LOCUS       sequence-402976-        8537 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For expression of N-flag tagged BoMo R2 protein in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8537)
  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 8537)
  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 BoMo N-1F
FEATURES             Location/Qualifiers
     source          1..8537
                     /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(4128..4145)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4134..4358
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4404..4832
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4491..4510)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4701..4722
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4841..4861)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4846..5175
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5026..5161
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5088..5107
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             5242..6036
                     /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(5296..5315)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5906..5925
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    6210..6331
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6247..6266)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6301..6320
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6380..6396)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6380..6396)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6393..6415)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6404..6420
                     /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(6428..6434,6435..6452,6453..6458))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6473..6494
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6611..6628)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6782..7370)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6862..6881)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7541..8332,8333..8401))
                     /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     8164..8183
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8402..8506)
                     /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 gtaccaagaa
      961 aagcaacaag gagaaccgtc cggaagcgag cggtctgccg ctggagagcg aacgtaccgg
     1021 cgataacccg accgtgcgtg gtagcgcggg tgcggacccg gttggtcagg atgcgccggg
     1081 ttggacctgc caattctgcg agcgtacctt tagcaccaac cgtggtctgg gcgtgcacaa
     1141 gcgtcgtgcg cacccggttg aaaccaacac cgacgcggcg ccgatgatgg tgaaacgtcg
     1201 ttggcacggc gaggaaatcg atctgctggc gcgtaccgag gcgcgtctgc tggcggaacg
     1261 tggccagtgc agcggtggcg acctgttcgg cgcgctgccg ggttttggtc gtaccctgga
     1321 ggcgattaaa ggtcaacgtc gtcgtgaacc gtatcgtgcg ctggttcagg cgcatctggc
     1381 gcgttttggt agccaaccgg gtccgagcag cggtggctgc agcgcggagc cggattttcg
     1441 tcgtgcgagc ggtgcggagg aagcgggcga ggaacgttgc gcggaagatg cggcggcgta
     1501 tgatccgagc gcggtgggtc aaatgagccc ggatgcggcg cgtgtgctga gcgaactgct
     1561 ggagggtgcg ggtcgtcgtc gtgcgtgccg tgcgatgcgt ccgaagaccg cgggtcgtcg
     1621 taacgacctg cacgacgatc gtaccgcgag cgcgcacaag accagccgtc agaaacgtcg
     1681 tgcggagtac gcgcgtgtgc aagaactgta taagaaatgc cgtagccgtg cggcggcgga
     1741 agtgatcgat ggtgcgtgcg gtggcgttgg tcacagcctg gaggaaatgg aaacctactg
     1801 gcgtccgatt ctggaacgtg tgagcgacgc gccgggtccg accccggagg cgctgcacgc
     1861 gctgggtcgt gcggaatggc acggtggcaa ccgtgattat acccagctgt ggaagccgat
     1921 cagcgttgag gaaattaaag cgagccgttt cgactggcgt accagcccgg gtccggatgg
     1981 tatccgtagc ggccagtggc gtgcggtgcc ggttcacctg aaggcggaaa tgttcaacgc
     2041 gtggatggcg cgtggcgaga tcccggaaat tctgcgtcaa tgccgtaccg tgtttgttcc
     2101 gaaagttgag cgtccgggtg gcccgggtga ataccgtccg atcagcattg cgagcatccc
     2161 gctgcgtcac ttccacagca ttctggcgcg tcgtctgctg gcgtgctgcc cgccggacgc
     2221 gcgtcagcgt ggctttatct gcgcggatgg taccctggag aacagcgcgg tgctggacgc
     2281 ggttctgggt gatagccgta agaaactgcg tgaatgccac gtggcggttc tggacttcgc
     2341 gaaggcgttt gataccgtga gccacgaggc gctggttgaa ctgctgcgtc tgcgtggcat
     2401 gccggagcag ttctgcggtt acattgcgca cctgtatgac accgcgagca ccaccctggc
     2461 ggtgaacaac gaaatgagca gcccggtgaa agttggccgt ggtgttcgtc aaggcgaccc
     2521 gctgagcccg atcctgttta acgtggttat ggatctgatt ctggcgagcc tgccggagcg
     2581 tgtgggttac cgtctggaga tggaactggt tagcgcgctg gcgtatgcgg acgatctggt
     2641 gctgctggcg ggcagcaagg ttggtatgca ggaaagcatc agcgcggtgg actgcgttgg
     2701 ccgtcaaatg ggtctgcgtc tgaactgccg taaaagcgcg gtgctgagca tgatcccgga
     2761 tggtcaccgt aagaaacacc actacctgac cgagcgtacc ttcaacattg gtggcaagcc
     2821 gctgcgtcag gtgagctgcg ttgaacgttg gcgttatctg ggcgtggact ttgaggcgag
     2881 cggttgcgtt accctggaac acagcatcag cagcgcgctg aacaacatta gccgtgcgcc
     2941 gctgaaaccg cagcaacgtc tggagatcct gcgtgcgcac ctgattccgc gtttccagca
     3001 cggctttgtt ctgggtaaca tcagcgacga tcgtctgcgt atgctggatg tgcagattcg
     3061 taaggcggtt ggtcaatggc tgcgtctgcc ggcggacgtg ccgaaagcgt actatcacgc
     3121 ggcggttcaa gatggtggcc tggcgatccc gagcgtgcgt gcgaccatcc cggacctgat
     3181 tgttcgtcgt tttggtggcc tggatagcag cccgtggagc gtggcgcgtg cggcggcgaa
     3241 gagcgacaaa attcgtaaga aactgcgttg ggcgtggaag cagctgcgtc gtttcagccg
     3301 tgtggatagc accacccaac gtccgagcgt tcgtctgttt tggcgtgagc acctgcacgc
     3361 gagcgttgac ggtcgtgagc tgcgtgaaag cacccgtacc ccgaccagca ccaaatggat
     3421 tcgtgaacgt tgcgcgcaga ttaccggtcg tgatttcgtg caatttgttc acacccacat
     3481 caacgcgctg ccgagccgta ttcgtggcag ccgtggccgt cgtggtggcg gtgagagcag
     3541 cctgacctgc cgtgcgggtt gcaaagtgcg tgaaaccacc gcgcacatcc tgcagcaatg
