtatct ctatcactga tagggatgtc aatctctatc
      901 actgataggg acctaggatg catataaatg tgagcggata acattgacat tgtgagcgga
      961 taacaagata ctgagcacac ccagtatcat tctcttcttc ctgcccacgc ggaaagagga
     1021 gaaaggtgta atgactagcc gtaaaggcga agagctgttc actggtgtcg tccctattct
     1081 ggtggaactg gatggtgatg tcaacggtca taagttttcc gtgcgtggcg agggtgaagg
     1141 tgacgcaact aatggtaaac tgacgctgaa gttcatctgt actactggta aactgccggt
     1201 accttggccg actctggtaa cgacgctgac ttatggtgtt cagtgctttg ctcgttatcc
     1261 ggaccatatg aagcagcatg acttcttcaa gtccgccatg ccggaaggct atgtgcagga
     1321 acgcacgatt tcctttaagg atgacggcac gtacaaaacg cgtgcggaag tgaaatttga
     1381 aggcgatacc ctggtaaacc gcattgagct gaaaggcatt gactttaaag aagacggcaa
     1441 tatcctgggc cataagctgg aatacaattt taacagccac aatgtttaca tcaccgccga
     1501 taaacaaaaa aatggcatta aagcgaattt taaaattcgc cacaacgtgg aggatggcag
     1561 cgtgcagctg gctgatcact accagcaaaa cactccaatc ggtgatggtc ctgttctgct
     1621 gccagacaat cactatctga gcacgcaaag cgttctgtct aaagatccga acgagaaacg
     1681 cgatcatatg gttctgctgg agttcgtaac cgcagcgggc atcacgcatg gtatggatga
     1741 actgtacaaa tgatgatact agtagcggcc gctgcaggag tcactaaggg ttagttagtt
     1801 agattagcag aaagtcaaaa gcctccgacc ggaggctttt gactaaaact tcccttgggg
     1861 ttatcattgg ggctcactca aaggcggtaa tcagataaaa aaaatcctta gctttcgcta
     1921 aggatgattt ctgctagtat tattagaaaa actcatcgag catcaaatga aactgcaatt
     1981 tattcatatc aggattatca ataccatatt tttgaaaaag ccgtttctgt aatgaaggag
     2041 aaaactcacc gaggcagttc caaagaatgg caaggtcctg gtaacggtct gcgattccga
     2101 cccgtccaac atcaatacaa cctattaatt tcccctcgtc aaaaataagg ttatcaagtg
     2161 agaaatcacc atgagtgacg actgaatccg gtgagaatgg caagagcttg tgcatttctt
     2221 tccagacttg ttcaacaggc cagccattac gctcgtcatc aaaatcactc gcatcaacca
     2281 aaccgttatt catgcgtgat tgcgcctgag caagacgaaa tacacgatcg ctgttaaaag
     2341 gacaattaca aacaggaatc gaatgtaacc ggcgcaggaa cacggccagc gcatcaacaa
     2401 tattttcacc tgaatcagga tattcttcta atacctggaa ggctgttttc ccaggaatcg
     2461 cggtggtgag taaccacgca tcatcaggag tacggataaa atgcttgatg gtcgggagag
     2521 gcataaactc cgtcagccag ttgagacgga ccatctcatc tgtaacatca ttggcaacgc
     2581 tacctttgcc atgtttcaga aacaactctg gcgcatcggg cttcccatac aagcgataga
     2641 ttgtcgcacc tgattgcccg acattatcgc gagcccattt atacccatat aaatcagcgt
     2701 ccatgttgga gtttaagcgc ggacgggagc aagacgtttc ccgttgaata tggctcataa
     2761 caccccttgt attactgttt atgtaagcag acagttttat tgttcatgat gatatatttt
     2821 tatcttgtgc aatgtaacat cagagatttt gagacacaac gtggctttgt tgaataaatc
     2881 gaacttttgc tgagttgaag gatcagctct agtagttaca ttgtcgatct gttcatggtg
     2941 aacagcttta aatgcaccaa aaactcgtaa aagctctgat gtatctatct tttttacacc
