LOCUS       sequence-203240-        7438 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral expression of Foxa1 DNA-binding domain (exon 2) targeting
            sgRNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7438)
  AUTHORS   Roe JS, Hwang CI, Somerville TDD, Milazzo JP, Lee EJ, Da Silva B, 
            Maiorino L, Tiriac H, Young CM, Miyabayashi K, Filippini D, 
            Creighton B, Burkhart RA, Buscaglia JM, Kim EJ, Grem JL, Lazenby AJ,
            Grunkemeyer JA, Hollingsworth MA, Grandgenett PM, Egeblad M, Park Y,
            Tuveson DA, Vakoc CR
  TITLE     Enhancer Reprogramming Promotes Pancreatic Cancer Metastasis.
  JOURNAL   Cell. 2017 Aug 24;170(5):875-888.e20. doi: 
            10.1016/j.cell.2017.07.007. Epub 2017 Jul 27.
  PUBMED    28757253
REFERENCE   2  (bases 1 to 7438)
  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"; doi: 
            "10.1016/j.cell.2017.07.007"; journalName: "Cell"; date: 
            "2017-08-24- 24"; volume: "170"; issue: "5"; pages: "875-888.e20"
COMMENT     Sequence Label: LRG/Foxa1 e2.4
FEATURES             Location/Qualifiers
     source          1..7438
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..227
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             228..408
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    455..580
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1073..1306
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1491..1535
                     /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"
     promoter        1713..1961
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     1713..1733
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     1884..1903
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        1983..2058
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     misc_feature    2106..2223
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2292..2503
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      2523..2532
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2532..2555
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             join(2556..2558,2559..2561,2562..3275)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(2556..2558,2559..2561,2562..3272)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2601..2622)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2862..2881)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3209..3230
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    3291..3879
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3344..3364)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3762..3773)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             3951..4184
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    4262..4383
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4299..4318)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4353..4372
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      4423..4558
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(4578..4597)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4579..4597)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4607..4624)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4607..4623)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4616..4638)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4765..5220
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4852..4871)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5062..5083
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5246..5350
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5351..5419,5420..6211)
                     /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(5569..5588)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6382..6970
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6871..6890
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7124..7141
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7258..7279
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7294..7299,7300..7317,7318..7324)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7332..7348
                     /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     7337..7359
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7356..7372
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7356..7372
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     7391..7411
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        7393..7411
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
ORIGIN
