LOCUS       sequence-479113-        7393 bp    DNA     circular SYN 21-JUL-2025
DEFINITION  NovaIscB mammalian expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7393)
  AUTHORS   Kannan S, Altae-Tran H, Zhu S, Xu P, Strebinger D, Oshiro R, Faure 
            G, Moeller L, Pham J, Mears KS, Ni HM, Macrae RK, Zhang F
  TITLE     Evolution-guided protein design of IscB for persistent epigenome 
            editing in vivo.
  JOURNAL   Nat Biotechnol. 2025 May 7. doi: 10.1038/s41587-025-02655-3.
  PUBMED    40335752
REFERENCE   2  (bases 1 to 7393)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jul 21, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Biotechnol. 2025 May 7. doi: 10.1038/s41587-025-02655-3."
COMMENT     Sequence Label: pHS_Nova
FEATURES             Location/Qualifiers
     source          1..7393
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     612..631
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(850..954)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(1029..1048)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        1220..1599
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1600..1803
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1754..1774
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1848..1867
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1848..1866
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      1902..1911
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1917..1937
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3789..3836
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             3843..3923
                     /codon_start=1
                     /product="three tandem HA epitope tags"
                     /label=3xHA
                     /translation="YPYDVPDYAYPYDVPDYAYPYDVPDYA"
     primer_bind     complement(3969..3986)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3975..4199
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4245..4673
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4332..4351)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4542..4563
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4682..4702)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4687..5016
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4867..5002
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4929..4948
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             5083..5877
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(5137..5156)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5747..5766
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    6051..6172
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6088..6107)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6142..6161
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6221..6237)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6221..6237)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6234..6256)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6245..6261
                     /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(6269..6275,6276..6293,6294..6299))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6314..6335
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6452..6469)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6623..7211)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6703..6722)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7382..7393,1..780,781..849))
                     /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"
ORIGIN
        1 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact
       61 ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat
      121 gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg
      181 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg
      241 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat
      301 tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc
      361 ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt
      421 cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc
      481 agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga
      541 gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc
      601 gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa
      661 acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta
      721 acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg
      781 agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg
      841 aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat
      901 gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt
      961 tccccgaaaa gtgccacctg acgtcgacgg atcgggagat ctcccgatcc cctatggtgc
     1021 actctcagta caatctgctc tgatgccgca tagttaagcc agtatctgct ccctgcttgt
     1081 gtgttggagg tcgctgagta gtgcgcgagc aaaatttaag ctacaacaag gcaaggcttg
     1141 accgacaatt gcatgaagaa tctgcttagg gttaggcgtt ttgcgctgct tcgcgatgta
     1201 cgggccagat atacgcgttg acattgatta ttgactagtt attaatagta atcaattacg
     1261 gggtcattag ttcatagccc atatatggag ttccgcgtta cataacttac ggtaaatggc
     1321 ccgcctggct gaccgcccaa cgacccccgc ccattgacgt caataatgac gtatgttccc
     1381 atagtaacgc caatagggac tttccattga cgtcaatggg tggagtattt acggtaaact
     1441 gcccacttgg cagtacatca agtgtatcat atgccaagta cgccccctat tgacgtcaat
     1501 gacggtaaat ggcccgcctg gcattatgcc cagtacatga ccttatggga ctttcctact
     1561 tggcagtaca tctacgtatt agtcatcgct attaccatgg tgatgcggtt ttggcagtac
     1621 atcaatgggc gtggatagcg gtttgactca cggggatttc caagtctcca ccccattgac
     1681 gtcaatggga gtttgttttg gcaccaaaat caacgggact ttccaaaatg tcgtaacaac
     1741 tccgccccat tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta tataagcaga
     1801 gctctctgga taactagaga acccactgct tactggctta tcgaaattaa tacgactcac
     1861 tatagggaga cccaagctgg ctagcgttta aacttaagct tgccaccatg ggatctccaa
     1921 aaaagaaacg gaaggtggga tctagatacg tgtacgtgct ggatgtggac ggcaagcctc
     1981 tgatgcctac atgcaggttc ggcaaggtgc ggagaatgct gaaatctggc caagctaagg
     2041 ccgtggacac cctgcctttc accatccagc tgacctacaa gcccagaacc agaatcctgc
     2101 aacctgtgac cctgggccag gaccccggca gaaccaatat cggaatggcc gccgtgcggt
     2161 tcgacggcaa ggaactggga agattccact gcatcacaag aaacaaggaa atccctaagc
