LOCUS       sequence-49422-e        5192 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 5192)
  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 5192)
  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: pRAJ31f
FEATURES             Location/Qualifiers
     source          1..5192
                     /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    907..925
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    932..950
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    989..1005
                     /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             1052..1063
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             1079..1789
                     /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    1796..1816
                     /label=BioBrick suffix
                     /note="universal suffix for all parts"
     terminator      1852..1883
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
     CDS             complement(1982..2797)
                     /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     2705..2724
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2789..2808
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             complement(3155..4105)
                     /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      4153..4375
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     terminator      complement(4867..4961)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     terminator      complement(5117..5146)
                     /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 aaaaaaagat cgtagccatg tagcaaatag ctactaagac
      781 caatacagtg acagcggata gggacttaat tgcggatgag acgacggtaa agcttaatat
      841 tgatcctagc aaccatttac cacataacta cgacccaggc gctggacgtt tgatgtgtgc
      901 tcagtatctc tatcactgat agggatgtca atctctatca ctgataggga cctaggatgc
      961 atataaatgt gagcggataa cattgacatt gtgagcggat aacaagatac tgagcacagt
     1021 ggatatgctt tacctcttgg tgagacgaca taaagaggag aaacataata tgactagccg
     1081 taaaggcgaa gagctgttca ctggtgtcgt ccctattctg gtggaactgg atggtgatgt
     1141 caacggtcat aagttttccg tgcgtggcga gggtgaaggt gacgcaacta atggtaaact
     1201 gacgctgaag ttcatctgta ctactggtaa actgccggta ccttggccga ctctggtaac
     1261 gacgctgact tatggtgttc agtgctttgc tcgttatccg gaccatatga agcagcatga
     1321 cttcttcaag tccgccatgc cggaaggcta tgtgcaggaa cgcacgattt cctttaagga
     1381 tgacggcacg tacaaaacgc gtgcggaagt gaaatttgaa ggcgataccc tggtaaaccg
     1441 cattgagctg aaaggcattg actttaaaga agacggcaat atcctgggcc ataagctgga
     1501 atacaatttt aacagccaca atgtttacat caccgccgat aaacaaaaaa atggcattaa
     1561 agcgaatttt aaaattcgcc acaacgtgga ggatggcagc gtgcagctgg ctgatcacta
     1621 ccagcaaaac actccaatcg gtgatggtcc tgttctgctg ccagacaatc actatctgag
     1681 cacgcaaagc gttctgtcta aagatccgaa cgagaaacgc gatcatatgg ttctgctgga
     1741 gttcgtaacc gcagcgggca tcacgcatgg tatggatgaa ctgtacaaat gatgatacta
     1801 gtagcggccg ctgcaggagt cactaagggt tagttagtta gattagcaga aagtcaaaag
     1861 cctccgaccg gaggcttttg actaaaactt cccttggggt tatcattggg gctcactcaa
     1921 aggcggtaat cagataaaaa aaatccttag ctttcgctaa ggatgatttc tgctagtatt
     1981 attagaaaaa ctcatcgagc atcaaatgaa actgcaattt attcatatca ggattatcaa
     2041 taccatattt ttgaaaaagc cgtttctgta atgaaggaga aaactcaccg aggcagttcc
     2101 aaagaatggc aaggtcctgg taacggtctg cgattccgac ccgtccaaca tcaatacaac
     2161 ctattaattt cccctcgtca aaaataaggt tatcaagtga gaaatcacca tgagtgacga
     2221 ctgaatccgg tgagaatggc aagagcttgt gcatttcttt ccagacttgt tcaacaggcc
     2281 agccattacg ctcgtcatca aaatcactcg catcaaccaa accgttattc atgcgtgatt
