LOCUS       sequence-491695-        6408 bp    DNA     circular SYN 04-NOV-2025
DEFINITION  The plasmid encode N-terminal his tagged zf438-T7 RNAP that pair 
            with pzfAAA-d1T7 promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6408)
  AUTHORS   Lee P-W, Mottaghi SS, Lugnier MG, Maerkl SJ
  TITLE     A T7 RNAP regulatory toolbox for cell-free network engineering and 
            biosensing applications
  JOURNAL   bioRxiv 2025.06.18.660355
REFERENCE   2  (bases 1 to 6408)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Nov 4, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "bioRxiv 
            2025.06.18.660355"
COMMENT     Sequence Label: pQE30-His-zfAAA-T7RNAP
FEATURES             Location/Qualifiers
     source          1..6408
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(171..962,963..1031))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     794..813
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1032..1136)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     1204..1222
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        join(1246..1260,1261..1266,1267..1283,1284..1290)
                     /label=T5 promoter
                     /note="bacteriophage T5 promoter for E. coli RNA 
                     polymerase, with embedded lac operator"
     protein_bind    1266..1282
                     /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)."
     protein_bind    1298..1314
                     /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             1333..1344
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             1354..1371
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      4391..4485
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             4529..5188
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(4595..4614)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      5253..5339
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     complement(5386..5408)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    5494..5634
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     5549..5568
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(5649..5666)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5820..6408)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5900..5919)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
       61 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
      121 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
      181 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
      241 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
      301 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
      361 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
      421 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
      481 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
      541 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
      601 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
      661 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
      721 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
      781 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
      841 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
      901 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
      961 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
     1021 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
     1081 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
     1141 tttccccgaa aagtgccacc tgacgtctaa gaaaccatta ttatcatgac attaacctat
     1201 aaaaataggc gtatcacgag gccctttcgt cttcacctcg agaaatcata aaaaatttat
     1261 ttgctttgtg agcggataac aattataata gattcaattg tgagcggata acaatttcac
     1321 acagaattca ttaaagagga gaaattaact atgcatcacc accatcatca catggaacgt
     1381 ccgtacgctt gcccggttga atcttgcgac cgtcgtttct ctcgttctga cgaactgacc
     1441 cgtcatatta gaattcatac tggacaaaaa ccattccaat gtagaatttg tatgagaaat
     1501 ttctctcgtt ctgaccacct gaccacccac atccgtaccc acaccggtga aaaaccgttc
     1561 gcttgcgaca tctgcggtcg taaattcgct cgttctgacg aacgtaaacg tcacaccaaa
     1621 atccacctgc gtcaggggtc cggaggcgga gggtccggag gcgggggatc cggtggtggt
     1681 ggttctatga acacgattaa catcgctaag aacgacttct ctgacatcga actggctgct
     1741 atcccgttca acactctggc tgaccattac ggtgagcgtt tagctcgcga acagttggcc
     1801 cttgagcatg agtcttacga gatgggtgaa gcacgcttcc gcaagatgtt tgagcgtcaa
     1861 cttaaagctg gtgaggttgc ggataacgct gccgccaagc ctctcatcac taccctactc
     1921 cctaagatga ttgcacgcat caacgactgg tttgaggaag tgaaagctaa gcgcggcaag
     1981 cgcccgacag ccttccagtt cctgcaagaa atcaagccgg aagccgtagc gtacatcacc
     2041 attaagacca ctctggcttg cctaaccagt gctgacaata caaccgttca ggctgtagca
     2101 agcgcaatcg gtcgggccat tgaggacgag gctcgcttcg gtcgtatccg tgaccttgaa
     2161 gctaagcact tcaagaaaaa cgttgaggaa caactcaaca agcgcgtagg gcacgtctac
     2221 aagaaagcat ttatgcaagt tgtcgaggct gacatgctct ctaagggtct actcggtggc
     2281 gaggcgtggt cttcgtggca taaggaagac tctattcatg taggagtacg ctgcatcgag
