LOCUS       sequence-402979-        8915 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For expression of N-flag tagged TaGu R2 protein in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8915)
  AUTHORS   Zhang X, Van Treeck B, Horton CA, McIntyre JJR, Palm SM, Shumate JL,
            Collins K
  TITLE     Harnessing eukaryotic retroelement proteins for transgene insertion 
            into human safe-harbor loci.
  JOURNAL   Nat Biotechnol. 2024 Feb 20. doi: 10.1038/s41587-024-02137-y.
  PUBMED    38379101
REFERENCE   2  (bases 1 to 8915)
  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 
            Biotechnol. 2024 Feb 20. doi: 10.1038/s41587-024-02137-y."
COMMENT     Sequence Label: pcDNA3.1 TaGu N-1F
FEATURES             Location/Qualifiers
     source          1..8915
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      917..926
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             926..949
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(4506..4523)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4512..4736
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4782..5210
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4869..4888)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5079..5100
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5219..5239)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5224..5553
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5404..5539
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5466..5485
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             5620..6414
                     /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(5674..5693)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6284..6303
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    6588..6709
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6625..6644)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6679..6698
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6758..6774)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6758..6774)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6771..6793)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6782..6798
                     /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(6806..6812,6813..6830,6831..6836))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6851..6872
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6989..7006)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(7160..7748)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(7240..7259)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7919..8710,8711..8779))
                     /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"
     primer_bind     8542..8561
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8780..8884)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttaaactt aagcttgcca ccatggatta caaggatgac gacgataagg gtaccgagaa
      961 ggtgatggtc accgttccgg acaagaaccc tccatgtcca tgctgcggca cccgggtgaa
     1021 cagcgtgctg aaccttatcg agcacctgaa ggtcagccac ggaaagcggg gcgtgtgctt
     1081 ccggtgcgcg aagtgtggga aggaaaatag caactaccac agcgtggtgt gccatttccc
     1141 aaaatgtaga ggacctgaga cagagaaagc tccagccggc gagtggattt gtgaagtgtg
     1201 caacagagac ttcaccacca aaatcggcct gggccagcat aagagattag ctcaccccgc
     1261 tgttcggaat caggagagaa tcgtcgccag ccagcctaag gaaactagca acagaggcgc
     1321 tcacaaaaga tgctggacca aagaggagga ggaactgctg atcagactgg aagcccagtt
     1381 tgagggcaac aagaacatca acaagctgat cgcagaacac atcaccacaa agaccgccaa
     1441 gcagatctcc gataagagga gactgctgag ccgcaagcct gccgaggagc ctagagagga
     1501 acctggcaca tgccaccaca ccagacgggc agctgccagc ctgagaacag agcccgagat
     1561 gtcccaccac gctcaggccg aagatcggga taatggccct ggccggagac ccctgcctgg
     1621 tagagccgcc gccggcggca gaacaatgga cgagatcagg agacaccctg acaagggcaa
     1681 tggacagcag cggcctacca aacagaagtc tgaggagcag ctgcaagcct actacaaaaa
     1741 gacactggag gagcggctga gcgccggagc cctgaatacc ttcccccgcg ccttcaagca
     1801 agtgatggaa ggcagagata tcaagctggt gatcaaccag acagcccagg actgcttcgg
     1861 ctgtctggag agcatctctc agatcagaac agcgaccaga gataagaaag ataccgtgac
     1921 cagagagaag cacccgaaga agccattcca aaagtggatg aaggaccggg ccataaagaa
     1981 gggcaactac ctgaggttcc agagattgtt ctacctggac cgaggcaaac tggccaagat
     2041 catcctggac gatatcgaat gcctgagctg cgacatcccc ctgtctgaga tctacagcgt
     2101 gtttaagaca agatgggaaa ccacaggaag cttcaaatct ctgggcgatt ttaaaaccta
     2161 cggcaaggcc gacaacactg ccttcagaga actgattaca gctaaggaaa tcgagaaaaa
     2221 cgtgcaggag atgagcaagg gcagcgcccc tggccctgac ggtatcaccc tgggagatgt
     2281 ggtgaaaatg gaccccgagt tcagccgcac aatggagatc ttcaacctgt ggctgaccac
     2341 tggcaaaatc cctgacatgg tgcggggatg ccgcaccgtg ctgataccta agagcagcaa
     2401 gccagataga ctgaaggata tcaacaactg gcggcctatc acaatcggca gcatcctgct
     2461 ccggctgttt agcagaatcg taaccgcccg tctgagtaag gcctgtcctc tgaaccccag
     2521 acagcggggc ttcatccggg ccgccggctg cagcgagaac ctcaaactgc tgcagaccat
     2581 catctggtcg gccaaacgag agcacagacc tctcggcgtg gtgttcgtgg acatcgccaa
     2641 agcttttgat accgtctccc accagcacat catccacgcc ctgcagcagc gggaagtgga
     2701 ccctcacatc gtgggactgg tgtccaacat gtacgagaac atcagcactt atatcaccac
     2761 caagagaaac acccacacag acaagatcca gatcagagtg ggcgttaagc agggcgatcc
     2821 tatgtctcct ctgctgttca acctggctat ggaccctctg ctctgtaaac tggaggagtc
     2881 tggcaaggga taccacagag gccaaagctc catcaccgct atggcctttg ccgacgacct
     2941 ggtgctgctg tctgattctt gggagaacat gaataccaac ataagcatcc tggaaacatt
     3001 ctgcaacctt acaggcctga aaacccaggg acaaaagtgc cacggctttt acatcaagcc
     3061 taccaaagac agctacacca tcaacgactg cgccgcctgg accatcaacg gcaccccttt
     3121 aaacatgatc gaccctggag aaagcgagaa gtatctgggc ctgcagttcg acccctggat
     3181 cggcatcgct agaagcggcc tgagcacaaa gctggacttt tggctgcaga gaattgacca
     3241 ggcccctctg aagcctctgc agaaaaccga catcctgaaa acctacacaa tccctcggct
     3301 gatctacatt gccgatcaca gcgaggtaaa gaccgccctg ctggaaaccc tggaccaaaa
     3361 gatcagaacc gccgtgaagg aatggctcca cctgcccccc tgcacctgcg acgctatcct
     3421 gtacagcagc accagggacg gcggcctggg catcaccaag ctggcgggcc tgatcccctc
     3481 cgtccaggct agacggctgc atagaatcgc ccagagcagc gatgacacaa tgaagtgctt
     3541 tatggaaaag gaaaagatgg aacagctgca caagaagctg tggattcagg ccggtggcga
