LOCUS       sequence-326711-        5507 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  HDR knock-in template for tagging the human endogenous SOX2 gene 
            with a tagBFP fluorescent marker linked by a self-cleaving P2A 
            peptide..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5507)
  AUTHORS   Abatti LE, Lado-Fernandez P, Huynh L, Collado M, Hoffman MM, 
            Mitchell JA
  TITLE     Epigenetic reprogramming of a distal developmental enhancer cluster 
            drives SOX2 overexpression in breast and lung adenocarcinoma.
  JOURNAL   Nucleic Acids Res. 2023 Sep 22:gkad734. doi: 10.1093/nar/gkad734.
  PUBMED    37738673
REFERENCE   2  (bases 1 to 5507)
  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: "Nucleic 
            Acids Res. 2023 Sep 22:gkad734. doi: 10.1093/nar/gkad734."
COMMENT     Sequence Label: SOX2-P2A-tagBFP-HDR
FEATURES             Location/Qualifiers
     source          1..5507
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     305..324
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        305..323
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1348..1404
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             1405..2103
                     /codon_start=1
                     /product="monomeric blue fluorescent protein"
                     /label=TagBFP
                     /note="mammalian codon-optimized"
                     /translation="SELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMRIKVV
                     EGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLTAT
                     QDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDMALKL
                     VGGSHLIANIKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVARYCD
                     LPSKLGHKLN"
     promoter        complement(join(3339..3345,3346..3363,3364..3369))
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    3384..3405
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3525..3542)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3696..4284)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3776..3795)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4455..5246,5247..5315))
                     /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     5078..5097
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5316..5420)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5488..5506
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 gcccctgcag ccgaattata ttatttttgc caaataattt ttaacaaaag ctctgaagtc
       61 ttcttcattt aaattcttag atgatacttc atctggaaaa ttgtcccaat tagtagcatc
      121 acgctgtgag taagttctaa accatttttt tattgttgta ttatctctaa tcttactact
      181 cgatgagttt tcggtattat ctctattttt aacttggagc aggttccatt cattgttttt
      241 ttcatcatag tgaataaaat caactgcttt aacacttgtg cctgaacacc atatccatcc
      301 ggcgtaatac gactcactat agggagagcg gccgccagat cttccggatg gctcgagttt
      361 ttcagcaaga tccggacagc gaactggagg ggggacatga tggagacgga gctgaagccg
      421 ccgggcccgc agcaaacttc ggggggcggc ggcggcaact ccaccgcggc ggcggccggc
      481 ggcaaccaga aaaacagccc ggaccgcgtc aagcggccca tgaatgcctt catggtgtgg
      541 tcccgcgggc agcggcgcaa gatggcccag gagaacccca agatgcacaa ctcggagatc
      601 agcaagcgcc tgggcgccga gtggaaactt ttgtcggaga cggagaagcg gccgttcatc
      661 gacgaggcta agcggctgcg agcgctgcac atgaaggagc acccggatta taaataccgg
      721 ccccggcgga aaaccaagac gctcatgaag aaggataagt acacgctgcc cggcgggctg
      781 ctggcccccg gcggcaatag catggcgagc ggggtcgggg tgggcgccgg cctgggcgcg
      841 ggcgtgaacc agcgcatgga cagttacgcg cacatgaacg gctggagcaa cggcagctac
      901 agcatgatgc aggaccagct gggctacccg cagcacccgg gcctcaatgc gcacggcgca
      961 gcgcagatgc agcccatgca ccgctacgac gtgagcgccc tgcagtacaa ctccatgacc
     1021 agctcgcaga cctacatgaa cggctcgccc acctacagca tgtcctactc gcagcagggc
     1081 acccctggca tggctcttgg ctccatgggt tcggtggtca agtccgaggc cagctccagc
     1141 ccccctgtgg ttacctcttc ctcccactcc agggcgccct gccaggccgg ggacctccgg
     1201 gacatgatca gcatgtatct ccccggcgcc gaggtgccgg aacccgccgc ccccagcaga
     1261 cttcacatgt cccagcacta ccagagcggc ccggtgcccg gcacggccat taacggcaca
     1321 ctgcccctct cacacatggg aagcggagct actaacttca gcctgctgaa gcaggctgga
     1381 gacgtggagg agaaccctgg acctagcgag ctgattaagg agaacatgca catgaagctg
     1441 tacatggagg gcaccgtgga caaccatcac ttcaagtgca catccgaggg cgaaggcaag
     1501 ccctacgagg gcacccagac catgagaatc aaggtggtcg agggcggccc tctccccttc
     1561 gccttcgaca tcctggctac tagcttcctc tacggcagca agaccttcat caaccacacc
