LOCUS       sequence-252988-        5940 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses 3xFlag NHP2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5940)
  AUTHORS   Egan ED, Collins K
  TITLE     Specificity and stoichiometry of subunit interactions in the human 
            telomerase holoenzyme assembled in vivo.
  JOURNAL   Mol Cell Biol. 2010 Jun;30(11):2775-86. doi: 10.1128/MCB.00151-10. 
            Epub 2010 Mar 29.
  PUBMED    20351177
REFERENCE   2  (bases 1 to 5940)
  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"; doi: 
            "10.1128/MCB.00151-10"; journalName: "Mol Cell Biol"; date: 
            "2010-06"; volume: "30"; issue: "11"; pages: "2775-86"
COMMENT     Sequence Label: pcDNA3.1 3xF-NHP2
FEATURES             Location/Qualifiers
     source          1..5940
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        7..386
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        387..590
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     541..561
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     635..654
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        635..653
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             676..741
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     complement(1303..1320)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1309..1533
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1579..2007
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1666..1685)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1876..1897
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2016..2036)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2021..2350
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2201..2336
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2263..2282
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2417..3211
                     /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(2471..2490)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3081..3100
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3385..3506
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3422..3441)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3476..3495
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3555..3571)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3555..3571)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3568..3590)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3579..3595
                     /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(3603..3609,3610..3627,3628..3633))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3648..3669
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3786..3803)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3957..4545)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4037..4056)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4716..5507,5508..5576))
                     /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     5339..5358
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5577..5681)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(5756..5775)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 cgcgttgaca ttgattattg actagttatt aatagtaatc aattacgggg tcattagttc
       61 atagcccata tatggagttc cgcgttacat aacttacggt aaatggcccg cctggctgac
      121 cgcccaacga cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa
      181 tagggacttt ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag
      241 tacatcaagt gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc
      301 ccgcctggca ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct
      361 acgtattagt catcgctatt accatggtga tgcggttttg gcagtacatc aatgggcgtg
      421 gatagcggtt tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt
      481 tgttttggca ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga
      541 cgcaaatggg cggtaggcgt gtacggtggg aggtctatat aagcagagct ctctggctaa
      601 ctagagaacc cactgcttac tggcttatcg aaattaatac gactcactat agggagaccc
      661 aagctggcta gcatggacta caaagaccat gacggtgatt ataaagatca tgacatcgat
      721 tacaaggatg acgatgacaa gaagcttatg accaaaataa aggcagatcc cgacgggccc
      781 gaggctcagg cggaggcgtg ttccggggag cgcacctacc aggagctgct ggtcaaccag
      841 aaccccatcg cgcagcccct ggcttctcgc cgcctcacgc ggaagctcta caaatgcatc
      901 aagaaagcgg tgaagcagaa gcagattcgg cgcggggtga aagaggttca gaaatttgtc
      961 aacaaaggag aaaaagggat catggttttg gcaggagaca cactgcccat tgaggtatac
     1021 tgccatctcc cagtcatgtg tgaggaccga aatttgccct atgtctatat cccctctaag
     1081 acggacctgg gtgcagccgc aggctccaag cgccccacct gtgtgataat ggtcaagccc
     1141 catgaggagt accaggaggc ttacgatgag tgcctggagg aggtgcagtc cctgccccta
     1201 cccctatgag gatccactag tccagtgtgg tggaattctg cagatatcca gcacagtggc
     1261 ggccgctcga gtctagaggg cccgtttaaa cccgctgatc agcctcgact gtgccttcta
     1321 gttgccagcc atctgttgtt tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca
     1381 ctcccactgt cctttcctaa taaaatgagg aaattgcatc gcattgtctg agtaggtgtc
     1441 attctattct ggggggtggg gtggggcagg acagcaaggg ggaggattgg gaagacaata
     1501 gcaggcatgc tggggatgcg gtgggctcta tggcttctga ggcggaaaga accagctggg