        1 tgtagtctta tgcaatactc ttgtagtctt gcaacatggt aacgatgagt tagcaacatg
       61 ccttacaagg agagaaaaag caccgtgcat gccgattggt ggaagtaagg tggtacgatc
      121 gtgccttatt aggaaggcaa cagacgggtc tgacatggat tggacgaacc actgaattgc
      181 cgcattgcag agatattgta tttaagtgcc tagctcgata cataaacggg tctctctggt
      241 tagaccagat ctgagcctgg gagctctctg gctaactagg gaacccactg cttaagcctc
      301 aataaagctt gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt gactctggta
      361 actagagatc cctcagaccc ttttagtcag tgtggaaaat ctctagcagt ggcgcccgaa
      421 cagggacttg aaagcgaaag ggaaaccaga ggagctctct cgacgcagga ctcggcttgc
      481 tgaagcgcgc acggcaagag gcgaggggcg gcgactggtg agtacgccaa aaattttgac
      541 tagcggaggc tagaaggaga gagatgggtg cgagagcgtc agtattaagc gggggagaat
      601 tagatcgcga tgggaaaaaa ttcggttaag gccaggggga aagaaaaaat ataaattaaa
      661 acatatagta tgggcaagca gggagctaga acgattcgca gttaatcctg gcctgttaga
      721 aacatcagaa ggctgtagac aaatactggg acagctacaa ccatcccttc agacaggatc
      781 agaagaactt agatcattat ataatacagt agcaaccctc tattgtgtgc atcaaaggat
      841 agagataaaa gacaccaagg aagctttaga caagatagag gaagagcaaa acaaaagtaa
      901 gaccaccgca cagcaagcgg ccgctgatct tcagacctgg aggaggagat atgagggaca
      961 attggagaag tgaattatat aaatataaag tagtaaaaat tgaaccatta ggagtagcac
     1021 ccaccaaggc aaagagaaga gtggtgcaga gagaaaaaag agcagtggga ataggagctt
     1081 tgttccttgg gttcttggga gcagcaggaa gcactatggg cgcagcgtca atgacgctga
     1141 cggtacaggc cagacaatta ttgtctggta tagtgcagca gcagaacaat ttgctgaggg
     1201 ctattgaggc gcaacagcat ctgttgcaac tcacagtctg gggcatcaag cagctccagg
     1261 caagaatcct ggctgtggaa agatacctaa aggatcaaca gctcctgggg atttggggtt
     1321 gctctggaaa actcatttgc accactgctg tgccttggaa tgctagttgg agtaataaat
     1381 ctctggaaca gatttggaat cacacgacct ggatggagtg ggacagagaa attaacaatt
     1441 acacaagctt aatacactcc ttaattgaag aatcgcaaaa ccagcaagaa aagaatgaac
     1501 aagaattatt ggaattagat aaatgggcaa gtttgtggaa ttggtttaac ataacaaatt
     1561 ggctgtggta tataaaatta ttcataatga tagtaggagg cttggtaggt ttaagaatag
     1621 tttttgctgt actttctata gtgaatagag ttaggcaggg atattcacca ttatcgtttc
     1681 agacccacct cccaaccccg aggggaccca gagagggcct atttcccatg attccttcat
     1741 atttgcatat acgatacaag gctgttagag agataattag aattaatttg actgtaaaca
     1801 caaagatatt agtacaaaat acgtgacgta gaaagtaata atttcttggg tagtttgcag
     1861 ttttaaaatt atgttttaaa atggactatc atatgcttac cgtaacttga aagtatttcg
     1921 atttcttggc tttatatatc ttgtggaaag gacgaaacac cgcgctgagc gagatctacc
     1981 aggttttaga gctagaaata gcaagttaaa ataaggctag tccgttatca acttgaaaaa
     2041 gtggcaccga gtcggtgctt ttttgaattc tagatcttga gacaaatggc agtattcatc
     2101 cacaatttta aaagaaaagg ggggattggg gggtacagtg caggggaaag aatagtagac
     2161 ataatagcaa cagacataca aactaaagaa ttacaaaaac aaattacaaa aattcaaaat
     2221 tttcgggttt attacaggga cagcagagat ccactttggc gccggctcga gtggctccgg
     2281 tgcccgtcag tgggcagagc gcacatcgcc cacagtcccc gagaagttgg ggggaggggt
     2341 cggcaattga accggtgcct agagaaggtg gcgcggggta aactgggaaa gtgatgtcgt
     2401 gtactggctc cgcctttttc ccgagggtgg gggagaaccg tatataagtg cagtagtcgc
     2461 cgtgaacgtt ctttttcgca acgggtttgc cgccagaaca caggtgtcgt gacgcgggat
     2521 ccgccaccat ggattacaaa gacgatgacg ataagatggt gagcaagggc gaggagctgt
     2581 tcaccggggt ggtgcccatc ctggtcgagc tggacggcga cgtaaacggc cacaagttca
     2641 gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa gctgaccctg aagttcatct
     2701 gcaccaccgg caagctgccc gtgccctggc ccaccctcgt gaccaccctg acctacggcg
     2761 tgcagtgctt cagccgctac cccgaccaca tgaagcagca cgacttcttc aagtccgcca
     2821 tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa ggacgacggc aactacaaga
     2881 cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa ccgcatcgag ctgaagggca
     2941 tcgacttcaa ggaggacggc aacatcctgg ggcacaagct ggagtacaac tacaacagcc
     3001 acaacgtcta tatcatggcc gacaagcaga agaacggcat caaggtgaac ttcaagatcc
     3061 gccacaacat cgaggacggc agcgtgcagc tcgccgacca ctaccagcag aacaccccca
     3121 tcggcgacgg ccccgtgctg ctgcccgaca accactacct gagcacccag tccgccctga
     3181 gcaaagaccc caacgagaag cgcgatcaca tggtcctgct ggagttcgtg accgccgccg
     3241 ggatcactct cggcatggac gagctgtaca agtagacgcg ttaagtcgac aatcaacctc
     3301 tggattacaa aatttgtgaa agattgactg gtattcttaa ctatgttgct ccttttacgc
     3361 tatgtggata cgctgcttta atgcctttgt atcatgctat tgcttcccgt atggctttca
     3421 ttttctcctc cttgtataaa tcctggttgc tgtctcttta tgaggagttg tggcccgttg
     3481 tcaggcaacg tggcgtggtg tgcactgtgt ttgctgacgc aacccccact ggttggggca
     3541 ttgccaccac ctgtcagctc ctttccggga ctttcgcttt ccccctccct attgccacgg
