LOCUS       sequence-124227-        4296 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses cisRAJ31-GFP-LAA, a cis-repressed mRNA regulated by 
            RAJ31..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4296)
  AUTHORS   Rostain W, Shen S, Cordero T, Rodrigo G, Jaramillo A
  TITLE     Engineering a Circular Riboregulator in Escherichia coli.
  JOURNAL   Biodes Res. 2020 Sep 12;2020:1916789. doi: 10.34133/2020/1916789. 
            eCollection 2020.
  PUBMED    37849901
REFERENCE   2  (bases 1 to 4296)
  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: "Biodes 
            Res."; date: "2020-09-12"; volume: "2020:1916789. doi"; pages: " 
            10.34133/2020/1916789. eCollection 2020"
COMMENT     Sequence Label: cisRAJ31-GFP-LAA
FEATURES             Location/Qualifiers
     source          1..4296
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(127..156)
                     /label=T3Te terminator
                     /note="phage T3 early transcription terminator"
     misc_feature    203..222
                     /label=BioBrick prefix
                     /note="BioBrick prefix for parts that start with ""ATG"""
     protein_bind    256..272
                     /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             319..330
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             346..1056
                     /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    1102..1122
                     /label=BioBrick suffix
                     /note="universal suffix for all parts"
     terminator      1158..1189
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
     CDS             complement(1288..2103)
                     /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     2011..2030
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2095..2114
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             complement(2461..3411)
                     /codon_start=1
                     /gene="rep101"
                     /product="RepA protein needed for replication with the 
                     pSC101 origin"
                     /label=Rep101
                     /translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
                     RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFGIFQWTNYAKFS
                     SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
                     EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
                     ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLHNGLRKTLHDALTAKIQLTSFEAKF
                     LSDMQSKYDLNGSFSWLTQKQRTTLENILAKYGRI"
     rep_origin      3459..3681
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     terminator      complement(4173..4267)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
ORIGIN
        1 ctctagctcc ggcaaaaaaa cgggcaaggt gtcaccaccc tgcccttttt ctttaaaacc
       61 gaaaagatta cttcgcgttt gccacctgac gtctaagaaa aggaatattc agcaatttgc
      121 ccgtgccgaa gaaaggccca cccgtgaagg tgagccagtg agttgattgc tacgtaatta
      181 gttagttagc ccttagtgac tcgaattcgc ggccgcttct aggatgcata taaatgtgag
      241 cggataacat tgacattgtg agcggataac aagatactga gcacagtgga tatgctttac
      301 ctcttggtga gacgacataa agaggagaaa cataatatga ctagccgtaa aggcgaagag
      361 ctgttcactg gtgtcgtccc tattctggtg gaactggatg gtgatgtcaa cggtcataag
      421 ttttccgtgc gtggcgaggg tgaaggtgac gcaactaatg gtaaactgac gctgaagttc
      481 atctgtacta ctggtaaact gccggtacct tggccgactc tggtaacgac gctgacttat
      541 ggtgttcagt gctttgctcg ttatccggac catatgaagc agcatgactt cttcaagtcc
      601 gccatgccgg aaggctatgt gcaggaacgc acgatttcct ttaaggatga cggcacgtac
      661 aaaacgcgtg cggaagtgaa atttgaaggc gataccctgg taaaccgcat tgagctgaaa
      721 ggcattgact ttaaagaaga cggcaatatc ctgggccata agctggaata caattttaac
      781 agccacaatg tttacatcac cgccgataaa caaaaaaatg gcattaaagc gaattttaaa
      841 attcgccaca acgtggagga tggcagcgtg cagctggctg atcactacca gcaaaacact
      901 ccaatcggtg atggtcctgt tctgctgcca gacaatcact atctgagcac gcaaagcgtt
      961 ctgtctaaag atccgaacga gaaacgcgat catatggttc tgctggagtt cgtaaccgca
