LOCUS       sequence-456531-        7378 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7378)
  AUTHORS   Mitsui K, Tokuzawa Y, Itoh H, Segawa K, Murakami M, Takahashi K, 
            Maruyama M, Maeda M, Yamanaka S
  TITLE     The homeoprotein Nanog is required for maintenance of pluripotency 
            in mouse epiblast and ES cells.
  JOURNAL   Cell. 2003 May 30. 113(5):631-42.
  PUBMED    12787504
REFERENCE   2  (bases 1 to 7378)
  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: "Cell. 2003 
            May 30. 113(5):631-42."
COMMENT     Sequence Label: pPyCAG-Nanog-IP
FEATURES             Location/Qualifiers
     source          1..7378
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    576..710
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     enhancer        724..1103
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1105..1382
                     /label=chicken beta-actin promoter
     intron          1383..2399
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     2323..2345
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     2407..2426
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     protein_bind    2456..2480
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(3409..3433)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_feature    join(3451..4025,4026..4028,4029..4037)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3618..3635)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3845..3864
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             4038..4637
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(4038..4057)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4534..4554
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     complement(4718..4735)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4724..4948
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4995..5011)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4995..5011)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5008..5030)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5019..5035
                     /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(5043..5049,5050..5067,5068..5073))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5088..5109
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        5168..5363
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      5214..5349
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    5369..5503
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5571..5588)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5742..6330)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5822..5841)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6501..7292,7293..7361))
                     /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     7124..7143
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 gaccgctttg gaaaaacaaa gactgtattt cctggaaatt aatgtttatt caataaactg
       61 tgtattcagc tatattcaca tagtggtgag gctgaaatga ggcgggaaga ggcggttggg
      121 gcttaattat atcaatttgg gtggccccac agcgcctcca aggcgccagt cctgttttga
      181 caagttgcct ctggaagcct ctcttctttt tctccagagt aagcggaggc caggggcccc
      241 cggcctctgc ttaatactaa aaaaaacagc tgttgtcata gtaatgattg ggtggaaaca
      301 ttccaggcct gggtggagag gctttttgct tcctcttgca aaaccacact gacattccag
      361 gcctgggtgg agaggctttt tgcttcctct tgcaaaacca cactgccctc tggagggcag
      421 ttgcctagca actaattaaa agaggatgtc gcacggccag ctgcggtcag ttagtcactt
      481 cctgcttaac tgacttgaca ttttctattt taagagtcgg gaggaaaatt actgtgttgg
      541 aggccctccg ccatcttctg aagctgaatc gaattaactt gtttattgca gcttataatg
      601 gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt
      661 ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctggatc tgatatcatc
      721 gtcgacattg attattgact agttattaat agtaatcaat tacggggtca ttagttcata
      781 gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc
      841 ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag
      901 ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac
      961 atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg
     1021 cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg
     1081 tattagtcat cgctattacc atggtcgagg tgagccccac gttctgcttc actctcccca
     1141 tctccccccc ctccccaccc ccaattttgt atttatttat tttttaatta ttttgtgcag
     1201 cgatgggggc gggggggggg ggggggcgcg cgccrggsgg ggsggggsgg ggsgrggggs
     1261 ggggcggggc gaggcggaga ggtgcggcgg cagccaatca gagcggcgcg ctccgaaagt
     1321 ttccttttat ggcgaggcgg cggcggcggc ggccctataa aaagcgaagc gcgcggcggg
     1381 cgggagtcgc tgcgcgctgc cttcgccccg tgccccgctc cgccgccgcc tcgcgccgcc
     1441 cgccccggct ctgactgacc gcgttactcc cacaggtgag cgggcgggac ggcccttctc
     1501 ctccgggctg taattagcgc ttggtttaat gacggcttgt ttcttttctg tggctgcgtg
     1561 aaagccttga ggggctccgg gagggccctt tgtgcggggg gagcggctcg gggggtgcgt
     1621 gcgtgtgtgt gtgcgtgggg agcgccgcgt gcggctccgc gctgcccggc ggctgtgagc
     1681 gctgcgggcg cggcgcgggg ctttgtgcgc tccgcagtgt gcgcgagggg agcgcggccg
     1741 ggggcggtgc cccgcggtgc ggggggggct gcgaggggaa caaaggctgc gtgcggggtg
     1801 tgtgcgtggg ggggtgagca gggggtgtgg gcgcgtcggt cgggctgcaa ccccccctgc
     1861 acccccctcc ccgagttgct gagcacggcc cggcttcggg tgcggggctc cgtacggggc
     1921 gtggcgcggg gctcgccgtg ccgggcgggg ggtggcggca ggtgggggtg ccgggcgggg
     1981 cggggccgcc tcgggccggg gagggctcgg gggaggggcg cggcggcccc cggagcgccg
     2041 gcggctgtcg aggcgcggcg agccgcagcc attgcctttt atggtaatcg tgcgagaggg
     2101 cgcagggact tcctttgtcc caaatctgtg cggagccgaa atctgggagg cgccgccgca
     2161 ccccctctag cgggcgcggg gcgaagcggt gcggcgccgg caggaaggaa atgggcgggg
     2221 agggccttcg tgcgtcgccg cgccgccgtc cccttctccc tctccagcct cggggctgtc
     2281 cgcgggggga cggctgcctt cgggggggac ggggcagggc ggggttcggc ttctggcgtg
     2341 tgaccggcgg ctctagagcc tctgctaacc atgttcatgc cttcttcttt ttcctacagc
