LOCUS       sequence-491694-        6193 bp    DNA     circular SYN 04-NOV-2025
DEFINITION  Express zf438-NB23 that bind to SARS-CoV-19 Spike protein 
            cooperatively with NB15-T7 RNAP..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6193)
  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 6193)
  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: pET-21(+)-His-zf438-NB23
FEATURES             Location/Qualifiers
     source          1..6193
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             66..83
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      815..862
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             869..886
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(939..957)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      953..1000
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1037..1492
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1124..1143)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1334..1355
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1518..1622
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1623..1691,1692..2483)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(1841..1860)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2654..3242
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3143..3162
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3396..3413
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3428..3567
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3493..3512)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3653..3675
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3669..3860)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4669..5751)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     5713..5732
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5752..5829)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6035..6054
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     6138..6157
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        6138..6156
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    6157..6181
                     /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)."
ORIGIN
        1 ggagaccaca acggtttccc tctagaaata attttgttta acttaagaag gaggaaaaaa
       61 aaatgcatca ccaccatcat cacggcggat tccagtgtcg catttgcatg cggaactttt
      121 cgcgccagga caggcttgac aggcataccc gtactcatac cggtgaaaaa ccgtttcagt
      181 gtcggatctg tatgcgaaat ttctcccaga aggagcactt ggcggggcat ctacgtacgc
      241 acaccggcga gaagccattc caatgccgaa tatgcatgcg caacttcagt cgccgcgaca
      301 acctgaaccg gcacctaaaa acccacggat ccggaggggg agcgcaggtg catctggtgg
      361 aaagcggcgg cgatctggtg cagccgggcg gcagcctgcg cctgagctgc gtggcgagcg
      421 gcagcggctt tgaaaacaac gcgattacct ggtatcgcca ggcgccgggc aaagaacgcg
      481 aactggtgag cggcattacc agcggcggca gcaccaacta tgcggatagc gtgaaaggcc
      541 gctttaccat tagccgcgat aacgcgaaaa acaccgtgta tctggaaatg aacagcctga
      601 aaccggaaga taccgcggtg tatctgtgcc aggcggtggc gtgggatagc cgccgccgca
      661 gcgtggtggc gttttggggc cagggcaccc aggtgaccgt gagctaataa cgactcaggc
      721 tgctacgcct gtgtactgga aaacaaaacc aaaacccaaa aaacaaaaaa ctgagcccat
      781 tggtatcgtg gaaggactct atcaaaaaaa aaaactagca taaccccttg gggcctctaa
      841 acgggtcttg aggggttttt tgctcgagca ccaccaccac caccactgag atccggctgc
      901 taacaaagcc cgaaaggaag ctgagttggc tgctgccacc gctgagcaat aactagcata
      961 accccttggg gcctctaaac gggtcttgag gggttttttg ctgaaaggag gaactatatc
     1021 cggattggcg aatgggacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt
     1081 acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc
     1141 ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct
     1201 ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat
     1261 ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc
     1321 acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc
     1381 tattcttttg atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg
     1441 atttaacaaa aatttaacgc gaattttaac aaaatattaa cgcttacaat ttaggtggca
     1501 cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata
     1561 tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga
     1621 gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc
     1681 ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg
     1741 cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc
     1801 ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat
     1861 cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact
     1921 tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat
     1981 tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga
     2041 tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc
     2101 ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga
     2161 tgcctgcagc aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag
