LOCUS       sequence-491521-        7667 bp    DNA     circular SYN 04-NOV-2025
DEFINITION  loxPcon DIAL Reporter Lentivirus expressing mCherry-HRasG12V in the 
            presence of ZFa .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7667)
  AUTHORS   Kabaria SR, Bae Y, Ehmann ME, Lende-Dorn BA, Beitz AM, Peterman EL, 
            Love KS, Ploessl DS, Galloway KE
  TITLE     Programmable promoter editing for precise control of transgene 
            expression.
  JOURNAL   Nat Biotechnol. 2025 Oct 13. doi: 10.1038/s41587-025-02854-y.
  PUBMED    41083901
REFERENCE   2  (bases 1 to 7667)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Nov 4, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Biotechnol. 2025 Oct 13. doi: 10.1038/s41587-025-02854-y."
COMMENT     Sequence Label: 
            LentiX_(loxPconDIAL-YB_TATA)-mCherry-HRasG12V-BGH_pKG3906
FEATURES             Location/Qualifiers
     source          1..7667
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     743..760
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    877..898
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(913..918,919..936,937..943)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    951..967
                     /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)."
     primer_bind     956..978
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     975..991
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     975..991
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     1010..1030
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        1012..1030
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        1058..1284
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             1285..1465
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1512..1637
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    2130..2363
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2548..2592
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    2859..2975
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     protein_bind    complement(3133..3166)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             3245..3952
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(3390..3408)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     3658..3677
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             join(3983..4015,4016..4018,4019..4552)
                     /codon_start=1
                     /product="human oncoprotein generated by the G12V mutation 
                     in the small GTPase H-Ras"
                     /label=H-Ras (G12V)
                     /translation="MTEYKLVVVGAVGVGKSALTIQLIQNHFVDEYDPTIEDSYRKQVV
                     IDGETCLLDILDTAGQEEYSAMRDQYMRTGEGFLCVFAINNTKSFEDIHQYREQIKRVK
                     DSDDVPMVLVGNKCDLAARTVESRQAQDLARSYGIPYIETSAKTRQGVEDAFYTLVREI
                     RQHKLRKLNPPDESGPGCMSCKCVLS"
     misc_feature    4563..5151
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4616..4636)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5034..5045)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             5243..5476
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    5548..5669
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5585..5604)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5639..5658
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      5709..5844
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(5864..5883)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(5865..5883)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5893..5910)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5893..5909)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5902..5924)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      6051..6506
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6138..6157)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6348..6369
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        6532..6636
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6637..6705,6706..7497)
                     /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     complement(6855..6874)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
      721 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat
