LOCUS       sequence-49421-e        4976 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 4976)
  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 4976)
  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: pRAJ31m
FEATURES             Location/Qualifiers
     source          1..4976
                     /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    773..789
                     /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             836..847
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             863..1573
                     /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    1580..1600
                     /label=BioBrick suffix
                     /note="universal suffix for all parts"
     terminator      1636..1667
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
     CDS             complement(1766..2581)
                     /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     2489..2508
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2573..2592
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             complement(2939..3889)
                     /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      3937..4159
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     terminator      complement(4651..4745)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     terminator      complement(4901..4930)
                     /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 atccgctagg atgcatataa atgtgagcgg ataacattga cattgtgagc
      781 ggataacaag atactgagca cagtggatat gctttacctc ttggtgagac gacataaaga
      841 ggagaaacat aatatgacta gccgtaaagg cgaagagctg ttcactggtg tcgtccctat
      901 tctggtggaa ctggatggtg atgtcaacgg tcataagttt tccgtgcgtg gcgagggtga
      961 aggtgacgca actaatggta aactgacgct gaagttcatc tgtactactg gtaaactgcc
     1021 ggtaccttgg ccgactctgg taacgacgct gacttatggt gttcagtgct ttgctcgtta
     1081 tccggaccat atgaagcagc atgacttctt caagtccgcc atgccggaag gctatgtgca
     1141 ggaacgcacg atttccttta aggatgacgg cacgtacaaa acgcgtgcgg aagtgaaatt
     1201 tgaaggcgat accctggtaa accgcattga gctgaaaggc attgacttta aagaagacgg
     1261 caatatcctg ggccataagc tggaatacaa ttttaacagc cacaatgttt acatcaccgc
     1321 cgataaacaa aaaaatggca ttaaagcgaa ttttaaaatt cgccacaacg tggaggatgg
     1381 cagcgtgcag ctggctgatc actaccagca aaacactcca atcggtgatg gtcctgttct
     1441 gctgccagac aatcactatc tgagcacgca aagcgttctg tctaaagatc cgaacgagaa
     1501 acgcgatcat atggttctgc tggagttcgt aaccgcagcg ggcatcacgc atggtatgga
     1561 tgaactgtac aaatgatgat actagtagcg gccgctgcag gagtcactaa gggttagtta
     1621 gttagattag cagaaagtca aaagcctccg accggaggct tttgactaaa acttcccttg
     1681 gggttatcat tggggctcac tcaaaggcgg taatcagata aaaaaaatcc ttagctttcg
     1741 ctaaggatga tttctgctag tattattaga aaaactcatc gagcatcaaa tgaaactgca
     1801 atttattcat atcaggatta tcaataccat atttttgaaa aagccgtttc tgtaatgaag
     1861 gagaaaactc accgaggcag ttccaaagaa tggcaaggtc ctggtaacgg tctgcgattc
     1921 cgacccgtcc aacatcaata caacctatta atttcccctc gtcaaaaata aggttatcaa
     1981 gtgagaaatc accatgagtg acgactgaat ccggtgagaa tggcaagagc ttgtgcattt
     2041 ctttccagac ttgttcaaca ggccagccat tacgctcgtc atcaaaatca ctcgcatcaa
     2101 ccaaaccgtt attcatgcgt gattgcgcct gagcaagacg aaatacacga tcgctgttaa
     2161 aaggacaatt acaaacagga atcgaatgta accggcgcag gaacacggcc agcgcatcaa
     2221 caatattttc acctgaatca ggatattctt ctaatacctg gaaggctgtt ttcccaggaa
