LOCUS       sequence-434856-        5516 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Recombinant expression of UNK ZnF1-3 with dual-affinity tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5516)
  AUTHORS   Harris SE, Alexis MS, Giri G, Cavazos FF Jr, Hu Y, Murn J, Aleman 
            MM, Burge CB, Dominguez D
  TITLE     Understanding species-specific and conserved RNA-protein 
            interactions in vivo and in vitro.
  JOURNAL   Nat Commun. 2024 Sep 27;15(1):8400. doi: 10.1038/s41467-024-52231-7.
  PUBMED    39333159
REFERENCE   2  (bases 1 to 5516)
  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"; journalName: "Nat 
            Commun."; date: "2024-09-27"; volume: "15(1):8400. doi"; pages: " 
            10.1038/s41467-024-52231-7"
COMMENT     Sequence Label: UNK_ZnF1-3_pGEX
FEATURES             Location/Qualifiers
     source          1..5516
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     27..49
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     41..58
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     42..58
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(74..90)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(74..90)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     protein_bind    98..114
                     /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)."
     promoter        complement(join(122..128,129..146,147..152))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     CDS             complement(201..1283)
                     /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     1245..1264
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(1284..1361)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     complement(1434..1451)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1605..2193)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1685..1704)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2364..3155,3156..3224))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     2987..3006
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3225..3329)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3397..3415
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     3560..3582
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4084..4197)
                     /codon_start=1
                     /product="streptavidin-binding peptide"
                     /label=SBP
                     /note="selected from a peptide library; binds streptavidin 
                     with nanomolar affinity (Keefe et al., 2001)"
                     /translation="MDEKTTGWRGGHVVEGLAGELEQLRARLEHHPQGQREP"
     CDS             complement(4207..4230)
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             complement(4237..4890)
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     complement(4257..4279)
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     protein_bind    4913..4929
                     /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)."
     promoter        complement(join(4937..4943,4944..4959,4960..4965))
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     CDS             complement(join(5067..5516,1..78))
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
ORIGIN
        1 attaagttgg gtaacgccag ggttttccca gtcacgacgt tgtaaaacga cggccagtga
       61 atccgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac
      121 acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac
      181 ttacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc
      241 tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctagcgtg
      301 gtttttcttt tcaccagtga gacgggcaac agctgattgc ccttcaccgc ctggccctga
      361 gagagttgca gcaagcggtc cacgctggtt tgccccagca ggcgaaaatc ctgtttgatg
      421 gtggttaacg gcgggatata acatgagctg tcttcggtat cgtcgtatcc cactaccgag
      481 atatccgcac caacgcgcag cccggactcg gtaatggcgc gcattgcgcc cagcgccatc
      541 tgatcgttgg caaccagcat cgcagtggga acgatgccct cattcagcat ttgcatggtt
      601 tgttgaaaac cggacatggc actccagtcg ccttcccgtt ccgctatcgg ctgaatttga
      661 ttgcgagtga gatatttatg ccagccagcc agacgcagac gcgccgagac agaacttaat
      721 gggcccgcta acagcgcgat ttgctggtga cccaatgcga ccagatgctc cacgcccagt
      781 cgcgtaccgt cttcatggga gaaaataata ctgttgatgg gtgtctggtc agagacatca
      841 agaaataacg ccggaacatt agtgcaggca gcttccacag caatggcatc ctggtcatcc
      901 agcggatagt taatgatcag cccactgacg cgttgcgcga gaagattgtg caccgccgct
      961 ttacaggctt cgacgccgct tcgttctacc atcgacacca ccacgctggc acccagttga
     1021 tcggcgcgag atttaatcgc cgcgacaatt tgcgacggcg cgtgcagggc cagactggag
     1081 gtggcaacgc caatcagcaa cgactgtttg cccgccagtt gttgtgccac gcggttggga
     1141 atgtaattca gctccgccat cgccgcttcc actttttccc gcgttttcgc agaaacgtgg
     1201 ctggcctggt tcaccacgcg ggaaacggtc tgataagaga caccggcata ctctgcgaca
     1261 tcgtataacg ttactggttt cacattcacc accctgaatt gactctcttc cgggcgctat