     1021 gcgggcatca cgcatggtat ggatgaactg tacaaagcta gcgcagcgaa cgacgaaaat
     1081 tacgcccttg cagcgtaata atactagtag cggccgctgc aggagtcact aagggttagt
     1141 tagttagatt agcagaaagt caaaagcctc cgaccggagg cttttgacta aaacttccct
     1201 tggggttatc attggggctc actcaaaggc ggtaatcaga taaaaaaaat ccttagcttt
     1261 cgctaaggat gatttctgct agtattatta gaaaaactca tcgagcatca aatgaaactg
     1321 caatttattc atatcaggat tatcaatacc atatttttga aaaagccgtt tctgtaatga
     1381 aggagaaaac tcaccgaggc agttccaaag aatggcaagg tcctggtaac ggtctgcgat
     1441 tccgacccgt ccaacatcaa tacaacctat taatttcccc tcgtcaaaaa taaggttatc
     1501 aagtgagaaa tcaccatgag tgacgactga atccggtgag aatggcaaga gcttgtgcat
     1561 ttctttccag acttgttcaa caggccagcc attacgctcg tcatcaaaat cactcgcatc
     1621 aaccaaaccg ttattcatgc gtgattgcgc ctgagcaaga cgaaatacac gatcgctgtt
     1681 aaaaggacaa ttacaaacag gaatcgaatg taaccggcgc aggaacacgg ccagcgcatc
     1741 aacaatattt tcacctgaat caggatattc ttctaatacc tggaaggctg ttttcccagg
     1801 aatcgcggtg gtgagtaacc acgcatcatc aggagtacgg ataaaatgct tgatggtcgg
     1861 gagaggcata aactccgtca gccagttgag acggaccatc tcatctgtaa catcattggc
     1921 aacgctacct ttgccatgtt tcagaaacaa ctctggcgca tcgggcttcc catacaagcg
     1981 atagattgtc gcacctgatt gcccgacatt atcgcgagcc catttatacc catataaatc
     2041 agcgtccatg ttggagttta agcgcggacg ggagcaagac gtttcccgtt gaatatggct
     2101 cataacaccc cttgtattac tgtttatgta agcagacagt tttattgttc atgatgatat
     2161 atttttatct tgtgcaatgt aacatcagag attttgagac acaacgtggc tttgttgaat
     2221 aaatcgaact tttgctgagt tgaaggatca gctctagtag ttacattgtc gatctgttca
     2281 tggtgaacag ctttgaatgc accaaaaact cgtaaaagct ctgatgtatc tatctttttt
     2341 acaccgtttt catctgtgca tatggacagt tttccctttg atatgtaacg gtgaacagtt
     2401 gttctacttt tgtttgttag tcttgatgct tcactgatag atacaagagc cataagaacc
     2461 tcagatcctt ccgtatttag ccagtatgtt ctctagtgtg gttcgttgtt tttgcgtgag
     2521 ccatgagaac gaaccattga gatcatactt actttgcatg tcactcaaaa attttgcctc
     2581 aaaactggtg agctgaattt ttgcagttaa agcatcgtgt agtgtttttc ttagtccgtt
     2641 atgtaggtag gaatctgatg taatggttgt tggtattttg tcaccattca tttttatctg
     2701 gttgttctca agttcggtta cgagatccat ttgtctatct agttcaactt ggaaaatcaa
     2761 cgtatcagtc gggcggcctc gcttatcaac caccaatttc atattgctgt aagtgtttaa
     2821 atctttactt attggtttca aaacccattg gttaagcctt ttaaactcat ggtagttatt
     2881 ttcaagcatt aacatgaact taaattcatc aaggctaatc tctatatttg ccttgtgagt
     2941 tttcttttgt gttagttctt ttaataacca ctcataaatc ctcatagagt atttgttttc
     3001 aaaagactta acatgttcca gattatattt tatgaatttt tttaactgga aaagataagg
     3061 caatatctct tcactaaaaa ctaattctaa tttttcgctt gagaacttgg catagtttgt
     3121 ccactggaaa atcccaaagc ctttaaccaa aggattcctg atttccacag ttctcgtcat
     3181 cagctctctg gttgctttag ctaatacacc ataagcattt tccctactga tgttcatcat
     3241 ctgagcgtat tggttataag tgaacgatac cgtccgttct ttccttgtag ggttttcaat
     3301 cgtggggttg agtagtgcca cacagcataa aattagcttg gtttcatgct ccgttaagtc
     3361 atagcgacta atcgctagtt catttgcttt gaaaacaact aattcagaca tacatctcaa
     3421 ttggtctagg tgattttaat cactatacca attgagatgg gctagtcaat gataattaca
     3481 tgtccttttc ctttgagttg tgggtatctg taaattctgc tagacctttg ctggaaaact
     3541 tgtaaattct gctagaccct ctgtaaattc cgctagacct ttgtgtgttt tttttgttta
     3601 tattcaagtg gttataattt atagaataaa gaaagaataa aaaaagataa aaagaataga
     3661 tcccagccct gtgtataact cactacttta gtcagttccg cagtattaca aaaggatgtc
     3721 gcaaacgctg tttgctcctc tacaaaacag accttaaaac cctaaaggct taagtagcac
     3781 cctcgcaagc tcgggcaaat cgctgaatat tccttttgtc tccgaccatc aggcacctga
     3841 gtcgctgtct ttttcgtgac attcagttcg ctgcgctcac ggctctggca gtgaatgggg
     3901 gtaaatggca ctacaggcgc cttttatgga ttcatgcaag gaaactaccc ataatacaag
     3961 aaaagcccgt cacgggcttc tcagggcgtt ttatggcggg tctgctatgt ggtgctatct
     4021 gactttttgc tgttcagcag ttcctgccct ctgattttcc agtctgacca cttcggatta
     4081 tcccgtgaca ggtcattcag actggctaat gcacccagta aggcagcggt atcatcaaca
     4141 ggcttacccg tcttactgtc cctagtgctt ggattctcac caataaaaaa cgcccggcgg
     4201 caaccgagcg ttctgaacaa atccagatgg agttctgagg tcattactgg atctatcaac
     4261 aggagtccaa gcgagctcgt aaacttggtc tgacag
//
