LOCUS       sequence-244180-        7104 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses human SOX2 and mCherry via t2A linker..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7104)
  AUTHORS   Dhaliwal NK, Abatti LE, Mitchell JA
  TITLE     KLF4 protein stability regulated by interaction with pluripotency 
            transcription factors overrides transcriptional control.
  JOURNAL   Genes Dev. 2019 Jun 20. pii: gad.324319.119. doi: 
            10.1101/gad.324319.119.
  PUBMED    31221664
REFERENCE   2  (bases 1 to 7104)
  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: "Genes Dev. 
            2019 Jun 20. pii: gad.324319.119. doi: 10.1101/gad.324319.119."
COMMENT     Sequence Label: Sox2-t2A-mCherry
FEATURES             Location/Qualifiers
     source          1..7104
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        50..429
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        430..633
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     584..604
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     630..654
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             1689..1742
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             1749..2453
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     complement(1891..1909)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2159..2178
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     complement(2490..2509)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    2534..2582
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      2784..3212
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2871..2890)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3081..3102
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3221..3241)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3226..3555
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3406..3541
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3468..3487
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3622..4416
                     /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(3676..3695)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4286..4305
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4592..4713
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4629..4648)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4683..4702
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4762..4778)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4762..4778)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4775..4797)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4786..4802
                     /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(4810..4816,4817..4834,4835..4840))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4855..4876
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4993..5010)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5164..5752)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5244..5263)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5923..6714,6715..6783))
                     /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     6546..6565
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6784..6888)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6963..6982)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gttaggcgtt ttgcgctgct tcgcgatgta cgggccagat atacgcgttg acattgatta
       61 ttgactagtt attaatagta atcaattacg gggtcattag ttcatagccc atatatggag
      121 ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc
      181 ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac tttccattga
      241 cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca agtgtatcat
      301 atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc
      361 cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt agtcatcgct
      421 attaccatgg tgatgcggtt ttggcagtac atcaatgggc gtggatagcg gtttgactca
      481 cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg gcaccaaaat
      541 caacgggact ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat gggcggtagg
      601 cgtgtacggt gggaggtcta tataagcaga gctcgtttag tgaaccgtca gatcgcctgg
      661 agacgccatc cacgctgttt tgacctccat agaagacacc gggaccgatc cagcctccgg
      721 actctagaat gtacaacatg atggagacgg agctgaagcc gccgggcccg cagcaaactt
      781 cggggggcgg cggcggcaac tccaccgcgg cggcggccgg cggcaaccag aaaaacagcc
      841 cggaccgcgt caagcggccc atgaatgcct tcatggtgtg gtcccgcggg cagcggcgca
      901 agatggccca ggagaacccc aagatgcaca actcggagat cagcaagcgc ctgggcgccg
      961 agtggaaact tttgtcggag acggagaagc ggccgttcat cgacgaggct aagcggctgc
     1021 gagcgctgca catgaaggag cacccggatt ataaataccg gccccggcgg aaaaccaaga
     1081 cgctcatgaa gaaggataag tacacgctgc ccggcgggct gctggccccc ggcggcaata
     1141 gcatggcgag cggggtcggg gtgggcgccg gcctgggcgc gggcgtgaac cagcgcatgg
     1201 acagttacgc gcacatgaac ggctggagca acggcagcta cagcatgatg caggaccagc
     1261 tgggctaccc gcagcacccg ggcctcaatg cgcacggcgc agcgcagatg cagcccatgc
     1321 accgctacga cgtgagcgcc ctgcagtaca actccatgac cagctcgcag acctacatga
     1381 acggctcgcc cacctacagc atgtcctact cgcagcaggg cacccctggc atggctcttg
     1441 gctccatggg ttcggtggtc aagtccgagg ccagctccag cccccctgtg gttacctctt
     1501 cctcccactc cagggcgccc tgccaggccg gggacctccg ggacatgatc agcatgtatc
     1561 tccccggcgc cgaggtgccg gaacccgccg cccccagcag acttcacatg tcccagcact
     1621 accagagcgg cccggtgccc ggcacggcca ttaacggcac actgcccctc tcacacatgt
     1681 tggctagcga gggcagagga agtcttctaa catgcggtga cgtggaggag aatcccggcc
     1741 ctggtaccgt gagcaagggc gaggaggata acatggccat catcaaggag ttcatgcgct
     1801 tcaaggtgca catggagggc tccgtgaacg gccacgagtt cgagatcgag ggcgagggcg
     1861 agggccgccc ctacgagggc acccagaccg ccaagctgaa ggtgaccaag ggtggccccc
     1921 tgcccttcgc ctgggacatc ctgtcccctc agttcatgta cggctccaag gcctacgtga
     1981 agcaccccgc cgacatcccc gactacttga agctgtcctt ccccgagggc ttcaagtggg
     2041 agcgcgtgat gaacttcgag gacggcggcg tggtgaccgt gacccaggac tcctccctgc
