LOCUS       sequence-491693-        6324 bp    DNA     circular SYN 04-NOV-2025
DEFINITION  Express spAb-T7 RNAP that bind to FC region of human IgG.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6324)
  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 6324)
  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-spAb-T7 RNAP
FEATURES             Location/Qualifiers
     source          1..6324
                     /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      4307..4401
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             4445..5104
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(4511..4530)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      5169..5255
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     complement(5302..5324)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    5410..5550
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     5465..5484
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(5565..5582)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5736..6324)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5816..5835)
                     /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 cggatccgcg
     1381 gataacaaat ttaacaaaga acagcagaac gcgttttatg aaattctgca tctgccgaac
     1441 ctgaacgaag aacagcgcaa cggctttatt cagagcctga aagatgatcc gagccagagc
     1501 gcgaacctgc tggcggaagc gaaaaaactg aacgatgcgc aggcgccgaa aggatccggg
     1561 ggcgggggat ccggaggcgg gggatccggt ggtgggggtt ctatgaacac gattaacatc
     1621 gctaagaacg acttctctga catcgaactg gctgctatcc cgttcaacac tctggctgac
     1681 cattacggtg agcgtttagc tcgcgaacag ttggcccttg agcatgagtc ttacgagatg
     1741 ggtgaagcac gcttccgcaa gatgtttgag cgtcaactta aagctggtga ggttgcggat
     1801 aacgctgccg ccaagcctct catcactacc ctactcccta agatgattgc acgcatcaac
     1861 gactggtttg aggaagtgaa agctaagcgc ggcaagcgcc cgacagcctt ccagttcctg
     1921 caagaaatca agccggaagc cgtagcgtac atcaccatta agaccactct ggcttgccta
     1981 accagtgctg acaatacaac cgttcaggct gtagcaagcg caatcggtcg ggccattgag
     2041 gacgaggctc gcttcggtcg tatccgtgac cttgaagcta agcacttcaa gaaaaacgtt
     2101 gaggaacaac tcaacaagcg cgtagggcac gtctacaaga aagcatttat gcaagttgtc
     2161 gaggctgaca tgctctctaa gggtctactc ggtggcgagg cgtggtcttc gtggcataag
     2221 gaagactcta ttcatgtagg agtacgctgc atcgagatgc tcattgagtc aaccggaatg
     2281 gttagcttac accgccaaaa tgctggcgta gtaggtcaag actctgagac tatcgaactc
     2341 gcacctgaat acgctgaggc tatcgcaacc cgtgcaggtg cgctggctgg catctctccg
     2401 atgttccaac cttgcgtagt tcctcctaag ccgtggactg gcattactgg tggtggctat
     2461 tgggctaacg gtcgtcgtcc tctggcgctg gtgcgtactc acagtaagaa agcactgatg
     2521 cgctacgaag acgtttacat gcctgaggtg tacaaagcga ttaacattgc gcaaaacacc
     2581 gcatggaaaa tcaacaagaa agtcctagcg gtcgccaacg taatcaccaa gtggaagcat
     2641 tgtccggtcg aggacatccc tgcgattgag cgtgaagaac tcccgatgaa accggaagac
     2701 atcgacatga atcctgaggc tctcaccgcg tggaaacgtg ctgccgctgc tgtgtaccgc
     2761 aaggacaagg ctcgcaagtc tcgccgtatc agccttgagt tcatgcttga gcaagccaat
     2821 aagtttgcta accataaggc catctggttc ccttacaaca tggactggcg cggtcgtgtt
     2881 tacgctgtgt caatgttcaa cccgcaaggt aacgatatga ccaaaggact gcttacgctg
     2941 gcgaaaggta aaccaatcgg taaggaaggt tactactggc tgaaaatcca cggtgcaaac
     3001 tgtgcgggtg tcgataaggt tccgttccct gagcgcatca agttcattga ggaaaaccac
     3061 gagaacatca tggcttgcgc taagtctcca ctggagaaca cttggtgggc tgagcaagat
     3121 tctccgttct gcttccttgc gttctgcttt gagtacgctg gggtacagca ccacggcctg
     3181 agctataact gctcccttcc gctggcgttt gacgggtctt gctctggcat ccagcacttc
     3241 tccgcgatgc tccgagatga ggtaggtggt cgcgcggtta acttgcttcc tagtgaaacc