     2401 tcctgggcaa cgtgctggtt attgtgctgt ctcatcattt tggcaaagaa ttatcacaag
     2461 tttgtacaaa aaagcaggct ctgacatgag tgtgggtctt cctggtcccc acagtttgcc
     2521 tagttctgag gaagcatcga attctgggaa cgcctcatca atgcctgcag tttttcatcc
     2581 cgagaactat tcttgcttac aagggtctgc tactgagatg ctctgcacag aggctgcctc
     2641 tcctcgccct tcctctgaag acctgcctct tcaaggcagc cctgattctt ctaccagtcc
     2701 caaacaaaag ctctcaagtc ctgaggctga caagggccct gaggaggagg agaacaaggt
     2761 ccttgccagg aagcagaaga tgcggactgt gttctctcag gcccagctgt gtgcactcaa
     2821 ggacaggttt cagaagcaga agtacctcag cctccagcag atgcaagaac tctcctccat
     2881 tctgaacctg agctataagc aggttaagac ctggtttcaa aaccaaagga tgaagtgcaa
     2941 gcggtggcag aaaaaccagt ggttgaagac tagcaatggt ctgattcaga agggctcagc
     3001 accagtggag tatcccagca tccattgcag ctatccccag ggctatctgg tgaacgcatc
     3061 tggaagcctt tccatgtggg gcagccagac ttggaccaac ccaacttgga gcagccagac
     3121 ctggaccaac ccaacttgga acaaccagac ctggaccaac ccaacttgga gcagccaggc
     3181 ctggaccgct cagtcctgga acggccagcc ttggaatgct gctccgctcc ataacttcgg
     3241 ggaggacttt ctgcagcctt acgtacagtt gcagcaaaac ttctctgcca gtgatttgga
     3301 ggtgaatttg gaagccacta gggaaagcca tgcgcatttt agcaccccac aagccttgga
     3361 attattcctg aactactctg tgactccacc aggtgaaata tgagacttac ccagctttct
     3421 tgtacaaagt ggtgataatt ccgccccccc cccctctccc tccccccccc ctaacgttac
     3481 tggccgaagc cgcttggaat aaggccggtg tgcgtttgtc tatatgttat tttccaccat
     3541 attgccgtct tttggcaatg tgagggcccg gaaacctggc cctgtcttct tgacgagcat
     3601 tcctaggggt ctttcccctc tcgccaaagg aatgcaaggt ctgttgaatg tcgtgaagga
     3661 agcagttcct ctggaagctt cttgaagaca aacaacgtct gtagcgaccc tttgcaggca
     3721 gcggaacccc ccacctggcg acaggtgcct ctgcggccaa aagccacgtg tataagatac
     3781 acctgcaaag gcggcacaac cccagtgcca cgttgtgagt tggatagttg tggaaagagt
     3841 caaatggctc tcctcaagcg tattcaacaa ggggctgaag gatgcccaga aggtacccca
     3901 ttgtatggga tctgatctgg ggcctcggtg cacatgcttt acatgtgttt agtcgaggtt
     3961 aaaaaaacgt ctaggccccc cgaaccacgg ggacgtggtt ttcctttgaa aaacacgatg
     4021 ataatatggc cacaaccatg accgagtaca agcccacggt gcgcctcgcc acccgcgacg
     4081 acgtccccag ggccgtacgc accctcgccg ccgcgttcgc cgactacccc gccacgcgcc
     4141 acaccgtcga tccggaccgc cacatcgagc gggtcaccga gctgcaagaa ctcttcctca
     4201 cgcgcgtcgg gctcgacatc ggcaaggtgt gggtcgcgga cgacggcgcc gcggtggcgg
     4261 tctggaccac gccggagagc gtcgaagcgg gggcggtgtt cgccgagatc ggcccgcgca
     4321 tggccgagtt gagcggttcc cggctggccg cgcagcaaca gatggaaggc ctcctggcgc
     4381 cgcaccggcc caaggagccc gcgtggttcc tggccaccgt cggcgtctcg cccgaccacc
     4441 agggcaaggg tctgggcagc gccgtcgtgc tccccggagt ggaggcggcc gagcgcgccg
     4501 gggtgcccgc cttcctggag acctccgcgc cccgcaacct ccccttctac gagcggctcg
     4561 gcttcaccgt caccgccgac gtcgaggtgc ccgaaggacc gcgcacctgg tgcatgaccc
     4621 gcaagcccgg tgcctgacgc ccgccccacg acccgcagcg cccgaccgaa aggagcgcac
     4681 gaccccatgc atcgatgatc tagagctcgc tgatcagcct cgactgtgcc ttctagttgc
     4741 cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc
     4801 actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct
     4861 attctggggg gtggggtggg gcaggacagc aagggggagg attgggaaga caatagcagg
     4921 catgctgggg atgcggtggg ctctatggct tctgaggcgg aaagaacctg cagcccaagc
     4981 ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca
     5041 cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa
     5101 ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag
     5161 cggatccgca tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc
     5221 ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt
     5281 atgcagaggc cgaggccgcc tcggcctctg agctattcca gaagtagtga ggaggctttt
     5341 ttggaggcct aggcttttgc aaaaagctaa cttgtttatt gcagcttata atggttacaa
     5401 ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg
     5461 tggtttgtcc aaactcatca atgtatctta tcatgtctgg atccgctgca ttaatgaatc
     5521 ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact
     5581 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta
     5641 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag
     5701 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
     5761 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
     5821 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg
     5881 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc
     5941 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac
     6001 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
     6061 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
     6121 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
     6181 agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
     6241 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag
     6301 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct
     6361 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg
     6421 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat
     6481 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc
     6541 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg
     6601 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct
     6661 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca
     6721 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg
     6781 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg
     6841 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc
     6901 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag
     6961 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg
     7021 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag
     7081 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat
     7141 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg
     7201 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca
     7261 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca
     7321 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaat
//