     1621 cagggcatcc ccgacttctt caagcagtcc ttccctgagg gcttcacatg ggagagagtc
     1681 accacatacg aagacggggg cgtgctgacc gctacccagg acaccagcct ccaggacggc
     1741 tgcctcatct acaacgtcaa gatcagaggg gtgaacttca catccaacgg ccctgtgatg
     1801 cagaagaaaa cactcggctg ggaggccttc accgagacgc tgtaccccgc tgacggcggc
     1861 ctggaaggca gaaacgacat ggccctgaag ctcgtgggcg ggagccatct gatcgcaaac
     1921 atcaagacca catatagatc caagaaaccc gctaagaacc tcaagatgcc tggcgtctac
     1981 tatgtggact acagactgga aagaatcaag gaggccaaca acgagaccta cgtcgagcag
     2041 cacgaggtgg cagtggccag atactgcgac ctccctagca aactggggca caagcttaat
     2101 tagtaaggag aaattttcaa agaaaaacga gggaaatggg aggggtgcaa aagaggagag
     2161 taagaaacag catggagaaa acccggtacg ctcaaaaaga aaaaggaaaa aaaaaaatcc
     2221 catcacccac agcaaatgac agctgcaaaa gagaacacca atcccatcca cactcacgca
     2281 aaaaccgcga tgccgacaag aaaactttta tgagagagat cctggacttc tttttggggg
     2341 actatttttg tacagagaaa acctggggag ggtggggagg gcgggggaat ggaccttgta
     2401 tagatctgga ggaaagaaag ctacgaaaaa ctttttaaaa gttctagtgg tacggtagga
     2461 gctttgcagg aagtttgcaa aagtctttac caataatatt tagagctagt ctccaagcga
     2521 cgaaaaaaat gttttaatat ttgcaagcaa cttttgtaca gtatttatcg agataaacat
     2581 ggcaatcaaa atgtccattg tttataagct gagaatttgc caatattttt caaggagagg
     2641 cttcttgctg aattttgatt ctgcagctga aatttaggac agttgcaaac gtgaaaagaa
     2701 gaaaattatt caaatttgga cattttaatt gtttaaaaat tgtacaaaag gaaaaaatta
     2761 gaataagtac tggcgaacca tctctgtggt cttgtttaaa aagggcaaaa gttttagact
     2821 gtactaaatt ttataactta ctgttaaaag caaaaatggc catgcaggtt gacaccgttg
     2881 gcccccctcc agttcgctgt ccggatcttt ctagaagatc tcctacaata ttctcagctg
     2941 ccatggaaaa tcgatgttct tcttttattc tctcaagatt ttcaggctgt atattaaaac
     3001 ttatattaag aactatgcta accacctcat caggaaccgt tgtaggtggc gtgggttttc
     3061 ttggcaatcg actctcatga aaactacgag ctaaatattc aatatgttcc tcttgaccaa
     3121 ctttattctg catttttttt gaacgaggtt tagagcaagc ttcaggaaac tgagacagga
     3181 attttattaa aaatttaaat tttgaagaaa gttcagggtt aatagcatcc attttttgct
     3241 ttgcaagttc ctcagcattc ttaacaaaag acgtctcttt tgacatgttt aaagtttaaa
     3301 cctcctgtgt gaaattatta tccgctcata attccacaca ttatacgagc cggaagcata
     3361 aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca
     3421 ctgccaattg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa
     3481 cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg
     3541 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
     3601 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     3661 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     3721 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     3781 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     3841 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     3901 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     3961 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     4021 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     4081 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
     4141 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     4201 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     4261 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     4321 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     4381 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     4441 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     4501 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     4561 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
     4621 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     4681 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     4741 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
     4801 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     4861 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     4921 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     4981 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     5041 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
     5101 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     5161 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     5221 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     5281 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     5341 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     5401 aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc
     5461 attattatca tgacattaac ctataaaaat aggcgtatca cgaggcc
//
