LOCUS       sequence-104608-        7359 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses Mouse Nanog R147A 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 7359)
  TITLE     Plasmids of NANOG HD Mutants
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7359)
  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            "Unpublished"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(R147A)-IP
FEATURES             Location/Qualifiers
     source          1..7359
                     /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..4006,4007..4009,4010..4018)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3599..3616)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3826..3845
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             4019..4618
                     /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(4019..4038)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4515..4535
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     complement(4699..4716)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4705..4929
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4976..4992)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4976..4992)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4989..5011)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5000..5016
                     /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(5024..5030,5031..5048,5049..5054))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5069..5090
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        5149..5344
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      5195..5330
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    5350..5484
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5552..5569)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5723..6311)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5803..5822)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6482..7273,7274..7342))
                     /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     7105..7124
                     /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 caggttaaga cctggtttca aaaccaagcg 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 cctaacgtta ctggccgaag ccgcttggaa
     3481 taaggccggt gtgcgtttgt ctatatgtta ttttccacca tattgccgtc ttttggcaat
     3541 gtgagggccc ggaaacctgg ccctgtcttc ttgacgagca ttcctagggg tctttcccct
     3601 ctcgccaaag gaatgcaagg tctgttgaat gtcgtgaagg aagcagttcc tctggaagct
     3661 tcttgaagac aaacaacgtc tgtagcgacc ctttgcaggc agcggaaccc cccacctggc
     3721 gacaggtgcc tctgcggcca aaagccacgt gtataagata cacctgcaaa ggcggcacaa
     3781 ccccagtgcc acgttgtgag ttggatagtt gtggaaagag tcaaatggct ctcctcaagc
     3841 gtattcaaca aggggctgaa ggatgcccag aaggtacccc attgtatggg atctgatctg
     3901 gggcctcggt gcacatgctt tacatgtgtt tagtcgaggt taaaaaaacg tctaggcccc
     3961 ccgaaccacg gggacgtggt tttcctttga aaaacacgat gataatatgg ccacaaccat
     4021 gaccgagtac aagcccacgg tgcgcctcgc cacccgcgac gacgtcccca gggccgtacg
     4081 caccctcgcc gccgcgttcg ccgactaccc cgccacgcgc cacaccgtcg atccggaccg
     4141 ccacatcgag cgggtcaccg agctgcaaga actcttcctc acgcgcgtcg ggctcgacat
     4201 cggcaaggtg tgggtcgcgg acgacggcgc cgcggtggcg gtctggacca cgccggagag
     4261 cgtcgaagcg ggggcggtgt tcgccgagat cggcccgcgc atggccgagt tgagcggttc
     4321 ccggctggcc gcgcagcaac agatggaagg cctcctggcg ccgcaccggc ccaaggagcc
     4381 cgcgtggttc ctggccaccg tcggcgtctc gcccgaccac cagggcaagg gtctgggcag
     4441 cgccgtcgtg ctccccggag tggaggcggc cgagcgcgcc ggggtgcccg ccttcctgga
     4501 gacctccgcg ccccgcaacc tccccttcta cgagcggctc ggcttcaccg tcaccgccga
     4561 cgtcgaggtg cccgaaggac cgcgcacctg gtgcatgacc cgcaagcccg gtgcctgacg
     4621 cccgccccac gacccgcagc gcccgaccga aaggagcgca cgaccccatg catcgatgat
     4681 ctagagctcg ctgatcagcc tcgactgtgc cttctagttg ccagccatct gttgtttgcc
     4741 cctcccccgt gccttccttg accctggaag gtgccactcc cactgtcctt tcctaataaa
     4801 atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc tattctgggg ggtggggtgg
     4861 ggcaggacag caagggggag gattgggaag acaatagcag gcatgctggg gatgcggtgg
     4921 gctctatggc ttctgaggcg gaaagaacct gcagcccaag cttggcgtaa tcatggtcat
     4981 agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa
     5041 gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc
     5101 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gcggatccgc atctcaatta
     5161 gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc
     5221 cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc
     5281 ctcggcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg
     5341 caaaaagcta acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca
     5401 aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc
     5461 aatgtatctt atcatgtctg gatccgctgc attaatgaat cggccaacgc gcggggagag
     5521 gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg
     5581 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat
     5641 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     5701 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     5761 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     5821 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     5881 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     5941 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     6001 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     6061 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     6121 cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct
     6181 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     6241 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     6301 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
     6361 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
     6421 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca
     6481 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca
     6541 tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc
     6601 ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa
     6661 accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc
     6721 agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca
     6781 acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat
     6841 tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag
     6901 cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac
     6961 tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt
     7021 ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt
     7081 gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc
     7141 tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat
     7201 ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca
     7261 gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga
     7321 cacggaaatg ttgaatactc atactcttcc tttttcaat
//