     3601 ccaccgtacc cacggcggtc gtatcctgcg tcacaacaag attgtgagct tcgttgcgaa
     3661 ggcgatggag gaaaacaaat ggaccgtgga gctggaaccg cgtctgcgta ccagcgttgg
     3721 cctgcgtaaa ccggacatca ttgcgagccg tgatggcgtg ggtgttatcg tggacgttca
     3781 ggtggttagc ggtcaacgta gcctggatga gctgcaccgt gaaaagcgta acaaatacgg
     3841 caaccacggt gagctggttg agctggttgc gggccgtctg ggtctgccga aagcggagtg
     3901 cgtgcgtgcg accagctgca ccattagctg gcgtggcgtt tggagcctga ccagctataa
     3961 agagctgcgt agcatcattg gtctgcgtga accgaccctg cagatcgtgc cgattctggc
     4021 gctgcgtggc agccacatga actggacccg ttttaaccaa atgaccagcg tgatgggtgg
     4081 cggtgttggt tagtaatcta gagggcccgt ttaaacccgc tgatcagcct cgactgtgcc
     4141 ttctagttgc cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg
     4201 tgccactccc actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag
     4261 gtgtcattct attctggggg gtggggtggg gcaggacagc aagggggagg attgggaaga
     4321 caatagcagg catgctgggg atgcggtggg ctctatggct tctgaggcgg aaagaaccag
     4381 ctggggctct agggggtatc cccacgcgcc ctgtagcggc gcattaagcg cggcgggtgt
     4441 ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc
     4501 tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg
     4561 gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta
     4621 gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt
     4681 ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat
     4741 ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa
     4801 tgagctgatt taacaaaaat ttaacgcgaa ttaattctgt ggaatgtgtg tcagttaggg
     4861 tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag
     4921 tcagcaacca ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg
     4981 catctcaatt agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact
     5041 ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag
     5101 gccgaggccg cctctgcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc
     5161 ctaggctttt gcaaaaagct cccgggagct tgtatatcca ttttcggatc tgatcaagag
     5221 acaggatgag gatcgtttcg catgattgaa caagatggat tgcacgcagg ttctccggcc
     5281 gcttgggtgg agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat
     5341 gccgccgtgt tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg
     5401 tccggtgccc tgaatgaact gcaggacgag gcagcgcggc tatcgtggct ggccacgacg
     5461 ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta
     5521 ttgggcgaag tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta
     5581 tccatcatgg ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc
     5641 gaccaccaag cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc
     5701 gatcaggatg atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg
     5761 ctcaaggcgc gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg
     5821 ccgaatatca tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt
     5881 gtggcggacc gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc
     5941 ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc
     6001 atcgccttct atcgccttct tgacgagttc ttctgagcgg gactctgggg ttcgaaatga
     6061 ccgaccaagc gacgcccaac ctgccatcac gagatttcga ttccaccgcc gccttctatg
     6121 aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg
     6181 atctcatgct ggagttcttc gcccacccca acttgtttat tgcagcttat aatggttaca
     6241 aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt
     6301 gtggtttgtc caaactcatc aatgtatctt atcatgtctg tataccgtcg acctctagct
     6361 agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa
     6421 ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga
     6481 gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt
     6541 gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct
     6601 cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat
     6661 cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga
     6721 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt
     6781 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt
     6841 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc
     6901 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
     6961 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct
     7021 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
     7081 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg
     7141 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc
     7201 ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta
     7261 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg
     7321 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
     7381 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg
     7441 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta
     7501 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
     7561 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     7621 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
     7681 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
     7741 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     7801 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
     7861 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     7921 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     7981 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     8041 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     8101 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
     8161 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     8221 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     8281 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     8341 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     8401 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
     8461 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
     8521 aagtgccacc tgacgtc
//