     3601 cagagagaac attcctagca tctgggaggc gccgcctagt agcgagcctc ccaacaacgt
     3661 gtctaccaat tctgagtggg aagcccctac acagaaggac aagttcccta agccttgtaa
     3721 ttggagaaag aacgagttca agaagtggac aaagctggcc tctcagggcc ggggaattgt
     3781 gaatttcgag cgggacaaga tcagcaatca ctggattcag tactacagaa gaatcccaca
     3841 cagaaagctg ctgacggccc ttcagctgag agccaacgtg taccccacgc gggagttcct
     3901 ggccagaggt agacaggacc agtacatcaa ggcctgcaga cattgtgatg ctgatatcga
     3961 gtcttgcgcc cacatcatcg gcaactgccc tgtgacacag gacgcgagaa tcaaaagaca
     4021 caactacatc tgcgagctgc tgctggaaga ggccaagaag aaggactggg tggtgttcaa
     4081 ggaaccccac atcagagaca gcaataagga actctataaa cctgacctga tcttcgtgaa
     4141 ggacgcccgg gccctggtcg tggacgtgac cgtgagatac gaggccgcca agtctagcct
     4201 ggaggaggct gccgccgaga aagtgcggaa gtacaagcac cttgaaacag aagtgcggca
     4261 cctgaccaac gccaaggacg tcacattcgt gggcttcccc ctgggcgcca gaggcaaatg
     4321 gcaccaggat aacttcaagc tgctgacaga gctgggcctg agcaagtccc gccaagtgaa
     4381 gatggccgag accttcagca ccgtggccct gttctcctct gtggacatcg tgcacatgtt
     4441 cgctagcaga gccagaaaga gcatggtgat gtagtctaga gggcccgttt aaacccgctg
     4501 atcagcctcg actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc
     4561 ttccttgacc ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc
     4621 atcgcattgt ctgagtaggt gtcattctat tctggggggt ggggtggggc aggacagcaa
     4681 gggggaggat tgggaagaca atagcaggca tgctggggat gcggtgggct ctatggcttc
     4741 tgaggcggaa agaaccagct ggggctctag ggggtatccc cacgcgccct gtagcggcgc
     4801 attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct
     4861 agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg
     4921 tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga
     4981 ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt
     5041 ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg
     5101 aacaacactc aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc
     5161 ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt aattctgtgg
     5221 aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa
     5281 agcatgcatc tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc
     5341 agaagtatgc aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg
     5401 cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt
     5461 ttttttattt atgcagaggc cgaggccgcc tctgcctctg agctattcca gaagtagtga
     5521 ggaggctttt ttggaggcct aggcttttgc aaaaagctcc cgggagcttg tatatccatt
     5581 ttcggatctg atcaagagac aggatgagga tcgtttcgca tgattgaaca agatggattg
     5641 cacgcaggtt ctccggccgc ttgggtggag aggctattcg gctatgactg ggcacaacag
     5701 acaatcggct gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg cccggttctt
     5761 tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc aggacgaggc agcgcggcta
     5821 tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg
     5881 ggaagggact ggctgctatt gggcgaagtg ccggggcagg atctcctgtc atctcacctt
     5941 gctcctgccg agaaagtatc catcatggct gatgcaatgc ggcggctgca tacgcttgat
     6001 ccggctacct gcccattcga ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg
     6061 atggaagccg gtcttgtcga tcaggatgat ctggacgaag agcatcaggg gctcgcgcca
     6121 gccgaactgt tcgccaggct caaggcgcgc atgcccgacg gcgaggatct cgtcgtgacc
     6181 catggcgatg cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc tggattcatc
     6241 gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc tacccgtgat
     6301 attgctgaag agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc
     6361 gctcccgatt cgcagcgcat cgccttctat cgccttcttg acgagttctt ctgagcggga
     6421 ctctggggtt cgaaatgacc gaccaagcga cgcccaacct gccatcacga gatttcgatt
     6481 ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga
     6541 tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccccaac ttgtttattg
     6601 cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt
     6661 tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgta
     6721 taccgtcgac ctctagctag agcttggcgt aatcatggtc atagctgttt cctgtgtgaa
     6781 attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct
     6841 ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc
     6901 agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg
     6961 gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
     7021 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag
     7081 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
     7141 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
     7201 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     7261 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     7321 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
     7381 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     7441 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     7501 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     7561 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg
     7621 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     7681 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     7741 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     7801 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa
     7861 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
     7921 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
     7981 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
     8041 gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc
     8101 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt
     8161 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg
     8221 ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca
     8281 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg
     8341 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca
     8401 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg
     8461 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct
     8521 cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca
     8581 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca
     8641 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg
     8701 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac
     8761 ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt
     8821 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc
     8881 cgcgcacatt tccccgaaaa gtgccacctg acgtc
//