     2341 atgctcattg agtcaaccgg aatggttagc ttacaccgcc aaaatgctgg cgtagtaggt
     2401 caagactctg agactatcga actcgcacct gaatacgctg aggctatcgc aacccgtgca
     2461 ggtgcgctgg ctggcatctc tccgatgttc caaccttgcg tagttcctcc taagccgtgg
     2521 actggcatta ctggtggtgg ctattgggct aacggtcgtc gtcctctggc gctggtgcgt
     2581 actcacagta agaaagcact gatgcgctac gaagacgttt acatgcctga ggtgtacaaa
     2641 gcgattaaca ttgcgcaaaa caccgcatgg aaaatcaaca agaaagtcct agcggtcgcc
     2701 aacgtaatca ccaagtggaa gcattgtccg gtcgaggaca tccctgcgat tgagcgtgaa
     2761 gaactcccga tgaaaccgga agacatcgac atgaatcctg aggctctcac cgcgtggaaa
     2821 cgtgctgccg ctgctgtgta ccgcaaggac aaggctcgca agtctcgccg tatcagcctt
     2881 gagttcatgc ttgagcaagc caataagttt gctaaccata aggccatctg gttcccttac
     2941 aacatggact ggcgcggtcg tgtttacgct gtgtcaatgt tcaacccgca aggtaacgat
     3001 atgaccaaag gactgcttac gctggcgaaa ggtaaaccaa tcggtaagga aggttactac
     3061 tggctgaaaa tccacggtgc aaactgtgcg ggtgtcgata aggttccgtt ccctgagcgc
     3121 atcaagttca ttgaggaaaa ccacgagaac atcatggctt gcgctaagtc tccactggag
     3181 aacacttggt gggctgagca agattctccg ttctgcttcc ttgcgttctg ctttgagtac
     3241 gctggggtac agcaccacgg cctgagctat aactgctccc ttccgctggc gtttgacggg
     3301 tcttgctctg gcatccagca cttctccgcg atgctccgag atgaggtagg tggtcgcgcg
     3361 gttaacttgc ttcctagtga aaccgttcag gacatctacg ggattgttgc taagaaagtc
     3421 aacgagattc tacaagcaga cgcaatcaat gggaccgata acgaagtagt taccgtgacc
     3481 gatgagaaca ctggtgaaat ctctgagaaa gtcaagctgg gcactaaggc actggctggt
     3541 caatggctgg cttacggtgt tactcgcagt gtgactaagc gttcagtcat gacgctggct
     3601 tacgggtcca aagagttcgg cttccgtcaa caagtgctgg aagataccat tcagccagct
     3661 attgattccg gcaagggtct gatgttcact cagccgaatc aggctgctgg atacatggct
     3721 aagctgattt gggaatctgt gagcgtgacg gtggtagctg cggttgaagc aatgaactgg
     3781 cttaagtctg ctgctaagct gctggctgct gaggtcaaag ataagaagac tggagagatt
     3841 cttcgcaagc gttgcgctgt gcattgggta actcctgatg gtttccctgt gtggcaggaa
     3901 tacaagaagc ctattcagac gcgcttgaac ctgatgttcc tcggtcagtt ccgcttacag
     3961 cctaccatta acaccaacaa agatagcgag attgatgcac acaaacagga gtctggtatc
     4021 gctcctaact ttgtacacag ccaagacggt agccaccttc gtaagactgt agtgtgggca
     4081 cacgagaagt acggaatcga atcttttgca ctgattcacg actccttcgg taccattccg
     4141 gctgacgctg cgaacctgtt caaagcagtg cgcgaaacta tggttgacac atatgagtct
     4201 tgtgatgtac tggctgattt ctacgaccag ttcgctgacc agttgcacga gtctcaattg
     4261 gacaaaatgc cagcacttcc ggctaaaggt aacttgaacc tccgtgacat cttagagtcg
     4321 gacttcgcgt tcgcgtaact gcaggcatcg cgttcgcgta actgcagcca agcttaatta
     4381 gctgagcttg gactcctgtt gatagatcca gtaatgacct cagaactcca tctggatttg
     4441 ttcagaacgc tcggttgccg ccgggcgttt tttattggtg agaatccaag ctagcttggc
     4501 gagattttca ggagctaagg aagctaaaat ggagaaaaaa atcactggat ataccaccgt
     4561 tgatatatcc caatggcatc gtaaagaaca ttttgaggca tttcagtcag ttgctcaatg
     4621 tacctataac cagaccgttc agctggatat tacggccttt ttaaagaccg taaagaaaaa
     4681 taagcacaag ttttatccgg cctttattca cattcttgcc cgcctgatga atgctcatcc
     4741 ggaatttcgt atggcaatga aagacggtga gctggtgata tgggatagtg ttcacccttg
     4801 ttacaccgtt ttccatgagc aaactgaaac gttttcatcg ctctggagtg aataccacga
     4861 cgatttccgg cagtttctac acatatattc gcaagatgtg gcgtgttacg gtgaaaacct
     4921 ggcctatttc cctaaagggt ttattgagaa tatgtttttc gtctcagcca atccctgggt
     4981 gagtttcacc agttttgatt taaacgtggc caatatggac aacttcttcg cccccgtttt
     5041 caccatgggc aaatattata cgcaaggcga caaggtgctg atgccgctgg cgattcaggt
     5101 tcatcatgcc gtttgtgatg gcttccatgt cggcagaatg cttaatgaat tacaacagta
     5161 ctgcgatgag tggcagggcg gggcgtaatt tttttaaggc agttattggt gcccttaaac
     5221 gcctggggta atgactctct agcttgaggc atcaaataaa acgaaaggct cagtcgaaag
     5281 actgggcctt tcgttttatc tgttgtttgt cggtgaacgc tctcctgagt aggacaaatc
     5341 cgccctctag agctgcctcg cgcgtttcgg tgatgacggt gaaaacctct gacacatgca
     5401 gctcccggag acggtcacag cttgtctgta agcggatgcc gggagcagac aagcccgtca
     5461 gggcgcgtca gcgggtgttg gcgggtgtcg gggcgcagcc atgacccagt cacgtagcga
     5521 tagcggagtg tatactggct taactatgcg gcatcagagc agattgtact gagagtgcac
     5581 catatgcggt gtgaaatacc gcacagatgc gtaaggagaa aataccgcat caggcgctct
     5641 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca
     5701 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac
     5761 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt
     5821 ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg
     5881 cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc
     5941 tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc
     6001 gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc
     6061 aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac
     6121 tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt
     6181 aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct
     6241 aactacggct acactagaag gacagtattt ggtatctgcg ctctgctgaa gccagttacc
     6301 ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt
     6361 ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaa
//
