LOCUS       sequence-263301-        5636 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  coding sequence for synthetic zinc finger ZF-ADD-L.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5636)
  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 5636)
  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-L
FEATURES             Location/Qualifiers
     source          1..5636
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    1..25
                     /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             40..62
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     primer_bind     complement(419..437)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      433..480
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      517..972
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(604..623)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     814..835
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        998..1102
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1103..1171,1172..1963)
                     /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(1321..1340)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2134..2722
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2623..2642
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2876..2893
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    2908..3047
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(2973..2992)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3133..3155
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3149..3340)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4149..5231)
                     /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     5193..5212
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5232..5309)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5515..5534
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     join(5618..5636,1)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5618..5636
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 ggaattgtga gcggataaca attcccctct agaaataatt ttgtttaact ttaagaagga
       61 gatatacata tggaacgtcc gtacgcttgc ccggttgaat cttgcgaccg tcgtttctct
      121 gacaaaacca aactgcgtgt tcatattaga attcatactg gacaaaaacc attccaatgt
      181 agaatttgta tgagaaattt ctctgttcgt cacaacctga cccgtcacat ccgtacccac
      241 accggtgaaa aaccgttcgc ttgcgacatc tgcggtcgta aattcgctcg ttctgacgaa
      301 cgtaaacgtc acaccaaaat ccacctgcgt cagggcagcg gcagcgtgaa agaaagcctg
      361 gtgtaataag atccggctgc taacaaagcc cgaaaggaag ctgagttggc tgctgccacc
      421 gctgagcaat aactagcata accccttggg gcctctaaac gggtcttgag gggttttttg
      481 ctgaaaggag gaactatatc cggattggcg aatgggacgc gccctgtagc ggcgcattaa
      541 gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc
      601 ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag
      661 ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca
      721 aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc
      781 gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa
      841 cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct
      901 attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa
      961 cgcttacaat ttaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt
     1021 tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca
     1081 ataatattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt
     1141 ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga
     1201 tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa
     1261 gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct
     1321 gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat
     1381 acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga
     1441 tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc
     1501 caacttactt ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat
     1561 gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa
     1621 cgacgagcgt gacaccacga tgcctgcagc aatggcaaca acgttgcgca aactattaac
     1681 tggcgaacta cttactctag cttcccggca acaattaata gactggatgg aggcggataa
     1741 agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc
     1801 tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc
     1861 ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag
     1921 acagatcgct gagataggtg cctcactgat taagcattgg taactgtcag accaagttta
     1981 ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa
     2041 gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc
     2101 gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat
     2161 ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga
     2221 gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt
     2281 ccttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata
     2341 cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac
     2401 cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg
     2461 ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg
     2521 tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag
     2581 cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct
     2641 ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc
     2701 aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt
     2761 ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg
     2821 tattaccgcc tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga
     2881 gtcagtgagc gaggaagcgg aagagcgcct gatgcggtat tttctcctta cgcatctgtg
     2941 cggtatttca caccgcaatg gtgcactctc agtacaatct gctctgatgc cgcatagtta
     3001 agccagtata cactccgcta tcgctacgtg actgggtcat ggctgcgccc cgacacccgc
     3061 caacacccgc tgacgcgccc tgacgggctt gtctgctccc ggcatccgct tacagacaag
     3121 ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg
     3181 cgaggcagct gcggtaaagc tcatcagcgt ggtcgtgaag cgattcacag atgtctgcct
     3241 gttcatccgc gtccagctcg ttgagtttct ccagaagcgt taatgtctgg cttctgataa
     3301 agcgggccat gttaagggcg gttttttcct gtttggtcac tgatgcctcc gtgtaagggg
     3361 gatttctgtt catgggggta atgataccga tgaaacgaga gaggatgctc acgatacggg
     3421 ttactgatga tgaacatgcc cggttactgg aacgttgtga gggtaaacaa ctggcggtat
     3481 ggatgcggcg ggaccagaga aaaatcactc agggtcaatg ccagcgcttc gttaatacag
     3541 atgtaggtgt tccacagggt agccagcagc atcctgcgat gcagatccgg aacataatgg
     3601 tgcagggcgc tgacttccgc gtttccagac tttacgaaac acggaaaccg aagaccattc
     3661 atgttgttgc tcaggtcgca gacgttttgc agcagcagtc gcttcacgtt cgctcgcgta
     3721 tcggtgattc attctgctaa ccagtaaggc aaccccgcca gcctagccgg gtcctcaacg
     3781 acaggagcac gatcatgcgc acccgtgggg ccgccatgcc ggcgataatg gcctgcttct
     3841 cgccgaaacg tttggtggcg ggaccagtga cgaaggcttg agcgagggcg tgcaagattc
     3901 cgaataccgc aagcgacagg ccgatcatcg tcgcgctcca gcgaaagcgg tcctcgccga
     3961 aaatgaccca gagcgctgcc ggcacctgtc ctacgagttg catgataaag aagacagtca
     4021 taagtgcggc gacgatagtc atgccccgcg cccaccggaa ggagctgact gggttgaagg
     4081 ctctcaaggg catcggtcga gatcccggtg cctaatgagt gagctaactt acattaattg
     4141 cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa
     4201 tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ccagggtggt ttttcttttc
     4261 accagtgaga cgggcaacag ctgattgccc ttcaccgcct ggccctgaga gagttgcagc
     4321 aagcggtcca cgctggtttg ccccagcagg cgaaaatcct gtttgatggt ggttaacggc
     4381 gggatataac atgagctgtc ttcggtatcg tcgtatccca ctaccgagat atccgcacca
     4441 acgcgcagcc cggactcggt aatggcgcgc attgcgccca gcgccatctg atcgttggca
     4501 accagcatcg cagtgggaac gatgccctca ttcagcattt gcatggtttg ttgaaaaccg
     4561 gacatggcac tccagtcgcc ttcccgttcc gctatcggct gaatttgatt gcgagtgaga
     4621 tatttatgcc agccagccag acgcagacgc gccgagacag aacttaatgg gcccgctaac
     4681 agcgcgattt gctggtgacc caatgcgacc agatgctcca cgcccagtcg cgtaccgtct
     4741 tcatgggaga aaataatact gttgatgggt gtctggtcag agacatcaag aaataacgcc
     4801 ggaacattag tgcaggcagc ttccacagca atggcatcct ggtcatccag cggatagtta
     4861 atgatcagcc cactgacgcg ttgcgcgaga agattgtgca ccgccgcttt acaggcttcg
     4921 acgccgcttc gttctaccat cgacaccacc acgctggcac ccagttgatc ggcgcgagat
     4981 ttaatcgccg cgacaatttg cgacggcgcg tgcagggcca gactggaggt ggcaacgcca
     5041 atcagcaacg actgtttgcc cgccagttgt tgtgccacgc ggttgggaat gtaattcagc
     5101 tccgccatcg ccgcttccac tttttcccgc gttttcgcag aaacgtggct ggcctggttc
     5161 accacgcggg aaacggtctg ataagagaca ccggcatact ctgcgacatc gtataacgtt
     5221 actggtttca cattcaccac cctgaattga ctctcttccg ggcgctatca tgccataccg
     5281 cgaaaggttt tgcgccattc gatggtgtcc gggatctcga cgctctccct tatgcgactc
     5341 ctgcattagg aagcagccca gtagtaggtt gaggccgttg agcaccgccg ccgcaaggaa
     5401 tagtgcatgc aaggagatgg cgcccaacag tcccccggcc acggggcctg ccaccatacc
     5461 cacgccgaaa caagcgctca tgagcccgaa gtggcgagcc cgatcttccc catcggtgat
     5521 gtcggcgata taggcgccag caaccgcacc tgtggcgccg gtgatgccgg ccacgatgcg
     5581 tccggcgtag aggatcgaga tctcgatccc gcgaaattaa tacgactcac tatagg
//
