LOCUS       sequence-86407-e        4613 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Produces one half (beta) of aminoglycoside phosphotransferase 
            (3′)-IIa when in the presence of T7 RNA Polymerase..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4613)
  AUTHORS   Schmidt CM, Shis DL, Nguyen-Huu TD, Bennett MR
  TITLE     Stable maintenance of multiple plasmids in E. coli using a single 
            selective marker.
  JOURNAL   ACS Synth Biol. 2012 Oct 19;1(10):445-50. doi: 10.1021/sb3000589. 
            Epub 2012 Sep 4.
  PUBMED    23656183
REFERENCE   2  (bases 1 to 4613)
  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"; doi: "10.1021/sb3000589";
            journalName: "ACS Synth Biol"; date: "2012-10-19- 19"; volume: "1"; 
            issue: "10"; pages: "445-50"
COMMENT     Sequence Label: pCS5
FEATURES             Location/Qualifiers
     source          1..4613
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(10..15,16..32,33..38)
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     CDS             86..1276
                     /codon_start=1
                     /gene="tet"
                     /product="tetracycline efflux protein"
                     /label=TcR
                     /note="confers resistance to tetracycline"
                     /translation="MKSNNALIVILGTVTLDAVGIGLVMPVLPGLLRDIVHSDSIASHY
                     GVLLALYALMQFLCAPVLGALSDRFGRRPVLLASLLGATIDYAIMATTPVLWILYAGRI
                     VAGITGATGAVAGAYIADITDGEDRARHFGLMSACFGVGMVAGPVAGGLLGAISLHAPF
                     LAAAVLNGLNLLLGCFLMQESHKGERRPMPLRAFNPVSSFRWARGMTIVAALMTVFFIM
                     QLVGQVPAALWVIFGEDRFRWSATMIGLSLAVFGILHALAQAFVTGPATKRFGEKQAII
                     AGMAADALGYVLLAFATRGWMAFPIMILLASGGIGMPALQAMLSRQVDDDHQGQLQGSL
                     AALTSLTSIIGPLIVTAIYAASASTWNGLAWIVGAALYLVCLPALRRGAWSRATST"
     primer_bind     complement(438..457)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     CDS             1915..2106
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(2100..2122)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    2208..2348
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     2263..2282
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(2363..2380)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2534..3122)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2614..2633)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3539..3558
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3539..3557
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    3558..3582
                     /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             3597..3619
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     primer_bind     4227..4246
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     CDS             4404..4421
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(4474..4492)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      4488..4535
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     4578..4596
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 ttctcatgtt tgacagctta tcatcgataa gctttaatgc ggtagtttat cacagttaaa
       61 ttgctaacgc agtcaggcac cgtgtatgaa atctaacaat gcgctcatcg tcatcctcgg
      121 caccgtcacc ctggatgctg taggcatagg cttggttatg ccggtactgc cgggcctctt
      181 gcgggatatc gtccattccg acagcatcgc cagtcactat ggcgtgctgc tagcgctata
      241 tgcgttgatg caatttctat gcgcacccgt tctcggagca ctgtccgacc gctttggccg
      301 ccgcccagtc ctgctcgctt cgctacttgg agccactatc gactacgcga tcatggcgac
      361 cacacccgtc ctgtggatcc tctacgccgg acgcatcgtg gccggcatca ccggcgccac
      421 aggtgcggtt gctggcgcct atatcgccga catcaccgat ggggaagatc gggctcgcca
      481 cttcgggctc atgagcgctt gtttcggcgt gggtatggtg gcaggccccg tggccggggg
      541 actgttgggc gccatctcct tgcatgcacc attccttgcg gcggcggtgc tcaacggcct
      601 caacctacta ctgggctgct tcctaatgca ggagtcgcat aagggagagc gtcgaccgat
      661 gcccttgaga gccttcaacc cagtcagctc cttccggtgg gcgcggggca tgactatcgt
      721 cgccgcactt atgactgtct tctttatcat gcaactcgta ggacaggtgc cggcagcgct
      781 ctgggtcatt ttcggcgagg accgctttcg ctggagcgcg acgatgatcg gcctgtcgct
      841 tgcggtattc ggaatcttgc acgccctcgc tcaagccttc gtcactggtc ccgccaccaa
      901 acgtttcggc gagaagcagg ccattatcgc cggcatggcg gccgacgcgc tgggctacgt
      961 cttgctggcg ttcgcgacgc gaggctggat ggccttcccc attatgattc ttctcgcttc
