LOCUS       sequence-74503-e        4343 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 4343)
  AUTHORS   Rhodius VA, Segall-Shapiro TH, Sharon BD, Ghodasara A, Orlova E, 
            Tabakh H, Burkhardt DH, Clancy K, Peterson TC, Gross CA, Voigt CA
  TITLE     Design of orthogonal genetic switches based on a crosstalk map of 
            sigmas, anti-sigmas, and promoters.
  JOURNAL   Mol Syst Biol. 2013 Oct 29;9:702. doi: 10.1038/msb.2013.58.
  PUBMED    24169405
REFERENCE   2  (bases 1 to 4343)
  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: "Mol Syst 
            Biol."; date: "2013-10-29"; volume: "9:702. doi"; pages: " 
            10.1038/msb.2013.58"
COMMENT     Sequence Label: pVRb11_3726
FEATURES             Location/Qualifiers
     source          1..4343
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             join(132..134,135..848)
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /note="mammalian codon-optimized"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     CDS             132..845
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Nager et al., 2011)"
                     /label=superfolder GFP
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(162..190)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     785..805
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     terminator      890..976
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(1511..2461)
                     /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      2509..2731
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     terminator      complement(3223..3317)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(3348..4142)
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3459..3478)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     4069..4088
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4321..4339
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgtcttcac ctcgaggcct ccacaccgct cgtcacatcc tgtgatccac tcttcatccc
       61 gctacgtaac acctctgcat cgcgaaccaa aaccagggat cctgtcaatt tccgcgatag
      121 aggaggtaaa gatgagcaaa ggagaagaac ttttcactgg agttgtccca attcttgttg
      181 aattagatgg tgatgttaat gggcacaaat tttctgtccg tggagagggt gaaggtgatg
      241 ctacaaacgg aaaactcacc cttaaattta tttgcactac tggaaaacta cctgttccgt
      301 ggccaacact tgtcactact ctgacctatg gtgttcaatg cttttcccgt tatccggatc
      361 acatgaaacg gcatgacttt ttcaagagtg ccatgcccga aggttatgta caggaacgca
      421 ctatatcttt caaagatgac gggacctaca agacgcgtgc tgaagtcaag tttgaaggtg
      481 atacccttgt taatcgtatc gagttaaagg gtattgattt taaagaagat ggaaacattc
      541 ttggacacaa actcgagtac aactttaact cacacaatgt atacatcacg gcagacaaac
      601 aaaagaatgg aatcaaagct aacttcaaaa ttcgccacaa cgttgaagat ggttccgttc
      661 aactagcaga ccattatcaa caaaatactc caattggcga tggccctgtc cttttaccag
      721 acaaccatta cctgtcgaca caatctgtcc tttcgaaaga tcccaacgaa aagcgtgacc
      781 acatggtcct tcttgagttt gtaactgctg ctgggattac acatggcatg gatgagctct
      841 acaaataata aatgtccaga cctgcaggca tgcaagcctc tagaggcatc aaataaaacg
      901 aaaggctcag tcgaaagact gggcctttcg ttttatctgt tgtttgtcgg tgaacgctct
      961 cctgagtagg acaaatccgc cgccctagac ctagggtacg ggttttgctg cccgcaaacg
     1021 ggctgttctg gtgttgctag tttgttatca gaatcgcaga tccggcttca ggtttgccgg
     1081 ctgaaagcgc tatttcttcc agaattgcca tgattttttc cccacgggag gcgtcactgg
     1141 ctcccgtgtt gtcggcagct ttgattcgat aagcagcatc gcctgtttca ggctgtctat
     1201 gtgtgactgt tgagctgtaa caagttgtct caggtgttca atttcatgtt ctagttgctt
     1261 tgttttactg gtttcacctg ttctattagg tgttacatgc tgttcatctg ttacattgtc
     1321 gatctgttca tggtgaacag ctttaaatgc accaaaaact cgtaaaagct ctgatgtatc