     2101 aggacggcga gttcatctac aaggtgaagc tgcgcggcac caacttcccc tccgacggcc
     2161 ccgtaatgca gaagaagacc atgggctggg aggcctcctc cgagcggatg taccccgagg
     2221 acggcgccct gaagggcgag atcaagcaga ggctgaagct gaaggacggc ggccactacg
     2281 acgctgaggt caagaccacc tacaaggcca agaagcccgt gcagctgccc ggcgcctaca
     2341 acgtcaacat caagttggac atcacctccc acaacgagga ctacaccatc gtggaacagt
     2401 acgaacgcgc cgagggccgc cactccaccg gcggcatgga cgagctgtac aagaccggtt
     2461 agtaatgagt ttaaacgggg gaggctaact gaaacacgga aggagacaat accggaagga
     2521 acccgcgcta tgacggcaat aaaaagacag aataaaacgc acgggtgttg ggtcgtttgt
     2581 tcataaacgc ggggttcggt cccagggctg gcactctgtc gataccccac cgagacccca
     2641 ttggggccaa tacgcccgcg tttcttcctt ttccccaccc caccccccaa gttcgggtga
     2701 aggcccaggg ctcgcagcca acgtcggggc ggcaggccct gccatagcag atctgcgcag
     2761 ctggggctct agggggtatc cccacgcgcc ctgtagcggc gcattaagcg cggcgggtgt
     2821 ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc
     2881 tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg
     2941 catcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta
     3001 gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt
     3061 ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat
     3121 ctcggtctat tcttttgatt tataagggat tttggggatt tcggcctatt ggttaaaaaa
     3181 tgagctgatt taacaaaaat ttaacgcgaa ttaattctgt ggaatgtgtg tcagttaggg
     3241 tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag
     3301 tcagcaacca ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg
     3361 catctcaatt agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact
     3421 ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag
     3481 gccgaggccg cctctgcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc
     3541 ctaggctttt gcaaaaagct cccgggagct tgtatatcca ttttcggatc tgatcaagag
     3601 acaggatgag gatcgtttcg catgattgaa caagatggat tgcacgcagg ttctccggcc
     3661 gcttgggtgg agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat
     3721 gccgccgtgt tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg
     3781 tccggtgccc tgaatgaact gcaggacgag gcagcgcggc tatcgtggct ggccacgacg
     3841 ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta
     3901 ttgggcgaag tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta
     3961 tccatcatgg ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc
     4021 gaccaccaag cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc
     4081 gatcaggatg atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg
     4141 ctcaaggcgc gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg
     4201 ccgaatatca tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt
     4261 gtggcggacc gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc
     4321 ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc
     4381 atcgccttct atcgccttct tgacgagttc ttctgagcgg gactctgggg ttcgcgaaat
     4441 gaccgaccaa gcgacgccca acctgccatc acgagatttc gattccaccg ccgccttcta
     4501 tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg
     4561 ggatctcatg ctggagttct tcgcccaccc caacttgttt attgcagctt ataatggtta
     4621 caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag
     4681 ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tgtataccgt cgacctctag
     4741 ctagagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac
     4801 aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt
     4861 gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc
     4921 gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg
     4981 ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt
     5041 atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa
     5101 gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
     5161 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
     5221 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
     5281 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
     5341 aagcgtggcg ctttctcaat gctcacgctg taggtatctc agttcggtgt aggtcgttcg
     5401 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
     5461 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
     5521 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
     5581 gcctaactac ggctacacta gaaggacagt atttggtatc tgcgctctgc tgaagccagt
     5641 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
     5701 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc
     5761 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt
     5821 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt
     5881 taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag
     5941 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt
     6001 cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg caatgatacc
     6061 gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc
     6121 cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg
     6181 ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac
     6241 aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg
     6301 atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc
     6361 tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact
     6421 gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc
     6481 aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat
     6541 acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc
     6601 ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac
     6661 tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa
     6721 aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact
     6781 catactcttc ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg
     6841 atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg
     6901 aaaagtgcca cctgacgtcg acggatcggg agatctcccg atcccctatg gtcgactctc
     6961 agtacaatct gctctgatgc cgcatagtta agccagtatc tgctccctgc ttgtgtgttg
     7021 gaggtcgctg agtagtgcgc gagcaaaatt taagctacaa caaggcaagg cttgaccgac
     7081 aattgcatga agaatctgct tagg
//