     2221 cttcccggca acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc
     2281 gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt
     2341 ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct
     2401 acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg
     2461 cctcactgat taagcattgg taactgtcag accaagttta ctcatatata ctttagattg
     2521 atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca
     2581 tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga
     2641 tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa
     2701 aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga
     2761 aggtaactgg cttcagcaga gcgcagatac caaatactgt ccttctagtg tagccgtagt
     2821 taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt
     2881 taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat
     2941 agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct
     3001 tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca
     3061 cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag
     3121 agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc
     3181 gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga
     3241 aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca
     3301 tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc tttgagtgag
     3361 ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg
     3421 aagagcgcct gatgcggtat tttctcctta cgcatctgtg cggtatttca caccgcaatg
     3481 gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagtata cactccgcta
     3541 tcgctacgtg actgggtcat ggctgcgccc cgacacccgc caacacccgc tgacgcgccc
     3601 tgacgggctt gtctgctccc ggcatccgct tacagacaag ctgtgaccgt ctccgggagc
     3661 tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg cgaggcagct gcggtaaagc
     3721 tcatcagcgt ggtcgtgaag cgattcacag atgtctgcct gttcatccgc gtccagctcg
     3781 ttgagtttct ccagaagcgt taatgtctgg cttctgataa agcgggccat gttaagggcg
     3841 gttttttcct gtttggtcac tgatgcctcc gtgtaagggg gatttctgtt catgggggta
     3901 atgataccga tgaaacgaga gaggatgctc acgatacggg ttactgatga tgaacatgcc
     3961 cggttactgg aacgttgtga gggtaaacaa ctggcggtat ggatgcggcg ggaccagaga
     4021 aaaatcactc agggtcaatg ccagcgcttc gttaatacag atgtaggtgt tccacagggt
     4081 agccagcagc atcctgcgat gcagatccgg aacataatgg tgcagggcgc tgacttccgc
     4141 gtttccagac tttacgaaac acggaaaccg aagaccattc atgttgttgc tcaggtcgca
     4201 gacgttttgc agcagcagtc gcttcacgtt cgctcgcgta tcggtgattc attctgctaa
     4261 ccagtaaggc aaccccgcca gcctagccgg gtcctcaacg acaggagcac gatcatgcgc
     4321 acccgtgggg ccgccatgcc ggcgataatg gcctgcttct cgccgaaacg tttggtggcg
     4381 ggaccagtga cgaaggcttg agcgagggcg tgcaagattc cgaataccgc aagcgacagg
     4441 ccgatcatcg tcgcgctcca gcgaaagcgg tcctcgccga aaatgaccca gagcgctgcc
     4501 ggcacctgtc ctacgagttg catgataaag aagacagtca taagtgcggc gacgatagtc
     4561 atgccccgcg cccaccggaa ggagctgact gggttgaagg ctctcaaggg catcggtcga
     4621 gatcccggtg cctaatgagt gagctaactt acattaattg cgttgcgctc actgcccgct
     4681 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga
     4741 ggcggtttgc gtattgggcg ccagggtggt ttttcttttc accagtgaga cgggcaacag
     4801 ctgattgccc ttcaccgcct ggccctgaga gagttgcagc aagcggtcca cgctggtttg
     4861 ccccagcagg cgaaaatcct gtttgatggt ggttaacggc gggatataac atgagctgtc
     4921 ttcggtatcg tcgtatccca ctaccgagat atccgcacca acgcgcagcc cggactcggt
     4981 aatggcgcgc attgcgccca gcgccatctg atcgttggca accagcatcg cagtgggaac
     5041 gatgccctca ttcagcattt gcatggtttg ttgaaaaccg gacatggcac tccagtcgcc
     5101 ttcccgttcc gctatcggct gaatttgatt gcgagtgaga tatttatgcc agccagccag
     5161 acgcagacgc gccgagacag aacttaatgg gcccgctaac agcgcgattt gctggtgacc
     5221 caatgcgacc agatgctcca cgcccagtcg cgtaccgtct tcatgggaga aaataatact
     5281 gttgatgggt gtctggtcag agacatcaag aaataacgcc ggaacattag tgcaggcagc
     5341 ttccacagca atggcatcct ggtcatccag cggatagtta atgatcagcc cactgacgcg
     5401 ttgcgcgaga agattgtgca ccgccgcttt acaggcttcg acgccgcttc gttctaccat
     5461 cgacaccacc acgctggcac ccagttgatc ggcgcgagat ttaatcgccg cgacaatttg
     5521 cgacggcgcg tgcagggcca gactggaggt ggcaacgcca atcagcaacg actgtttgcc
     5581 cgccagttgt tgtgccacgc ggttgggaat gtaattcagc tccgccatcg ccgcttccac
     5641 tttttcccgc gttttcgcag aaacgtggct ggcctggttc accacgcggg aaacggtctg
     5701 ataagagaca ccggcatact ctgcgacatc gtataacgtt actggtttca cattcaccac
     5761 cctgaattga ctctcttccg ggcgctatca tgccataccg cgaaaggttt tgcgccattc
     5821 gatggtgtcc gggatctcga cgctctccct tatgcgactc ctgcattagg aagcagccca
     5881 gtagtaggtt gaggccgttg agcaccgccg ccgcaaggaa tggtgcatgc aaggagatgg
     5941 cgcccaacag tcccccggcc acggggcctg ccaccatacc cacgccgaaa caagcgctca
     6001 tgagcccgaa gtggcgagcc cgatcttccc catcggtgat gtcggcgata taggcgccag
     6061 caaccgcacc tgtggcgccg gtgatgccgg ccacgatgcg tccggcgtag aggatcgaga
     6121 tctcgatccc gcgaaattaa tacgactcac tataggggaa ttgtgagcgg ataacaattc
     6181 ccctctagga tcc
//