     3301 gttcaggaca tctacgggat tgttgctaag aaagtcaacg agattctaca agcagacgca
     3361 atcaatggga ccgataacga agtagttacc gtgaccgatg agaacactgg tgaaatctct
     3421 gagaaagtca agctgggcac taaggcactg gctggtcaat ggctggctta cggtgttact
     3481 cgcagtgtga ctaagcgttc agtcatgacg ctggcttacg ggtccaaaga gttcggcttc
     3541 cgtcaacaag tgctggaaga taccattcag ccagctattg attccggcaa gggtctgatg
     3601 ttcactcagc cgaatcaggc tgctggatac atggctaagc tgatttggga atctgtgagc
     3661 gtgacggtgg tagctgcggt tgaagcaatg aactggctta agtctgctgc taagctgctg
     3721 gctgctgagg tcaaagataa gaagactgga gagattcttc gcaagcgttg cgctgtgcat
     3781 tgggtaactc ctgatggttt ccctgtgtgg caggaataca agaagcctat tcagacgcgc
     3841 ttgaacctga tgttcctcgg tcagttccgc ttacagccta ccattaacac caacaaagat
     3901 agcgagattg atgcacacaa acaggagtct ggtatcgctc ctaactttgt acacagccaa
     3961 gacggtagcc accttcgtaa gactgtagtg tgggcacacg agaagtacgg aatcgaatct
     4021 tttgcactga ttcacgactc cttcggtacc attccggctg acgctgcgaa cctgttcaaa
     4081 gcagtgcgcg aaactatggt tgacacatat gagtcttgtg atgtactggc tgatttctac
     4141 gaccagttcg ctgaccagtt gcacgagtct caattggaca aaatgccagc acttccggct
     4201 aaaggtaact tgaacctccg tgacatctta gagtcggact tcgcgttcgc gtaactgcag
     4261 gcatcgcgtt cgcgtaactg cagccaagct taattagctg agcttggact cctgttgata
     4321 gatccagtaa tgacctcaga actccatctg gatttgttca gaacgctcgg ttgccgccgg
     4381 gcgtttttta ttggtgagaa tccaagctag cttggcgaga ttttcaggag ctaaggaagc
     4441 taaaatggag aaaaaaatca ctggatatac caccgttgat atatcccaat ggcatcgtaa
     4501 agaacatttt gaggcatttc agtcagttgc tcaatgtacc tataaccaga ccgttcagct
     4561 ggatattacg gcctttttaa agaccgtaaa gaaaaataag cacaagtttt atccggcctt
     4621 tattcacatt cttgcccgcc tgatgaatgc tcatccggaa tttcgtatgg caatgaaaga
     4681 cggtgagctg gtgatatggg atagtgttca cccttgttac accgttttcc atgagcaaac
     4741 tgaaacgttt tcatcgctct ggagtgaata ccacgacgat ttccggcagt ttctacacat
     4801 atattcgcaa gatgtggcgt gttacggtga aaacctggcc tatttcccta aagggtttat
     4861 tgagaatatg tttttcgtct cagccaatcc ctgggtgagt ttcaccagtt ttgatttaaa
     4921 cgtggccaat atggacaact tcttcgcccc cgttttcacc atgggcaaat attatacgca
     4981 aggcgacaag gtgctgatgc cgctggcgat tcaggttcat catgccgttt gtgatggctt
     5041 ccatgtcggc agaatgctta atgaattaca acagtactgc gatgagtggc agggcggggc
     5101 gtaatttttt taaggcagtt attggtgccc ttaaacgcct ggggtaatga ctctctagct
     5161 tgaggcatca aataaaacga aaggctcagt cgaaagactg ggcctttcgt tttatctgtt
     5221 gtttgtcggt gaacgctctc ctgagtagga caaatccgcc ctctagagct gcctcgcgcg
     5281 tttcggtgat gacggtgaaa acctctgaca catgcagctc ccggagacgg tcacagcttg
     5341 tctgtaagcg gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg
     5401 gtgtcggggc gcagccatga cccagtcacg tagcgatagc ggagtgtata ctggcttaac
     5461 tatgcggcat cagagcagat tgtactgaga gtgcaccata tgcggtgtga aataccgcac
     5521 agatgcgtaa ggagaaaata ccgcatcagg cgctcttccg cttcctcgct cactgactcg
     5581 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
     5641 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     5701 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     5761 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     5821 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     5881 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     5941 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     6001 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     6061 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     6121 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaaggaca
     6181 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     6241 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     6301 acgcgcagaa aaaaaggatc tcaa
//