      781 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt
      841 tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc tcactcatta
      901 ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg
      961 ataacaattt cacacaggaa acagctatga ccatgattac gccaagcgcg caattaaccc
     1021 tcactaaagg gaacaaaagc tggagctgca agcttaatgt agtcttatgc aatactcttg
     1081 tagtcttgca acatggtaac gatgagttag caacatgcct tacaaggaga gaaaaagcac
     1141 cgtgcatgcc gattggtgga agtaaggtgg tacgatcgtg ccttattagg aaggcaacag
     1201 acgggtctga catggattgg acgaaccact gaattgccgc attgcagaga tattgtattt
     1261 aagtgcctag ctcgatacat aaacgggtct ctctggttag accagatctg agcctgggag
     1321 ctctctggct aactagggaa cccactgctt aagcctcaat aaagcttgcc ttgagtgctt
     1381 caagtagtgt gtgcccgtct gttgtgtgac tctggtaact agagatccct cagacccttt
     1441 tagtcagtgt ggaaaatctc tagcagtggc gcccgaacag ggacttgaaa gcgaaaggga
     1501 aaccagagga gctctctcga cgcaggactc ggcttgctga agcgcgcacg gcaagaggcg
     1561 aggggcggcg actggtgagt acgccaaaaa ttttgactag cggaggctag aaggagagag
     1621 atgggtgcga gagcgtcagt attaagcggg ggagaattag atcgcgatgg gaaaaaattc
     1681 ggttaaggcc agggggaaag aaaaaatata aattaaaaca tatagtatgg gcaagcaggg
     1741 agctagaacg attcgcagtt aatcctggcc tgttagaaac atcagaaggc tgtagacaaa
     1801 tactgggaca gctacaacca tcccttcaga caggatcaga agaacttaga tcattatata
     1861 atacagtagc aaccctctat tgtgtgcatc aaaggataga gataaaagac accaaggaag
     1921 ctttagacaa gatagaggaa gagcaaaaca aaagtaagac caccgcacag caagcggccg
     1981 ctgatcttca gacctggagg aggagatatg agggacaatt ggagaagtga attatataaa
     2041 tataaagtag taaaaattga accattagga gtagcaccca ccaaggcaaa gagaagagtg
     2101 gtgcagagag aaaaaagagc agtgggaata ggagctttgt tccttgggtt cttgggagca
     2161 gcaggaagca ctatgggcgc agcgtcaatg acgctgacgg tacaggccag acaattattg
     2221 tctggtatag tgcagcagca gaacaatttg ctgagggcta ttgaggcgca acagcatctg
     2281 ttgcaactca cagtctgggg catcaagcag ctccaggcaa gaatcctggc tgtggaaaga
     2341 tacctaaagg atcaacagct cctggggatt tggggttgct ctggaaaact catttgcacc
     2401 actgctgtgc cttggaatgc tagttggagt aataaatctc tggaacagat ttggaatcac
     2461 acgacctgga tggagtggga cagagaaatt aacaattaca caagcttaat acactcctta
     2521 attgaagaat cgcaaaacca gcaagaaaag aatgaacaag aattattgga attagataaa
     2581 tgggcaagtt tgtggaattg gtttaacata acaaattggc tgtggtatat aaaattattc
     2641 ataatgatag taggaggctt ggtaggttta agaatagttt ttgctgtact ttctatagtg
     2701 aatagagtta ggcagggata ttcaccatta tcgtttcaga cccacctccc aaccccgagg
     2761 ggacccgaca ggcccgaagg aatagaagaa gaaggtggag agagagacag agacagatcc
     2821 attcgattag tgaacggatc tcgacggtat cggttaactt ttaaaagaaa aggggggatt
     2881 ggggggtaca gtgcagggga aagaatagta gacataatag caacagacat acaaactaaa
     2941 gaattacaaa aacaaattac aaaattcaaa attttatcgt ccagtgtggt ggctacttac
     3001 tcggatcggg agatctgaat tggagtgagg acgtgtagag tgaggacgtg cggagtgagg
     3061 acgtgtagag tgaggacgtg cggagtgagg acgtgtagag tgaggacgtg ccctcctctg
     3121 gatcaaatcc gaataacttc gtatagcata cattatacga agttatcgga tttgatctga
     3181 tcaagcaact ctagagggta tataatgggg gccactagtc tactaccaga gctcatcgct
     3241 agcaatggtg agcaagggcg aggaggataa catggccatc atcaaggagt tcatgcgctt
     3301 caaggtgcac atggagggct ccgtgaacgg ccacgagttc gagatcgagg gcgagggcga
     3361 gggccgcccc tacgagggca cccagaccgc caagctgaag gtgaccaagg gtggccccct
     3421 gcccttcgcc tgggacatcc tgtcccctca gttcatgtac ggctccaagg cctacgtgaa
     3481 gcaccccgcc gacatccccg actacttgaa gctgtccttc cccgagggct tcaagtggga
     3541 gcgcgtgatg aacttcgagg acggcggcgt ggtgaccgtg acccaggact cctccctgca
     3601 ggacggcgag ttcatctaca aggtgaagct gcgcggcacc aacttcccct ccgacggccc
     3661 cgtaatgcag aagaagacca tgggctggga ggcctcctcc gagcggatgt accccgagga
     3721 cggcgccctg aagggcgaga tcaagcagag gctgaagctg aaggacggcg gccactacga
     3781 cgctgaggtc aagaccacct acaaggccaa gaagcccgtg cagctgcccg gcgcctacaa
     3841 cgtcaacatc aagttggaca tcacctccca caacgaggac tacaccatcg tggaacagta
     3901 cgaacgcgcc gagggccgcc actccaccgg cggcatggac gagctgtaca agggtggtgg
     3961 gggaagcgga ggaggaggtt cgatgaccga atacaagctg gtggtggtgg gcgccgtcgg
     4021 tgtgggcaag agtgcgctga ccatccagct gatccagaac cattttgtgg acgaatacga
     4081 ccccactata gaggattcct accggaagca ggtggtcatt gatggggaga cgtgcctgtt
     4141 ggacatcctg gataccgccg gccaggagga gtacagcgcc atgcgggacc agtacatgcg
