LOCUS       sequence-244079-        5611 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  coding sequence for synthetic zinc finger ZF-ADD.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5611)
  AUTHORS   Swank Z, Laohakunakorn N, Maerkl SJ
  TITLE     Cell-free gene-regulatory network engineering with synthetic 
            transcription factors.
  JOURNAL   Proc Natl Acad Sci U S A. 2019 Mar 26;116(13):5892-5901. doi: 
            10.1073/pnas.1816591116. Epub 2019 Mar 8.
  PUBMED    30850530
REFERENCE   2  (bases 1 to 5611)
  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: 
            "10.1073/pnas.1816591116"; journalName: "Proc Natl Acad Sci U S A"; 
            date: "2019-03-26- 26"; volume: "116"; issue: "13"; pages: 
            "5892-5901"
COMMENT     Sequence Label: pZF-ADD
FEATURES             Location/Qualifiers
     source          1..5611
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     131..153
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(147..338)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(1147..2229)
                     /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     2191..2210
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(2230..2307)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     2513..2532
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     2620..2639
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2620..2638
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    2639..2663
                     /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             2678..2700
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     primer_bind     complement(3024..3042)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      3038..3085
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      3122..3577
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3209..3228)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3419..3440
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3604..3708
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3709..3777,3778..4569)
                     /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(3927..3946)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4740..5328
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5229..5248
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5482..5499
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    join(5514..5611,1..45)
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5582..5601)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 ccagtataca ctccgctatc gctacgtgac tgggtcatgg ctgcgccccg acacccgcca
       61 acacccgctg acgcgccctg acgggcttgt ctgctcccgg catccgctta cagacaagct
      121 gtgaccgtct ccgggagctg catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg
      181 aggcagctgc ggtaaagctc atcagcgtgg tcgtgaagcg attcacagat gtctgcctgt
      241 tcatccgcgt ccagctcgtt gagtttctcc agaagcgtta atgtctggct tctgataaag
      301 cgggccatgt taagggcggt tttttcctgt ttggtcactg atgcctccgt gtaaggggga
      361 tttctgttca tgggggtaat gataccgatg aaacgagaga ggatgctcac gatacgggtt
      421 actgatgatg aacatgcccg gttactggaa cgttgtgagg gtaaacaact ggcggtatgg
      481 atgcggcggg accagagaaa aatcactcag ggtcaatgcc agcgcttcgt taatacagat
      541 gtaggtgttc cacagggtag ccagcagcat cctgcgatgc agatccggaa cataatggtg
      601 cagggcgctg acttccgcgt ttccagactt tacgaaacac ggaaaccgaa gaccattcat
      661 gttgttgctc aggtcgcaga cgttttgcag cagcagtcgc ttcacgttcg ctcgcgtatc
      721 ggtgattcat tctgctaacc agtaaggcaa ccccgccagc ctagccgggt cctcaacgac
      781 aggagcacga tcatgcgcac ccgtggggcc gccatgccgg cgataatggc ctgcttctcg
      841 ccgaaacgtt tggtggcggg accagtgacg aaggcttgag cgagggcgtg caagattccg
      901 aataccgcaa gcgacaggcc gatcatcgtc gcgctccagc gaaagcggtc ctcgccgaaa
      961 atgacccaga gcgctgccgg cacctgtcct acgagttgca tgataaagaa gacagtcata
     1021 agtgcggcga cgatagtcat gccccgcgcc caccggaagg agctgactgg gttgaaggct
     1081 ctcaagggca tcggtcgaga tcccggtgcc taatgagtga gctaacttac attaattgcg
     1141 ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc
     1201 ggccaacgcg cggggagagg cggtttgcgt attgggcgcc agggtggttt ttcttttcac
     1261 cagtgagacg ggcaacagct gattgccctt caccgcctgg ccctgagaga gttgcagcaa
     1321 gcggtccacg ctggtttgcc ccagcaggcg aaaatcctgt ttgatggtgg ttaacggcgg
     1381 gatataacat gagctgtctt cggtatcgtc gtatcccact accgagatat ccgcaccaac
     1441 gcgcagcccg gactcggtaa tggcgcgcat tgcgcccagc gccatctgat cgttggcaac
     1501 cagcatcgca gtgggaacga tgccctcatt cagcatttgc atggtttgtt gaaaaccgga
     1561 catggcactc cagtcgcctt cccgttccgc tatcggctga atttgattgc gagtgagata
     1621 tttatgccag ccagccagac gcagacgcgc cgagacagaa cttaatgggc ccgctaacag