     3001 gttttcatct gtgcatatgg acagttttcc ctttgatatc taacggtgaa cagttgttct
     3061 acttttgttt gttagtcttg atgcttcact gatagataca agagccataa gaacctcaga
     3121 tccttccgta tttagccagt atgttctcta gtgtggttcg ttgtttttgc gtgagccatg
     3181 agaacgaacc attgagatca tgcttacttt gcatgtcact caaaaatttt gcctcaaaac
     3241 tggtgagctg aatttttgca gttaaagcat cgtgtagtgt ttttcttagt ccgttacgta
     3301 ggtaggaatc tgatgtaatg gttgttggta ttttgtcacc attcattttt atctggttgt
     3361 tctcaagttc ggttacgaga tccatttgtc tatctagttc aacttggaaa atcaacgtat
     3421 cagtcgggcg gcctcgctta tcaaccacca atttcatatt gctgtaagtg tttaaatctt
     3481 tacttattgg tttcaaaacc cattggttaa gccttttaaa ctcatggtag ttattttcaa
     3541 gcattaacat gaacttaaat tcatcaaggc taatctctat atttgccttg tgagttttct
     3601 tttgtgttag ttcttttaat aaccactcat aaatcctcat agagtatttg ttttcaaaag
     3661 acttaacatg ttccagatta tattttatga atttttttaa ctggaaaaga taaggcaata
     3721 tctcttcact aaaaactaat tctaattttt cgcttgagaa cttggcatag tttgtccact
     3781 ggaaaatctc aaagccttta accaaaggat tcctgatttc cacagttctc gtcatcagct
     3841 ctctggttgc tttagctaat acaccataag cattttccct actgatgttc atcatctgag
     3901 cgtattggtt ataagtgaac gataccgtcc gttctttcct tgtagggttt tcaatcgtgg
     3961 ggttgagtag tgccacacag cataaaatta gcttggtttc atgctccgtt aagtcatagc
     4021 gactaatcgc tagttcattt gctttgaaaa caactaattc agacatacat ctcaattggt
     4081 ctaggtgatt ttaatcacta taccaattga gatgggctag tcaatgataa ttacatgtcc
     4141 ttttcctttg agttgtgggt atctgtaaat tctgctagac ctttgctgga aaacttgtaa
     4201 attctgctag accctctgta aattccgcta gacctttgtg tgtttttttt gtttatattc
     4261 aagtggttat aatttataga ataaagaaag aataaaaaaa gataaaaaga atagatccca
     4321 gccctgtgta taactcacta ctttagtcag ttccgcagta ttacaaaagg atgtcgcaaa
     4381 cgctgtttgc tcctctacaa aacagacctt aaaaccctaa aggcttaagt agcaccctcg
     4441 caagctcggg caaatcgctg aatattcctt ttgtctccga ccatcaggca cctgagtcgc
     4501 tgtctttttc gtgacattca gttcgctgcg ctcacggctc tggcagtgaa tgggggtaaa
     4561 tggcactaca ggcgcctttt atggattcat gcaaggaaac tacccataat acaagaaaag
     4621 cccgtcacgg gcttctcagg gcgttttatg gcgggtctgc tatgtggtgc tatctgactt
     4681 tttgctgttc agcagttcct gccctctgat tttccagtct gaccacttcg gattatcccg
     4741 tgacaggtca ttcagactgg ctaatgcacc cagtaaggca gcggtatcat caacaggctt
     4801 acccgtctta ctgtccctag tgcttggatt ctcaccaata aaaaacgccc ggcggcaacc
     4861 gagcgttctg aacaaatcca gatggagttc tgaggtcatt actggatcta tcaacaggag
     4921 tccaagcgag ctcgtaaact tggtctgaca gctctagctc cggcaaaaaa acgggcaagg
     4981 tgtcaccacc ctgccctttt tctttaaaac cgaaaagatt acttcgcgtt tgccacctga
     5041 cgtctaagaa aaggaatatt cagcaatttg cccgtgccga agaaaggccc acccgtgaag
     5101 gtgagccagt gagttgattg ctacgtaatt agttagttag cccttagtga ctc
//