     4201 caccggggag ggcttcctgt gtgtgtttgc catcaacaac accaagtctt ttgaggacat
     4261 ccaccagtac agggagcaga tcaaacgggt gaaggactcg gatgacgtgc ccatggtgct
     4321 ggtggggaac aagtgtgacc tggctgcacg cactgtggaa tctcggcagg ctcaggacct
     4381 cgcccgaagc tacggcatcc cctacatcga gacctcggcc aagacccggc agggagtgga
     4441 ggatgccttc tacacgttgg tgcgtgagat ccggcagcac aagctgcgga agctgaaccc
     4501 tcctgatgag agtggccccg gctgcatgag ctgcaagtgt gtgctctcct aacaacgccg
     4561 ataatcaacc tctggattac aaaatttgtg aaagattgac tggtattctt aactatgttg
     4621 ctccttttac gctatgtgga tacgctgctt taatgccttt gtatcatgct attgcttccc
     4681 gtatggcttt cattttctcc tccttgtata aatcctggtt gctgtctctt tatgaggagt
     4741 tgtggcccgt tgtcaggcaa cgtggcgtgg tgtgcactgt gtttgctgac gcaaccccca
     4801 ctggttgggg cattgccacc acctgtcagc tcctttccgg gactttcgct ttccccctcc
     4861 ctattgccac ggcggaactc atcgccgcct gccttgcccg ctgctggaca ggggctcggc
     4921 tgttgggcac tgacaattcc gtggtgttgt cggggaaatc atcgtccttt ccttggctgc
     4981 tcgcctgtgt tgccacctgg attctgcgcg ggacgtcctt ctgctacgtc ccttcggccc
     5041 tcaatccagc ggaccttcct tcccgcggcc tgctgccggc tctgcggcct cttccgcgtc
     5101 ttcgccttcg ccctcagacg agtcggatct ccctttgggc cgcctccccg ccgctcgctg
     5161 atatagggcc cggtaccttt aagaccaatg acttacaagg cagctgtaga tcttagccac
     5221 tttttaaaag aaaagggggg actggaaggg ctagctcact cccaacgaag acaagatctg
     5281 ctttttgctt gtactgggtc tctctggtta gaccagatct gagcctggga gctctctggc
     5341 taactaggga acccactgct taagcctcaa taaagcttgc cttgagtgct tcaagtagtg
     5401 tgtgcccgtc tgttgtgtga ctctggtaac tagagatccc tcagaccctt ttagtcagtg
     5461 tggaaaatct ctagcagtag tagttcatgt catcttatta ttcagtattt ataacttgca
     5521 aagaaatgaa tatcagagag tgagaggaac ttgtttattg cagcttataa tggttacaaa
     5581 taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt
     5641 ggtttgtcca aactcatcaa tgtatcttat catgtctggc tctagctatc ccgcccctaa
     5701 ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc catggctgac
     5761 taattttttt tatttatgca gaggccgagg ccgcctcggc ctctgagcta ttccagaagt
     5821 agtgaggagg cttttttgga ggcctaggga cgtacccaat tcgccctata gtgagtcgta
     5881 ttacgcgcgc tcactggccg tcgttttaca acgtcgtgac tgggaaaacc ctggcgttac
     5941 ccaacttaat cgccttgcag cacatccccc tttcgccagc tggcgtaata gcgaagaggc
     6001 ccgcaccgat cgcccttccc aacagttgcg cagcctgaat ggcgaatggg acgcgccctg
     6061 tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc
     6121 cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg
     6181 ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg
     6241 gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg
     6301 atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt
     6361 ccaaactgga acaacactca accctatctc ggtctattct tttgatttat aagggatttt
     6421 gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaattt
     6481 taacaaaata ttaacgctta caatttaggt ggcacttttc ggggaaatgt gcgcggaacc
     6541 cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc
     6601 tgataaatgc ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc
     6661 gcccttattc ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg
     6721 gtgaaagtaa aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat
     6781 ctcaacagcg gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc
     6841 acttttaaag ttctgctatg tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa
     6901 ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa
     6961 aagcatctta cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt
     7021 gataacactg cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct
     7081 tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat
     7141 gaagccatac caaacgacga gcgtgacacc acgatgcctg tagcaatggc aacaacgttg
     7201 cgcaaactat taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg
     7261 atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt
     7321 attgctgata aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg
     7381 ccagatggta agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg
     7441 gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg
     7501 tcagaccaag tttactcata tatactttag attgatttaa aacttcattt ttaatttaaa
     7561 aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt
     7621 tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttc
//