     1681 cgcgatttgc tggtgaccca atgcgaccag atgctccacg cccagtcgcg taccgtcttc
     1741 atgggagaaa ataatactgt tgatgggtgt ctggtcagag acatcaagaa ataacgccgg
     1801 aacattagtg caggcagctt ccacagcaat ggcatcctgg tcatccagcg gatagttaat
     1861 gatcagccca ctgacgcgtt gcgcgagaag attgtgcacc gccgctttac aggcttcgac
     1921 gccgcttcgt tctaccatcg acaccaccac gctggcaccc agttgatcgg cgcgagattt
     1981 aatcgccgcg acaatttgcg acggcgcgtg cagggccaga ctggaggtgg caacgccaat
     2041 cagcaacgac tgtttgcccg ccagttgttg tgccacgcgg ttgggaatgt aattcagctc
     2101 cgccatcgcc gcttccactt tttcccgcgt tttcgcagaa acgtggctgg cctggttcac
     2161 cacgcgggaa acggtctgat aagagacacc ggcatactct gcgacatcgt ataacgttac
     2221 tggtttcaca ttcaccaccc tgaattgact ctcttccggg cgctatcatg ccataccgcg
     2281 aaaggttttg cgccattcga tggtgtccgg gatctcgacg ctctccctta tgcgactcct
     2341 gcattaggaa gcagcccagt agtaggttga ggccgttgag caccgccgcc gcaaggaatg
     2401 gtgcatgcaa ggagatggcg cccaacagtc ccccggccac ggggcctgcc accataccca
     2461 cgccgaaaca agcgctcatg agcccgaagt ggcgagcccg atcttcccca tcggtgatgt
     2521 cggcgatata ggcgccagca accgcacctg tggcgccggt gatgccggcc acgatgcgtc
     2581 cggcgtagag gatcgagatc gatctcgatc ccgcgaaatt aatacgactc actatagggg
     2641 aattgtgagc ggataacaat tcccctctag aaataatttt gtttaacttt aagaaggaga
     2701 tatacatatg gaacgtccgt acgcttgccc ggttgaatct tgcgaccgtc gtttctctga
     2761 caaaaccaaa ctgcgtgttc atattagaat tcatactgga caaaaaccat tccaatgtag
     2821 aatttgtatg agaaatttct ctgttcgtca caacctgacc cgtcacatcc gtacccacac
     2881 cggtgaaaaa ccgttcgctt gcgacatctg cggtcgtaaa ttcgctcgtt ctgacgaacg
     2941 taaacgtcac accaaaatcc acctgcgtca gtaagatccg gctgctaaca aagcccgaaa
     3001 ggaagctgag ttggctgctg ccaccgctga gcaataacta gcataacccc ttggggcctc
     3061 taaacgggtc ttgaggggtt ttttgctgaa aggaggaact atatccggat tggcgaatgg
     3121 gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc
     3181 gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc
     3241 acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt
     3301 agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg
     3361 ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt
     3421 ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta
     3481 taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt
     3541 aacgcgaatt ttaacaaaat attaacgttt acaatttcag gtggcacttt tcggggaaat
     3601 gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg
     3661 agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa
     3721 catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac
     3781 ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac
     3841 atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt
     3901 ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc
     3961 gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca
     4021 ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc
     4081 ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag
     4141 gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa
     4201 ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgcagcaatg
     4261 gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa
     4321 ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg
     4381 gctggctggt ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt
     4441 gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt
     4501 caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag
     4561 cattggtaac tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat
     4621 ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct
     4681 taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct
     4741 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca
     4801 gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc
     4861 agcagagcgc agataccaaa tactgtcctt ctagtgtagc cgtagttagg ccaccacttc
     4921 aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct
     4981 gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag
     5041 gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc
     5101 tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg
     5161 agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag
     5221 cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt
     5281 gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac
     5341 gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg
     5401 ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc
     5461 cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga gcgcctgatg
     5521 cggtattttc tccttacgca tctgtgcggt atttcacacc gcatatatgg tgcactctca
     5581 gtacaatctg ctctgatgcc gcatagttaa g
//