     1321 catgccatac cgcgaaaggt tttgcgccat tcgatggtgt cggaatttat gcggtgtgaa
     1381 ataccgcaca gatgcgtaag gagaaaatac cgcatcaggc gctcttccgc ttcctcgctc
     1441 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
     1501 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc
     1561 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     1621 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     1681 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     1741 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
     1801 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     1861 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     1921 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     1981 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     2041 agaaggacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     2101 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     2161 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     2221 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     2281 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
     2341 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
     2401 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
     2461 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
     2521 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
     2581 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
     2641 tcgccagtta atagtttgcg caacgttgtt gccattgctg caggcatcgt ggtgtcacgc
     2701 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
     2761 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
     2821 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
     2881 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
     2941 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa cacgggataa taccgcgcca
     3001 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
     3061 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
     3121 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
     3181 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
     3241 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
     3301 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc
     3361 taagaaacca ttattatcat gacattaacc tataaaaata ggcgtatcac gaggcccttt
     3421 cgtcttcaag aattatacac tccgctatcg ctacgtgact gggtcatggc tgcgccccga
     3481 cacccgccaa cacccgctga cgcgccctga cgggcttgtc tgctcccggc atccgcttac
     3541 agacaagctg tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg
     3601 aaacgcgcga ggcagatcgt cagtcagtca cgatgcggcc gctcaattct gcaaggcctc
     3661 catggcctga agctccctga tgtcgtagac aggggagcgg aggtcatggg gcccgtgggc
     3721 aaaagcgcag tgcaggccgt ttttggtgca gttgcctttc gagtctgtct cgtggatgca
     3781 gattccagtt ttgtagtaac gaaggtggta cctgcgctca gtgtcccctg tggttctgtg
     3841 caggaatggg cactcgtcgc cctccgggca gaggcctgta gcctcgtcgt acttggtgca
     3901 gtagacgtca gggctgtaat tgaaggtgcc gtcccgacgg cggatggacc ggcggcgccg
     3961 ctggttcacg aagtgccagt ggaagcaggt gtagggccga tgctgcgtgc atttgtgttg
     4021 cacaaagagt gggcactgct ctgtgcggaa ttctttcagg tacgtgtagt gctgcgggga
     4081 tcccggttca cgctgaccct gcgggtggtg ttccagacga gcacgcagct gttccagttc
     4141 accagccaga ccttcaacaa cgtgaccacc acgccaaccg gtggtttttt cgtccattaa
     4201 tcccaggggc ccctggaaca gaacttccag atccgatttt ggaggatggt cgccaccacc
     4261 aaacgtggct tgccagccct gcaaaggcca tgctatatac ttgctggatt tcaagtactt
     4321 atcaatttgt gggatagctt caatacgttt tttaaaacaa actaattttg ggaacgcatc
     4381 caggcacatt gggtccatgt ataaaacaac atcaagagcg tcatacaaca tgaagtcagg
     4441 atgggttaca tgatcaccat ttaaatatgt tttatgacat aaacgatctt cgaacatttt
     4501 cagcatttca ggtagcttgc taagaaaatc aactttgaga gtttcaaagt ctttactata
     4561 tgcaattctc gaaacaccgt atctaatatc caaaaccgct ccttcaagca ttgaaatctc
     4621 tgcacgctct tttggacaac cacccaacat gttgtgcttg tcagctatat aacgtatgat
     4681 ggccatagac tgtgttaatt taacatcacc atcaatataa taaggaagat tgggaaactc
     4741 caaacccaat tcaaactttt tgtttcgcca tttatcacct tcatcgcgct catacaaatg
     4801 ctcttcatat ttttcttcaa gatattccaa aagaagtcga gtgggttgca caaggccctt
     4861 aattttccaa taacctagta taggggacat gaatactgtt tcctgtgtga aattgttatc
     4921 cgctcacaat tccacacatt atacgagccg atgattaatt gtcaacagct catttcagaa
     4981 tatttgccag aaccgttatg atgtcggcgc aaaaaacatt atccagaacg ggagtgcgcc
     5041 ttgagcgaca cgaattatgc agtgatttac gacctgcaca gccataccac agcttccgat
     5101 ggctgcctga cgccagaagc attggtgcac cgtgcagtcg ataagctaat tccaacgcca
     5161 tcaaaaataa ttcgcgtctg gccttcctgt agccagcttt catcaacatt aaatgtgagc
     5221 gagtaacaac ccgtcggatt ctccgtggga acaaacggcg gattgaccgt aatgggatag
     5281 gtcacgttgg tgtagatggg cgcatcgtaa ccgtgcatct gccagtttga ggggacgacg
     5341 acagtatcgg cctcaggaag atcgcactcc agccagcttt ccggcaccgc ttctggtgcc
     5401 ggaaaccagg caaagcgcca ttcgccattc aggctgcgca actgttggga agggcgatcg
     5461 gtgcgggcct cttcgctatt acgccagctg gcgaaagggg gatgtgctgc aaggcg
//