     2341 gcgcctgagc aagacgaaat acacgatcgc tgttaaaagg acaattacaa acaggaatcg
     2401 aatgtaaccg gcgcaggaac acggccagcg catcaacaat attttcacct gaatcaggat
     2461 attcttctaa tacctggaag gctgttttcc caggaatcgc ggtggtgagt aaccacgcat
     2521 catcaggagt acggataaaa tgcttgatgg tcgggagagg cataaactcc gtcagccagt
     2581 tgagacggac catctcatct gtaacatcat tggcaacgct acctttgcca tgtttcagaa
     2641 acaactctgg cgcatcgggc ttcccataca agcgatagat tgtcgcacct gattgcccga
     2701 cattatcgcg agcccattta tacccatata aatcagcgtc catgttggag tttaagcgcg
     2761 gacgggagca agacgtttcc cgttgaatat ggctcataac accccttgta ttactgttta
     2821 tgtaagcaga cagttttatt gttcatgatg atatattttt atcttgtgca atgtaacatc
     2881 agagattttg agacacaacg tggctttgtt gaataaatcg aacttttgct gagttgaagg
     2941 atcagctcta gtagttacat tgtcgatctg ttcatggtga acagctttaa atgcaccaaa
     3001 aactcgtaaa agctctgatg tatctatctt ttttacaccg ttttcatctg tgcatatgga
     3061 cagttttccc tttgatatct aacggtgaac agttgttcta cttttgtttg ttagtcttga
     3121 tgcttcactg atagatacaa gagccataag aacctcagat ccttccgtat ttagccagta
     3181 tgttctctag tgtggttcgt tgtttttgcg tgagccatga gaacgaacca ttgagatcat
     3241 gcttactttg catgtcactc aaaaattttg cctcaaaact ggtgagctga atttttgcag
     3301 ttaaagcatc gtgtagtgtt tttcttagtc cgttacgtag gtaggaatct gatgtaatgg
     3361 ttgttggtat tttgtcacca ttcattttta tctggttgtt ctcaagttcg gttacgagat
     3421 ccatttgtct atctagttca acttggaaaa tcaacgtatc agtcgggcgg cctcgcttat
     3481 caaccaccaa tttcatattg ctgtaagtgt ttaaatcttt acttattggt ttcaaaaccc
     3541 attggttaag ccttttaaac tcatggtagt tattttcaag cattaacatg aacttaaatt
     3601 catcaaggct aatctctata tttgccttgt gagttttctt ttgtgttagt tcttttaata
     3661 accactcata aatcctcata gagtatttgt tttcaaaaga cttaacatgt tccagattat
     3721 attttatgaa tttttttaac tggaaaagat aaggcaatat ctcttcacta aaaactaatt
     3781 ctaatttttc gcttgagaac ttggcatagt ttgtccactg gaaaatctca aagcctttaa
     3841 ccaaaggatt cctgatttcc acagttctcg tcatcagctc tctggttgct ttagctaata
     3901 caccataagc attttcccta ctgatgttca tcatctgagc gtattggtta taagtgaacg
     3961 ataccgtccg ttctttcctt gtagggtttt caatcgtggg gttgagtagt gccacacagc
     4021 ataaaattag cttggtttca tgctccgtta agtcatagcg actaatcgct agttcatttg
     4081 ctttgaaaac aactaattca gacatacatc tcaattggtc taggtgattt taatcactat
     4141 accaattgag atgggctagt caatgataat tacatgtcct tttcctttga gttgtgggta
     4201 tctgtaaatt ctgctagacc tttgctggaa aacttgtaaa ttctgctaga ccctctgtaa
     4261 attccgctag acctttgtgt gttttttttg tttatattca agtggttata atttatagaa
     4321 taaagaaaga ataaaaaaag ataaaaagaa tagatcccag ccctgtgtat aactcactac
     4381 tttagtcagt tccgcagtat tacaaaagga tgtcgcaaac gctgtttgct cctctacaaa
     4441 acagacctta aaaccctaaa ggcttaagta gcaccctcgc aagctcgggc aaatcgctga
     4501 atattccttt tgtctccgac catcaggcac ctgagtcgct gtctttttcg tgacattcag
     4561 ttcgctgcgc tcacggctct ggcagtgaat gggggtaaat ggcactacag gcgcctttta
     4621 tggattcatg caaggaaact acccataata caagaaaagc ccgtcacggg cttctcaggg
     4681 cgttttatgg cgggtctgct atgtggtgct atctgacttt ttgctgttca gcagttcctg
     4741 ccctctgatt ttccagtctg accacttcgg attatcccgt gacaggtcat tcagactggc
     4801 taatgcaccc agtaaggcag cggtatcatc aacaggctta cccgtcttac tgtccctagt
     4861 gcttggattc tcaccaataa aaaacgcccg gcggcaaccg agcgttctga acaaatccag
     4921 atggagttct gaggtcatta ctggatctat caacaggagt ccaagcgagc tcgtaaactt
     4981 ggtctgacag ctctagctcc ggcaaaaaaa cgggcaaggt gtcaccaccc tgcccttttt
     5041 ctttaaaacc gaaaagatta cttcgcgttt gccacctgac gtctaagaaa aggaatattc
     5101 agcaatttgc ccgtgccgaa gaaaggccca cccgtgaagg tgagccagtg agttgattgc
     5161 tacgtaatta gttagttagc ccttagtgac tc
//
