LOCUS       sequence-224734-        8407 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Constitutive expression of a SOX2 protein tagged with SNAP-tag and a
            mitotic degron (MD).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8407)
  AUTHORS   Deluz C, Friman ET, Strebinger D, Benke A, Raccaud M, Callegari A, 
            Leleu M, Manley S, Suter DM
  TITLE     A role for mitotic bookmarking of SOX2 in pluripotency and 
            differentiation.
  JOURNAL   Genes Dev. 2016 Nov 15;30(22):2538-2550. doi: 
            10.1101/gad.289256.116. Epub 2016 Dec 5.
  PUBMED    27920086
REFERENCE   2  (bases 1 to 8407)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1101/gad.289256.116"; journalName: "Genes Dev"; date: 
            "2016-11-15- 15"; volume: "30"; issue: "22"; pages: "2538-2550"
COMMENT     Sequence Label: pLV-PGK-SNAP-MD-SOX2
FEATURES             Location/Qualifiers
     source          1..8407
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        21..531
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     450..468
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             546..1091
                     /codon_start=1
                     /product="human O6-alkylguanine-DNA-alkyltransferase 
                     variant that forms covalent bonds with benzylguanine 
                     derivatives"
                     /label=SNAP-tag(R)
                     /note="fast-reacting variant, optimized for mammalian 
                     expression"
                     /translation="MDKDCEMKRTTLDSPLGKLELSGCEQGLHRIIFLGKGTSAADAVE
                     VPAPAAVLGGPEPLMQATAWLNAYFHQPEAIEEFPVPALHHPVFQQESFTRQVLWKLLK
                     VVKFGEVISYSHLAALAGNPAATAAVKTALSGNPVPILIPCHRVVQGDLDVGGYEGGLA
                     VKEWLLAHEGHRLGKPGLG"
     misc_feature    2307..2895
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2360..2380)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2778..2789)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             2982..3215
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    3287..3408
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3324..3343)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3378..3397
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      3448..3583
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(3603..3622)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3604..3622)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3632..3649)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3632..3648)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3641..3663)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3790..4245
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3877..3896)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4087..4108
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4271..4375
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4376..4444,4445..5236)
                     /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(4594..4613)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5407..5995
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5896..5915
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6149..6166
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    6283..6304
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6319..6324,6325..6342,6343..6349)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6357..6373
                     /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     6362..6384
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6381..6397
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6381..6397
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     6416..6436
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        6418..6436
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        6464..6690
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             6691..6871
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    6918..7043
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    7536..7769
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             7954..7998
                     /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    8265..8382
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
ORIGIN
        1 agcttgatat cgaattccac ggggttgggg ttgcgccttt tccaaggcag ccctgggttt
       61 gcgcagggac gcggctgctc tgggcgtggt tccgggaaac gcagcggcgc cgaccctggg
      121 tctcgcacat tcttcacgtc cgttcgcagc gtcacccgga tcttcgccgc tacccttgtg
      181 ggccccccgg cgacgcttcc tgctccgccc ctaagtcggg aaggttcctt gcggttcgcg
      241 gcgtgccgga cgtgacaaac ggaagccgca cgtctcacta gtaccctcgc agacggacag
      301 cgccagggag caatggcagc gcgccgaccg cgatgggctg tggccaatag cggctgctca
      361 gcagggcgcg ccgagagcag cggccgggaa ggggcggtgc gggaggcggg gtgtggggcg