     2281 tcgcggtggt gagtaaccac gcatcatcag gagtacggat aaaatgcttg atggtcggga
     2341 gaggcataaa ctccgtcagc cagttgagac ggaccatctc atctgtaaca tcattggcaa
     2401 cgctaccttt gccatgtttc agaaacaact ctggcgcatc gggcttccca tacaagcgat
     2461 agattgtcgc acctgattgc ccgacattat cgcgagccca tttataccca tataaatcag
     2521 cgtccatgtt ggagtttaag cgcggacggg agcaagacgt ttcccgttga atatggctca
     2581 taacacccct tgtattactg tttatgtaag cagacagttt tattgttcat gatgatatat
     2641 ttttatcttg tgcaatgtaa catcagagat tttgagacac aacgtggctt tgttgaataa
     2701 atcgaacttt tgctgagttg aaggatcagc tctagtagtt acattgtcga tctgttcatg
     2761 gtgaacagct ttaaatgcac caaaaactcg taaaagctct gatgtatcta tcttttttac
     2821 accgttttca tctgtgcata tggacagttt tccctttgat atctaacggt gaacagttgt
     2881 tctacttttg tttgttagtc ttgatgcttc actgatagat acaagagcca taagaacctc
     2941 agatccttcc gtatttagcc agtatgttct ctagtgtggt tcgttgtttt tgcgtgagcc
     3001 atgagaacga accattgaga tcatgcttac tttgcatgtc actcaaaaat tttgcctcaa
     3061 aactggtgag ctgaattttt gcagttaaag catcgtgtag tgtttttctt agtccgttac
     3121 gtaggtagga atctgatgta atggttgttg gtattttgtc accattcatt tttatctggt
     3181 tgttctcaag ttcggttacg agatccattt gtctatctag ttcaacttgg aaaatcaacg
     3241 tatcagtcgg gcggcctcgc ttatcaacca ccaatttcat attgctgtaa gtgtttaaat
     3301 ctttacttat tggtttcaaa acccattggt taagcctttt aaactcatgg tagttatttt
     3361 caagcattaa catgaactta aattcatcaa ggctaatctc tatatttgcc ttgtgagttt
     3421 tcttttgtgt tagttctttt aataaccact cataaatcct catagagtat ttgttttcaa
     3481 aagacttaac atgttccaga ttatatttta tgaatttttt taactggaaa agataaggca
     3541 atatctcttc actaaaaact aattctaatt tttcgcttga gaacttggca tagtttgtcc
     3601 actggaaaat ctcaaagcct ttaaccaaag gattcctgat ttccacagtt ctcgtcatca
     3661 gctctctggt tgctttagct aatacaccat aagcattttc cctactgatg ttcatcatct
     3721 gagcgtattg gttataagtg aacgataccg tccgttcttt ccttgtaggg ttttcaatcg
     3781 tggggttgag tagtgccaca cagcataaaa ttagcttggt ttcatgctcc gttaagtcat
     3841 agcgactaat cgctagttca tttgctttga aaacaactaa ttcagacata catctcaatt
     3901 ggtctaggtg attttaatca ctataccaat tgagatgggc tagtcaatga taattacatg
     3961 tccttttcct ttgagttgtg ggtatctgta aattctgcta gacctttgct ggaaaacttg
     4021 taaattctgc tagaccctct gtaaattccg ctagaccttt gtgtgttttt tttgtttata
     4081 ttcaagtggt tataatttat agaataaaga aagaataaaa aaagataaaa agaatagatc
     4141 ccagccctgt gtataactca ctactttagt cagttccgca gtattacaaa aggatgtcgc
     4201 aaacgctgtt tgctcctcta caaaacagac cttaaaaccc taaaggctta agtagcaccc
     4261 tcgcaagctc gggcaaatcg ctgaatattc cttttgtctc cgaccatcag gcacctgagt
     4321 cgctgtcttt ttcgtgacat tcagttcgct gcgctcacgg ctctggcagt gaatgggggt
     4381 aaatggcact acaggcgcct tttatggatt catgcaagga aactacccat aatacaagaa
     4441 aagcccgtca cgggcttctc agggcgtttt atggcgggtc tgctatgtgg tgctatctga
     4501 ctttttgctg ttcagcagtt cctgccctct gattttccag tctgaccact tcggattatc
     4561 ccgtgacagg tcattcagac tggctaatgc acccagtaag gcagcggtat catcaacagg
     4621 cttacccgtc ttactgtccc tagtgcttgg attctcacca ataaaaaacg cccggcggca
     4681 accgagcgtt ctgaacaaat ccagatggag ttctgaggtc attactggat ctatcaacag
     4741 gagtccaagc gagctcgtaa acttggtctg acagctctag ctccggcaaa aaaacgggca
     4801 aggtgtcacc accctgccct ttttctttaa aaccgaaaag attacttcgc gtttgccacc
     4861 tgacgtctaa gaaaaggaat attcagcaat ttgcccgtgc cgaagaaagg cccacccgtg
     4921 aaggtgagcc agtgagttga ttgctacgta attagttagt tagcccttag tgactc
//