     1561 gctctagggg gtatccccac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg
     1621 ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct
     1681 tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc
     1741 ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg
     1801 atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt
     1861 ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg
     1921 tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc
     1981 tgatttaaca aaaatttaac gcgaattaat tctgtggaat gtgtgtcagt tagggtgtgg
     2041 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc
     2101 aaccaggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     2161 caattagtca gcaaccatag tcccgcccct aactccgccc atcccgcccc taactccgcc
     2221 cagttccgcc cattctccgc cccatggctg actaattttt tttatttatg cagaggccga
     2281 ggccgcctct gcctctgagc tattccagaa gtagtgagga ggcttttttg gaggcctagg
     2341 cttttgcaaa aagctcccgg gagcttgtat atccattttc ggatctgatc aagagacagg
     2401 atgaggatcg tttcgcatga ttgaacaaga tggattgcac gcaggttctc cggccgcttg
     2461 ggtggagagg ctattcggct atgactgggc acaacagaca atcggctgct ctgatgccgc
     2521 cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg acctgtccgg
     2581 tgccctgaat gaactgcagg acgaggcagc gcggctatcg tggctggcca cgacgggcgt
     2641 tccttgcgca gctgtgctcg acgttgtcac tgaagcggga agggactggc tgctattggg
     2701 cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct cctgccgaga aagtatccat
     2761 catggctgat gcaatgcggc ggctgcatac gcttgatccg gctacctgcc cattcgacca
     2821 ccaagcgaaa catcgcatcg agcgagcacg tactcggatg gaagccggtc ttgtcgatca
     2881 ggatgatctg gacgaagagc atcaggggct cgcgccagcc gaactgttcg ccaggctcaa
     2941 ggcgcgcatg cccgacggcg aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa
     3001 tatcatggtg gaaaatggcc gcttttctgg attcatcgac tgtggccggc tgggtgtggc
     3061 ggaccgctat caggacatag cgttggctac ccgtgatatt gctgaagagc ttggcggcga
     3121 atgggctgac cgcttcctcg tgctttacgg tatcgccgct cccgattcgc agcgcatcgc
     3181 cttctatcgc cttcttgacg agttcttctg agcgggactc tggggttcga aatgaccgac
     3241 caagcgacgc ccaacctgcc atcacgagat ttcgattcca ccgccgcctt ctatgaaagg
     3301 ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc
     3361 atgctggagt tcttcgccca ccccaacttg tttattgcag cttataatgg ttacaaataa
     3421 agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt
     3481 ttgtccaaac tcatcaatgt atcttatcat gtctgtatac cgtcgacctc tagctagagc
     3541 ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca
     3601 cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa
     3661 ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag
     3721 ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc
     3781 gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct
     3841 cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg
     3901 tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc
     3961 cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga
     4021 aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct
     4081 cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg
     4141 gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag
     4201 ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat
     4261 cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac
     4321 aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac
     4381 tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc
     4441 ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt
     4501 tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc
     4561 ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg
     4621 agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca
     4681 atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca
     4741 cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag
     4801 ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac
     4861 ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc
     4921 agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct
     4981 agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc
     5041 gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg
     5101 cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc
     5161 gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat
     5221 tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag
     5281 tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat
     5341 aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg
     5401 cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca
     5461 cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga
     5521 aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc
     5581 ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag cggatacata
     5641 tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg
     5701 ccacctgacg tcgacggatc gggagatctc ccgatcccct atggtgcact ctcagtacaa
     5761 tctgctctga tgccgcatag ttaagccagt atctgctccc tgcttgtgtg ttggaggtcg
     5821 ctgagtagtg cgcgagcaaa atttaagcta caacaaggca aggcttgacc gacaattgca
     5881 tgaagaatct gcttagggtt aggcgttttg cgctgcttcg cgatgtacgg gccagatata
//