      421 gtagtgtggg ccctgttcct gcccgcgcgg tgttccgcat tctgcaagcc tccggagcgc
      481 acgtcggcag tcggctccct cgttgaccga atcaccgacc tctctcccca gggggatcca
      541 ccggtatgga caaagactgc gaaatgaagc gcaccaccct ggatagccct ctgggcaagc
      601 tggaactgtc tgggtgcgaa cagggcctgc accgtatcat cttcctgggc aaaggaacat
      661 ctgccgccga cgccgtggaa gtgcctgccc cagccgccgt gctgggcgga ccagagccac
      721 tgatgcaggc caccgcctgg ctcaacgcct actttcacca gcctgaggcc atcgaggagt
      781 tccctgtgcc agccctgcac cacccagtgt tccagcagga gagctttacc cgccaggtgc
      841 tgtggaaact gctgaaagtg gtgaagttcg gagaggtcat cagctacagc cacctggccg
      901 ccctggccgg caatcccgcc gccaccgccg ccgtgaaaac cgccctgagc ggaaatcccg
      961 tgcccattct gatcccctgc caccgggtgg tgcagggcga cctggacgtg gggggctacg
     1021 agggcgggct cgccgtgaaa gagtggctgc tggcccacga gggccacaga ctgggcaagc
     1081 ctgggctggg ttctagagca gaaaataagg ctaaagtctc aatggccggc gctaagcgcg
     1141 ttcccgtgac cgtaactgcc gcttctaaac ccgggttgcg cccgaggact gctcttggag
     1201 atattggcaa caaggtgtct gaagagctgc aggctcgcgt gccactgaag cgagaggcta
     1261 agacgctggg aactggaaag ggtacggtta aggccctgcc gaagccagtg gagaaagttc
     1321 ctgtctgcga accggctaga atgtataaca tgatggagac ggagctgaag ccgccgggcc
     1381 cgcagcaagc ttcggggggc ggcggcggag gaggcaacgc cacggcggcg gcgaccggcg
     1441 gcaaccagaa gaacagcccg gaccgcgtca agaggcccat gaacgccttc atggtatggt
     1501 cccgggggca gcggcgtaag atggcccagg agaaccccaa gatgcacaac tcggagatca
     1561 gcaagcgcct gggcgcggag tggaaacttt tgtccgagac cgagaagcgg ccgttcatcg
     1621 acgaggccaa gcggctgcgc gctctgcaca tgaaggagca cccggattat aaataccggc
     1681 cgcggcggaa aaccaagacg ctcatgaaga aggataagta cacgcttccc ggaggcttgc
     1741 tggcccccgg cgggaacagc atggcgagcg gggttggggt gggcgccggc ctgggtgcgg
     1801 gcgtgaacca gcgcatggac agctacgcgc acatgaacgg ctggagcaac ggcagctaca
     1861 gcatgatgca ggagcagctg ggctacccgc agcacccggg cctcaacgct cacggcgcgg
     1921 cacagatgca accgatgcac cgctacgacg tcagcgccct gcagtacaac tccatgacca
     1981 gctcgcagac ctacatgaac ggctcgccca cctacagcat gtcctactcg cagcagggca
     2041 cccccggtat ggcgctgggc tccatgggct ctgtggtcaa gtccgaggcc agctccagcc
     2101 cccccgtggt tacctcttcc tcccactcca gggcgccctg ccaggccggg gacctccggg
     2161 acatgatcag catgtacctc cccggcgccg aggtgccgga gcccgctgcg cccagtagac
     2221 tgcacatggc ccagcactac cagagcggcc cggtgcccgg cacggccatt aacggcacac
     2281 tgcccctgtc gcacatgtga gtcgacaatc aacctctgga ttacaaaatt tgtgaaagat
     2341 tgactggtat tcttaactat gttgctcctt ttacgctatg tggatacgct gctttaatgc
     2401 ctttgtatca tgctattgct tcccgtatgg ctttcatttt ctcctccttg tataaatcct
     2461 ggttgctgtc tctttatgag gagttgtggc ccgttgtcag gcaacgtggc gtggtgtgca
     2521 ctgtgtttgc tgacgcaacc cccactggtt ggggcattgc caccacctgt cagctccttt
     2581 ccgggacttt cgctttcccc ctccctattg ccacggcgga actcatcgcc gcctgccttg
     2641 cccgctgctg gacaggggct cggctgttgg gcactgacaa ttccgtggtg ttgtcgggga
     2701 agctgacgtc ctttccatgg ctgctcgcct gtgttgccac ctggattctg cgcgggacgt
     2761 ccttctgcta cgtcccttcg gccctcaatc cagcggacct tccttcccgc ggcctgctgc
     2821 cggctctgcg gcctcttccg cgtcttcgcc ttcgccctca gacgagtcgg atctcccttt
     2881 gggccgcctc cccgcctgga attcgagctc ggtaccttta agaccaatga cttacaaggc
     2941 agctgtagat cttagccact ttttaaaaga aaagggggga ctggaagggc taattcactc
     3001 ccaacgaaga caagatctgc tttttgcttg tactgggtct ctctggttag accagatctg
     3061 agcctgggag ctctctggct aactagggaa cccactgctt aagcctcaat aaagcttgcc
     3121 ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac tctggtaact agagatccct
     3181 cagacccttt tagtcagtgt ggaaaatctc tagcagtagt agttcatgtc atcttattat
     3241 tcagtattta taacttgcaa agaaatgaat atcagagagt gagaggaact tgtttattgc
     3301 agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt
     3361 ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctggct
     3421 ctagctatcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat
     3481 tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc cgcctcggcc
     3541 tctgagctat tccagaagta gtgaggaggc ttttttggag gcctagggac gtacccaatt
     3601 cgccctatag tgagtcgtat tacgcgcgct cactggccgt cgttttacaa cgtcgtgact
     3661 gggaaaaccc tggcgttacc caacttaatc gccttgcagc acatccccct ttcgccagct
     3721 ggcgtaatag cgaagaggcc cgcaccgatc gcccttccca acagttgcgc agcctgaatg
     3781 gcgaatggga cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca
     3841 gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct
     3901 ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt
     3961 tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac
     4021 gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct
     4081 ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt
     4141 ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac
     4201 aaaaatttaa cgcgaatttt aacaaaatat taacgcttac aatttaggtg gcacttttcg
     4261 gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc
     4321 gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag
     4381 tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt
     4441 tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt
     4501 gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga
     4561 acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat
     4621 tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga
     4681 gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag
     4741 tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg
     4801 accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg
     4861 ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt
     4921 agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg
     4981 gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc
     5041 ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg
     5101 tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac
     5161 ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact
     5221 gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa
     5281 acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa
     5341 aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg
     5401 atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc
     5461 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac
     5521 tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca
     5581 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt
     5641 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
     5701 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg
     5761 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
     5821 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
     5881 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
     5941 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
     6001 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt
     6061 tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac
     6121 cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg
     6181 cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt cattaatgca gctggcacga
     6241 caggtttccc gactggaaag cgggcagtga gcgcaacgca attaatgtga gttagctcac
     6301 tcattaggca ccccaggctt tacactttat gcttccggct cgtatgttgt gtggaattgt
     6361 gagcggataa caatttcaca caggaaacag ctatgaccat gattacgcca agcgcgcaat
     6421 taaccctcac taaagggaac aaaagctgga gctgcaagct taatgtagtc ttatgcaata
     6481 ctcttgtagt cttgcaacat ggtaacgatg agttagcaac atgccttaca aggagagaaa
     6541 aagcaccgtg catgccgatt ggtggaagta aggtggtacg atcgtgcctt attaggaagg
     6601 caacagacgg gtctgacatg gattggacga accactgaat tgccgcattg cagagatatt
     6661 gtatttaagt gcctagctcg atacataaac gggtctctct ggttagacca gatctgagcc
     6721 tgggagctct ctggctaact agggaaccca ctgcttaagc ctcaataaag cttgccttga
     6781 gtgcttcaag tagtgtgtgc ccgtctgttg tgtgactctg gtaactagag atccctcaga
     6841 cccttttagt cagtgtggaa aatctctagc agtggcgccc gaacagggac ttgaaagcga
     6901 aagggaaacc agaggagctc tctcgacgca ggactcggct tgctgaagcg cgcacggcaa
     6961 gaggcgaggg gcggcgactg gtgagtacgc caaaaatttt gactagcgga ggctagaagg
     7021 agagagatgg gtgcgagagc gtcagtatta agcgggggag aattagatcg cgatgggaaa
     7081 aaattcggtt aaggccaggg ggaaagaaaa aatataaatt aaaacatata gtatgggcaa
     7141 gcagggagct agaacgattc gcagttaatc ctggcctgtt agaaacatca gaaggctgta
     7201 gacaaatact gggacagcta caaccatccc ttcagacagg atcagaagaa cttagatcat
     7261 tatataatac agtagcaacc ctctattgtg tgcatcaaag gatagagata aaagacacca
     7321 aggaagcttt agacaagata gaggaagagc aaaacaaaag taagaccacc gcacagcaag
     7381 cggccgctga tcttcagacc tggaggagga gatatgaggg acaattggag aagtgaatta
     7441 tataaatata aagtagtaaa aattgaacca ttaggagtag cacccaccaa ggcaaagaga
     7501 agagtggtgc agagagaaaa aagagcagtg ggaataggag ctttgttcct tgggttcttg
     7561 ggagcagcag gaagcactat gggcgcagcg tcaatgacgc tgacggtaca ggccagacaa
     7621 ttattgtctg gtatagtgca gcagcagaac aatttgctga gggctattga ggcgcaacag
     7681 catctgttgc aactcacagt ctggggcatc aagcagctcc aggcaagaat cctggctgtg
     7741 gaaagatacc taaaggatca acagctcctg gggatttggg gttgctctgg aaaactcatt
     7801 tgcaccactg ctgtgccttg gaatgctagt tggagtaata aatctctgga acagatttgg
     7861 aatcacacga cctggatgga gtgggacaga gaaattaaca attacacaag cttaatacac
     7921 tccttaattg aagaatcgca aaaccagcaa gaaaagaatg aacaagaatt attggaatta
     7981 gataaatggg caagtttgtg gaattggttt aacataacaa attggctgtg gtatataaaa
     8041 ttattcataa tgatagtagg aggcttggta ggtttaagaa tagtttttgc tgtactttct
     8101 atagtgaata gagttaggca gggatattca ccattatcgt ttcagaccca cctcccaacc
     8161 ccgaggggac ccgacaggcc cgaaggaata gaagaagaag gtggagagag agacagagac
     8221 agatccattc gattagtgaa cggatctcga cggtatcggt taacttttaa aagaaaaggg
     8281 gggattgggg ggtacagtgc aggggaaaga atagtagaca taatagcaac agacatacaa
     8341 actaaagaat tacaaaaaca aattacaaaa attcaaaatt ttatcgatca cgagactagc
     8401 ctcgaga
//
