LOCUS       sequence-104602-        7360 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses Mouse Nanog T141A Mutants. It is modified from 
            pPyCAG-Nanog-IP (Addgene Plasmid #13838)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7360)
  TITLE     Plasmids of NANOG HD Mutants
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7360)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pPyCAG-Nanog(T141A)-IP
FEATURES             Location/Qualifiers
     source          1..7360
                     /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    2437..2461
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(3390..3414)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_feature    join(3432..4007,4008..4010,4011..4019)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3600..3617)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3827..3846
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             4020..4619
                     /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(4020..4039)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4516..4536
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     complement(4700..4717)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4706..4930
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4977..4993)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4977..4993)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4990..5012)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5001..5017
                     /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(5025..5031,5032..5049,5050..5055))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5070..5091
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        5150..5345
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      5196..5331
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    5351..5485
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5553..5570)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5724..6312)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5804..5823)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6483..7274,7275..7343))
                     /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     7106..7125
                     /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 cgccaggcgg ggcggggcgg ggcgaggggc
     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 ttcntacagc
     2401 tcctgggcaa cgtgctggtt attgtgctga attatcacaa gtttgtacaa aaaagcaggc
     2461 tctgacatga gtgtgggtct tcctggtccc cacagtttgc ctagttctga ggaagcatcg
     2521 aattctggga acgcctcatc aatgcctgca gtttttcatc ccgagaacta ttcttgctta
     2581 caagggtctg ctactgagat gctctgcaca gaggctgcct ctcctcgccc ttcctctgaa
     2641 gacctgcctc ttcaaggcag ccctgattct tctaccagtc ccaaacaaaa gctctcaagt
     2701 cctgaggctg acaagggccc tgaggaggag gagaacaagg tccttgccag gaagcagaag
     2761 atgcggactg tgttctctca ggcccagctg tgtgcactca aggacaggtt tcagaagcag
     2821 aagtacctca gcctccagca gatgcaagaa ctctcctcca ttctgaacct gagctataag
     2881 caggttaagg cctggtttca aaaccaaagg atgaagtgca agcggtggca gaaaaaccag
     2941 tggttgaaga ctagcaatgg tctgattcag aagggctcag caccagtgga gtatcccagc
     3001 atccattgca gctatcccca gggctatctg gtgaacgcat ctggaagcct ttccatgtgg
     3061 ggcagccaga cttggaccaa cccaacttgg agcagccaga cctggaccaa cccaacttgg
     3121 aacaaccaga cctggaccaa cccaacttgg agcagccagg cctggaccgc tcagtcctgg
     3181 aacggccagc cttggaatgc tgctccgctc cataacttcg gggaggactt tctgcagcct
     3241 tacgtacagt tgcagcaaaa cttctctgcc agtgatttgg aggtgaattt ggaagccact
     3301 agggaaagcc atgcgcattt tagcacccca caagccttgg aattattcct gaactactct
     3361 gtgactccac caggtgaaat atgagactta cccagctttc ttgtacaaag tggtgataat
     3421 tccgcccccc ccccctctcc ctcccccccc ccctaacgtt actggccgaa gccgcttgga
     3481 ataaggccgg tgtgcgtttg tctatatgtt attttccacc atattgccgt cttttggcaa
     3541 tgtgagggcc cggaaacctg gccctgtctt cttgacgagc attcctaggg gtctttcccc
     3601 tctcgccaaa ggaatgcaag gtctgttgaa tgtcgtgaag gaagcagttc ctctggaagc
     3661 ttcttgaaga caaacaacgt ctgtagcgac cctttgcagg cagcggaacc ccccacctgg
     3721 cgacaggtgc ctctgcggcc aaaagccacg tgtataagat acacctgcaa aggcggcaca
     3781 accccagtgc cacgttgtga gttggatagt tgtggaaaga gtcaaatggc tctcctcaag
     3841 cgtattcaac aaggggctga aggatgccca gaaggtaccc cattgtatgg gatctgatct
     3901 ggggcctcgg tgcacatgct ttacatgtgt ttagtcgagg ttaaaaaaac gtctaggccc
     3961 cccgaaccac ggggacgtgg ttttcctttg aaaaacacga tgataatatg gccacaacca
     4021 tgaccgagta caagcccacg gtgcgcctcg ccacccgcga cgacgtcccc agggccgtac
     4081 gcaccctcgc cgccgcgttc gccgactacc ccgccacgcg ccacaccgtc gatccggacc
     4141 gccacatcga gcgggtcacc gagctgcaag aactcttcct cacgcgcgtc gggctcgaca
     4201 tcggcaaggt gtgggtcgcg gacgacggcg ccgcggtggc ggtctggacc acgccggaga
     4261 gcgtcgaagc gggggcggtg ttcgccgaga tcggcccgcg catggccgag ttgagcggtt
     4321 cccggctggc cgcgcagcaa cagatggaag gcctcctggc gccgcaccgg cccaaggagc
     4381 ccgcgtggtt cctggccacc gtcggcgtct cgcccgacca ccagggcaag ggtctgggca
     4441 gcgccgtcgt gctccccgga gtggaggcgg ccgagcgcgc cggggtgccc gccttcctgg
     4501 agacctccgc gccccgcaac ctccccttct acgagcggct cggcttcacc gtcaccgccg
     4561 acgtcgaggt gcccgaagga ccgcgcacct ggtgcatgac ccgcaagccc ggtgcctgac
     4621 gcccgcccca cgacccgcag cgcccgaccg aaaggagcgc acgaccccat gcatcgatga
     4681 tctagagctc gctgatcagc ctcgactgtg ccttctagtt gccagccatc tgttgtttgc
     4741 ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct ttcctaataa
     4801 aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg gggtggggtg
     4861 gggcaggaca gcaaggggga ggattgggaa gacaatagca ggcatgctgg ggatgcggtg
     4921 ggctctatgg cttctgaggc ggaaagaacc tgcagcccaa gcttggcgta atcatggtca
     4981 tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga
     5041 agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg
     5101 cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agcggatccg catctcaatt
     5161 agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt
     5221 ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg
     5281 cctcggcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt
     5341 gcaaaaagct aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac
     5401 aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat
     5461 caatgtatct tatcatgtct ggatccgctg cattaatgaa tcggccaacg cgcggggaga
     5521 ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc
     5581 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
     5641 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     5701 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     5761 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     5821 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     5881 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     5941 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     6001 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     6061 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     6121 acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc
     6181 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     6241 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     6301 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     6361 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     6421 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
     6481 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     6541 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
     6601 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata
     6661 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     6721 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     6781 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     6841 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     6901 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     6961 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     7021 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     7081 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     7141 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     7201 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     7261 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     7321 acacggaaat gttgaatact catactcttc ctttttcaat
//