     3601 cggaactcat cgccgcctgc cttgcccgct gctggacagg ggctcggctg ttgggcactg
     3661 acaattccgt ggtgttgtcg gggaaatcat cgtcctttcc ttggctgctc gcctgtgttg
     3721 ccacctggat tctgcgcggg acgtccttct gctacgtccc ttcggccctc aatccagcgg
     3781 accttccttc ccgcggcctg ctgccggctc tgcggcctct tccgcgtctt cgccttcgcc
     3841 ctcagacgag tcggatctcc ctttgggccg cctccccgcg tcgactttaa gaccaatgac
     3901 ttacaaggca gctgtagatc ttagccactt tttaaaagaa aaggggggac tggaagggct
     3961 aattcactcc caacgaagac aagatctgct ttttgcttgt actgggtctc tctggttaga
     4021 ccagatctga gcctgggagc tctctggcta actagggaac ccactgctta agcctcaata
     4081 aagcttgcct tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta
     4141 gagatccctc agaccctttt agtcagtgtg gaaaatctct agcagtacgt atagtagttc
     4201 atgtcatctt attattcagt atttataact tgcaaagaaa tgaatatcag agagtgagag
     4261 gaacttgttt attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac
     4321 aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc
     4381 ttatcatgtc tggctctagc tatcccgccc ctaactccgc ccatcccgcc cctaactccg
     4441 cccagttccg cccattctcc gccccatggc tgactaattt tttttattta tgcagaggcc
     4501 gaggccgcct cggcctctga gctattccag aagtagtgag gaggcttttt tggaggccta
     4561 gggacgtacc caattcgccc tatagtgagt cgtattacgc gcgctcactg gccgtcgttt
     4621 tacaacgtcg tgactgggaa aaccctggcg ttacccaact taatcgcctt gcagcacatc
     4681 cccctttcgc cagctggcgt aatagcgaag aggcccgcac cgatcgccct tcccaacagt
     4741 tgcgcagcct gaatggcgaa tgggacgcgc cctgtagcgg cgcattaagc gcggcgggtg
     4801 tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg
     4861 ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg
     4921 ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt
     4981 agggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt
     5041 tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta
     5101 tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa
     5161 atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg cttacaattt
     5221 aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca
     5281 ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa
     5341 aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt
     5401 ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca
     5461 gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag
     5521 ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc
     5581 ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac actattctca
     5641 gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt
     5701 aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct
     5761 gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt
     5821 aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga
     5881 caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact
     5941 tactctagct tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc
     6001 acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga
     6061 gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt
     6121 agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga
     6181 gataggtgcc tcactgatta agcattggta actgtcagac caagtttact catatatact
     6241 ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga
     6301 taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt
     6361 agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca
     6421 aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct
     6481 ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc ttctagtgta
     6541 gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct
     6601 aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc
     6661 aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca
     6721 gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga
     6781 aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg
     6841 aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt
     6901 cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag
     6961 cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt
     7021 tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt
     7081 tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga
     7141 ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc cgattcatta
     7201 atgcagctgg cacgacaggt ttcccgactg gaaagcgggc agtgagcgca acgcaattaa
     7261 tgtgagttag ctcactcatt aggcacccca ggctttacac tttatgcttc cggctcgtat
     7321 gttgtgtgga attgtgagcg gataacaatt tcacacagga aacagctatg accatgatta
     7381 cgccaagcgc gcaattaacc ctcactaaag ggaacaaaag ctggagctgc aagcttaa
//