     2221 tgatggccga cagaatggca gccagaaagg cctcccggag aggcgagaga ctggccagaa
     2281 agcgcctggc ccggaaactg cacaccaccg ccaagcacct gaatggcaga atcctgccag
     2341 gatgtagcga gcccatcgcc gtgaaggaca tcatcaacac agagagcaga ttcaacaaca
     2401 gaatcctgac caaatgcaaa gtgtgcggca agaacacccc tctgagaaga aacgtgcggg
     2461 aactcctgct ggaaaacatc gtgaagaaca tcagactgca cggccctggc ggagagatcg
     2521 tgggcctgca ccccgacctg aaggaaacac tcaagagaac catcctggaa ggccagcagg
     2581 gcaatatcaa caagctgttc aggaaactta gaaaggtgta caagatcacc ctgaaccaga
     2641 aagactggcc tggcaaaaac ctgaccgata tcgctaagaa taagctgcca ggccggctgc
     2701 ccttctgcaa ggagcacttc gccgagaacg agaagttcac caccatcgag aagagcacct
     2761 tccggctgac ccctaccgcc acccagctgc tgcggaccca catcaacctg tttagaaagc
     2821 tgagcggcat cctgcccgtg accgacgtgg ccgtcgagct gaacaagttc gcctttatgc
     2881 agctggacaa ccctgagatg aagaagcggg aaatcgactt ctgccacggc ccactgtgcg
     2941 gcaccggcgg cctggaagcc gctgttaagg agcagcagga tggaaaatgc ctgctgtgtg
     3001 gaaaagaatc tatcggccac taccaccaca tcgtgcctag gtcccgtaga ggcagcaaca
     3061 ccatcgctaa tatcgccggc ctttgtccta agtgccacga gctcgtgcac aaggacgccg
     3121 atacagccga gtctctgaca gagatgaaaa ccggcctgat gaagaagtat ggaggcacca
     3181 gcgtgctgaa ccagatcatc cccaagctgg tggaaaccct ggccgacctg tttcctggcc
     3241 acttccacgt gacaaacggc tggaacacca aggagttcag agagaaacat catctggaaa
     3301 aggatcacga cgtggatgct tactgcatcg cctgcagcca cctgaagcct gaggaaaccc
     3361 tggtggaaac tgagcctttc gagatcctgc agttcagaaa gcacaacaga gccatcattc
     3421 accaccagac cgagcggacg tacaagctgg acggcgtgac agtggctaaa aatagaaaga
     3481 aacggatgga acagaaaacc gacagcctgg aagattggta cgtggacatg gccaaggaac
     3541 acggcaaaac acaggccgat gccatgcgga gcagactgac cgtgatcaag agcacacggt
     3601 attacaacac ccctggcaga atgatgccag gcacagtttt cctgtacgag ggcaagagat
     3661 acgtgatgac cggccagatt acaaacggca agtactacag agcctacggc caggagaagc
     3721 ggaacttccc cgctgtcaag gtgaggattc tgacaaagaa caccggacta gtctttgtgg
     3781 ccggatccaa aaggccggcg gccacgaaaa aggccggcca ggcaaaaaag aaaaagggat
     3841 cttacccata cgatgttcca gattacgctt acccatacga tgttccagat tacgcttacc
     3901 catacgatgt tccagattac gcttgaggcc gctcgagtct agagggcccg tttaaacccg
     3961 ctgatcagcc tcgactgtgc cttctagttg ccagccatct gttgtttgcc cctcccccgt
     4021 gccttccttg accctggaag gtgccactcc cactgtcctt tcctaataaa atgaggaaat
     4081 tgcatcgcat tgtctgagta ggtgtcattc tattctgggg ggtggggtgg ggcaggacag
     4141 caagggggag gattgggaag acaatagcag gcatgctggg gatgcggtgg gctctatggc
     4201 ttctgaggcg gaaagaacca gctggggctc tagggggtat ccccacgcgc cctgtagcgg
     4261 cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc
     4321 cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc
     4381 ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct
     4441 cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt gggccatcgc cctgatagac
     4501 ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac
     4561 tggaacaaca ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat
     4621 ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attaattctg
     4681 tggaatgtgt gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg
     4741 caaagcatgc atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca
     4801 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact
     4861 ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta
     4921 atttttttta tttatgcaga ggccgaggcc gcctctgcct ctgagctatt ccagaagtag
     4981 tgaggaggct tttttggagg cctaggcttt tgcaaaaagc tcccgggagc ttgtatatcc
     5041 attttcggat ctgatcaaga gacaggatga ggatcgtttc gcatgattga acaagatgga
     5101 ttgcacgcag gttctccggc cgcttgggtg gagaggctat tcggctatga ctgggcacaa
     5161 cagacaatcg gctgctctga tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt
     5221 ctttttgtca agaccgacct gtccggtgcc ctgaatgaac tgcaggacga ggcagcgcgg
     5281 ctatcgtggc tggccacgac gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa
     5341 gcgggaaggg actggctgct attgggcgaa gtgccggggc aggatctcct gtcatctcac
     5401 cttgctcctg ccgagaaagt atccatcatg gctgatgcaa tgcggcggct gcatacgctt
     5461 gatccggcta cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact
     5521 cggatggaag ccggtcttgt cgatcaggat gatctggacg aagagcatca ggggctcgcg
     5581 ccagccgaac tgttcgccag gctcaaggcg cgcatgcccg acggcgagga tctcgtcgtg
     5641 acccatggcg atgcctgctt gccgaatatc atggtggaaa atggccgctt ttctggattc
     5701 atcgactgtg gccggctggg tgtggcggac cgctatcagg acatagcgtt ggctacccgt
     5761 gatattgctg aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc
     5821 gccgctcccg attcgcagcg catcgccttc tatcgccttc ttgacgagtt cttctgagcg
     5881 ggactctggg gttcgaaatg accgaccaag cgacgcccaa cctgccatca cgagatttcg
     5941 attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct
     6001 ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccacccc aacttgttta
     6061 ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat
     6121 ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct
     6181 gtataccgtc gacctctagc tagagcttgg cgtaatcatg gtcatagctg tttcctgtgt
     6241 gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag
     6301 cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt
     6361 tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag
     6421 gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg
     6481 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat
     6541 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     6601 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     6661 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     6721 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     6781 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     6841 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     6901 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     6961 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     7021 cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct
     7081 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     7141 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     7201 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
     7261 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
     7321 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca
     7381 gttaccaatg ctt
//