     1021 cggcggcatc gggatgcccg cgttgcaggc catgctgtcc aggcaggtag atgacgacca
     1081 tcagggacag cttcaaggat cgctcgcggc tcttaccagc ctaacttcga tcattggacc
     1141 gctgatcgtc acggcgattt atgccgcctc ggcgagcaca tggaacgggt tggcatggat
     1201 tgtaggcgcc gccctatacc ttgtctgcct ccccgcgttg cgtcgcggtg catggagccg
     1261 ggccacctcg acctgaatgg aagccggcgg cacctcgcta acggattcac cactccaaga
     1321 attggagcca atcaattctt gcggagaact gtgaatgcgc aaaccaaccc ttggcagaac
     1381 atatccatcg cgtccgccat ctccagcagc cgcacgcggc gcatctcggg cagcgttggg
     1441 tcctggccac gggtgcgcat gatcgtgctc ctgtcgttga ggacccggct aggctggcgg
     1501 ggttgcctta ctggttagca gaatgaatca ccgatacgcg agcgaacgtg aagcgactgc
     1561 tgctgcaaaa cgtctgcgac ctgagcaaca acatgaatgg tcttcggttt ccgtgtttcg
     1621 taaagtctgg aaacgcggaa gtcagcgccc tgcaccatta tgttccggat ctgcatcgca
     1681 ggatgctgct ggctaccctg tggaacacct acatctgtat taacgaagcg ctggcattga
     1741 ccctgagtga tttttctctg gtcccgccgc atccataccg ccagttgttt accctcacaa
     1801 cgttccagta accgggcatg ttcatcatca gtaacccgta tcgtgagcat cctctctcgt
     1861 ttcatcggta tcattacccc catgaacaga aatccccctt acacggaggc atcagtgacc
     1921 aaacaggaaa aaaccgccct taacatggcc cgctttatca gaagccagac attaacgctt
     1981 ctggagaaac tcaacgagct ggacgcggat gaacaggcag acatctgtga atcgcttcac
     2041 gaccacgctg atgagcttta ccgcagctgc ctcgcgcgtt tcggtgatga cggtgaaaac
     2101 ctctgacaca tgcagctccc ggagacggtc acagcttgtc tgtaagcgga tgccgggagc
     2161 agacaagccc gtcagggcgc gtcagcgggt gttggcgggt gtcggggcgc agccatgacc
     2221 cagtcacgta gcgatagcgg agtgtatact ggcttaacta tgcggcatca gagcagattg
     2281 tactgagagt gcaccatatg cggtgtgaaa taccgcacag atgcgtaagg agaaaatacc
     2341 gcatcaggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
     2401 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
     2461 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     2521 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     2581 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     2641 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     2701 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     2761 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     2821 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     2881 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     2941 tgaagtggtg gcctaactac ggctacacta gaaggacagt atttggtatc tgcgctctgc
     3001 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     3061 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     3121 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     3181 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     3241 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     3301 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     3361 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
     3421 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
     3481 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     3541 atacgactca ctatagggga attgtgagcg gataacaatt cccctctaga aataattttg
     3601 tttaacttta agaaggagat ataccatggc tagcgcacag ctggagaaga aactgcaggc
     3661 tctggagaag aaactggcac agctggagtg gaaaaaccag gcactggaga agaaactggc
     3721 acagggtgga agcggggaac tgcaggacga ggcagcgcgg ctatcgtggc tggccacgac
     3781 gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg actggctgct
     3841 attgggcgaa gtgccggggc aggatctcct gtcatctcac cttgctcctg ccgagaaagt
     3901 atccatcatg gctgatgcaa tgcggcggct gcatacgctt gatccggcta cctgcccatt
     3961 cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact cggatggaag ccggtcttgt
     4021 cgatcaggat gatctggacg aagagcatca ggggctcgcg ccagccgaac tgttcgccag
     4081 gctcaaggcg agcatgcccg acggcgagga tctcgtcgtg acccacggcg atgcctgctt
     4141 gccgaatatc atggaggaaa atggccgctt ttctggattc atcgactgtg gccggctggg
     4201 tgtggcggac cgctatcagg acatagcgtt ggctacccgt gatattgccg aagagcttgg
     4261 cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc gccgctcccg attcgcagcg
     4321 catcgccttc tatcgccttc ttgacgagtt cttctaagga tccgaattcg agctccgtcg
     4381 acaagcttgc ggccgcactc gagcaccacc accaccacca ctgagatccg gctgctaaca
     4441 aagcccgaaa ggaagctgag ttggctgctg ccaccgctga gcaataacta gcataacccc
     4501 ttggggcctc taaacgggtc ttgaggggtt ttttggacgt ctaagaaacc attattatca
     4561 tgacattaac ctataaaaat aggcgtatca cgaggccctt tcgtcttcaa gaa
//