     1381 tatctttttt acaccgtttt catctgtgca tatggacagt tttccctttg atatctaacg
     1441 gtgaacagtt gttctacttt tgtttgttag tcttgatgct tcactgatag atacaagagc
     1501 cataagaacc tcagatcctt ccgtatttag ccagtatgtt ctctagtgtg gttcgttgtt
     1561 tttgcgtgag ccatgagaac gaaccattga gatcatgctt actttgcatg tcactcaaaa
     1621 attttgcctc aaaactggtg agctgaattt ttgcagttaa agcatcgtgt agtgtttttc
     1681 ttagtccgtt acgtaggtag gaatctgatg taatggttgt tggtattttg tcaccattca
     1741 tttttatctg gttgttctca agttcggtta cgagatccat ttgtctatct agttcaactt
     1801 ggaaaatcaa cgtatcagtc gggcggcctc gcttatcaac caccaatttc atattgctgt
     1861 aagtgtttaa atctttactt attggtttca aaacccattg gttaagcctt ttaaactcat
     1921 ggtagttatt ttcaagcatt aacatgaact taaattcatc aaggctaatc tctatatttg
     1981 ccttgtgagt tttcttttgt gttagttctt ttaataacca ctcataaatc ctcatagagt
     2041 atttgttttc aaaagactta acatgttcca gattatattt tatgaatttt tttaactgga
     2101 aaagataagg caatatctct tcactaaaaa ctaattctaa tttttcgctt gagaacttgg
     2161 catagtttgt ccactggaaa atctcaaagc ctttaaccaa aggattcctg atttccacag
     2221 ttctcgtcat cagctctctg gttgctttag ctaatacacc ataagcattt tccctactga
     2281 tgttcatcat ctgagcgtat tggttataag tgaacgatac cgtccgttct ttccttgtag
     2341 ggttttcaat cgtggggttg agtagtgcca cacagcataa aattagcttg gtttcatgct
     2401 ccgttaagtc atagcgacta atcgctagtt catttgcttt gaaaacaact aattcagaca
     2461 tacatctcaa ttggtctagg tgattttaat cactatacca attgagatgg gctagtcaat
     2521 gataattact agtccttttc ctttgagttg tgggtatctg taaattctgc tagacctttg
     2581 ctggaaaact tgtaaattct gctagaccct ctgtaaattc cgctagacct ttgtgtgttt
     2641 tttttgttta tattcaagtg gttataattt atagaataaa gaaagaataa aaaaagataa
     2701 aaagaataga tcccagccct gtgtataact cactacttta gtcagttccg cagtattaca
     2761 aaaggatgtc gcaaacgctg tttgctcctc tacaaaacag accttaaaac cctaaaggct
     2821 taagtagcac cctcgcaagc tcgggcaaat cgctgaatat tccttttgtc tccgaccatc
     2881 aggcacctga gtcgctgtct ttttcgtgac attcagttcg ctgcgctcac ggctctggca
     2941 gtgaatgggg gtaaatggca ctacaggcgc cttttatgga ttcatgcaag gaaactaccc
     3001 ataatacaag aaaagcccgt cacgggcttc tcagggcgtt ttatggcggg tctgctatgt
     3061 ggtgctatct gactttttgc tgttcagcag ttcctgccct ctgattttcc agtctgacca
     3121 cttcggatta tcccgtgaca ggtcattcag actggctaat gcacccagta aggcagcggt
     3181 atcatcaaca ggcttacccg tcttactgtc cctagtgctt ggattctcac caataaaaaa
     3241 cgcccggcgg caaccgagcg ttctgaacaa atccagatgg agttctgagg tcattactgg
     3301 atctatcaac aggagtccaa gcgagctctc gaaccccaga gtcccgctca gaagaactcg
     3361 tcaagaaggc gatagaaggc gatgcgctgc gaatcgggag cggcgatacc gtaaagcacg
     3421 aggaagcggt cagcccattc gccgccaagc tcttcagcaa tatcacgggt agccaacgct
     3481 atgtcctgat agcggtccgc cacacccagc cggccacagt cgatgaatcc agaaaagcgg
     3541 ccattttcca ccatgatatt cggcaagcag gcatcgccat gggtcacgac gagatcctcg
     3601 ccgtcgggca tgcgcgcctt gagcctggcg aacagttcgg ctggcgcgag cccctgatgc
     3661 tcttcgtcca gatcatcctg atcgacaaga ccggcttcca tccgagtacg tgctcgctcg
     3721 atgcgatgtt tcgcttggtg gtcgaatggg caggtagccg gatcaagcgt atgcagccgc
     3781 cgcattgcat cagccatgat ggatactttc tcggcaggag caaggtgaga tgacaggaga
     3841 tcctgccccg gcacttcgcc caatagcagc cagtcccttc ccgcttcagt gacaacgtcg
     3901 agcacagctg cgcaaggaac gcccgtcgtg gccagccacg atagccgcgc tgcctcgtcc
     3961 tgcagttcat tcagggcacc ggacaggtcg gtcttgacaa aaagaaccgg gcgcccctgc
     4021 gctgacagcc ggaacacggc ggcatcagag cagccgattg tctgttgtgc ccagtcatag
     4081 ccgaatagcc tctccaccca agcggccgga gaacctgcgt gcaatccatc ttgttcaatc
     4141 atgcgaaacg atcctcatcc tgtctcttga tcagatcttg atcccctgcg ccatcagatc
     4201 cttggcggca agaaagccat ccagtttact ttgcagggct tcccaacctt accagagggc
     4261 gccccagctg gcaattccga cgtctaagaa accattatta tcatgacatt aacctataaa
     4321 aataggcgta tcacgaggcc ctt
//
