LOCUS       sequence-253013-        6132 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses 3xFlag GAR1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6132)
  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 6132)
  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-GAR1
FEATURES             Location/Qualifiers
     source          1..6132
                     /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(1495..1512)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1501..1725
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1771..2199
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1858..1877)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2068..2089
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2208..2228)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2213..2542
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2393..2528
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2609..3403
                     /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(2663..2682)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3273..3292
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3577..3698
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3614..3633)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3668..3687
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3747..3763)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3747..3763)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3760..3782)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3771..3787
                     /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(3795..3801,3802..3819,3820..3825))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3840..3861
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3978..3995)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4149..4737)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4229..4248)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4908..5699,5700..5768))
                     /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     5531..5550
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5769..5873)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(5948..5967)
                     /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 tcttttcgag gcggaggtcg tggaggcttt
      781 aatcgaggtg gtggaggtgg cggcttcaac cgaggtggca gcagcaacca cttccgaggt
      841 ggaggcggcg gtggaggcgg cggcaatttc agaggcggcg gcaggggagg atttggacga
      901 gggggtggcc gcggaggctt taacaaaggc caagaccaag gacctccaga acgtgtagtc
      961 ttattaggag agttcctgca tccctgtgaa gatgacatag tttgtaaatg taccacagat
     1021 gaaaataagg tgccttattt caatgctcct gtttacttag aaaacaaaga acaaattgga
     1081 aaagtggatg aaatatttgg acaactcaga gatttttatt tttcagttaa gttgtcagaa
     1141 aacatgaagg cttcatcctt taaaaaacta cagaagtttt atatagaccc atataagctg
     1201 ctgccactgc agaggttttt acctcgacct ccaggtgaga aaggacctcc aagaggtggt
     1261 ggcaggggag gccgaggagg aggaagagga ggaggtggca gaggtggtgg cagaggcggt
     1321 ggttttagag gtggaagagg aggtggaggt gggggcttca gaggaggaag aggtggtggt
     1381 ttcagaggga gaggacatta aggatccact agtccagtgt ggtggaattc tgcagatatc
     1441 cagcacagtg gcggccgctc gagtctagag ggcccgttta aacccgctga tcagcctcga
     1501 ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc
     1561 tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc
     1621 tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt
     1681 gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct gaggcggaaa
     1741 gaaccagctg gggctctagg gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg
     1801 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc
     1861 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa
     1921 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac
     1981 ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt
     2041 tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca
     2101 accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt
     2161 taaaaaatga gctgatttaa caaaaattta acgcgaatta attctgtgga atgtgtgtca
     2221 gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     2281 caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     2341 aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc
     2401 cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt tttttattta
     2461 tgcagaggcc gaggccgcct ctgcctctga gctattccag aagtagtgag gaggcttttt
     2521 tggaggccta ggcttttgca aaaagctccc gggagcttgt atatccattt tcggatctga
     2581 tcaagagaca ggatgaggat cgtttcgcat gattgaacaa gatggattgc acgcaggttc
     2641 tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga caatcggctg
     2701 ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac
     2761 cgacctgtcc ggtgccctga atgaactgca ggacgaggca gcgcggctat cgtggctggc
     2821 cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg gaagggactg
     2881 gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg ctcctgccga
     2941 gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc cggctacctg
     3001 cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga tggaagccgg
     3061 tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt
     3121 cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc
     3181 ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg actgtggccg
     3241 gctgggtgtg gcggaccgct atcaggacat agcgttggct acccgtgata ttgctgaaga
     3301 gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg ctcccgattc
     3361 gcagcgcatc gccttctatc gccttcttga cgagttcttc tgagcgggac tctggggttc
     3421 gaaatgaccg accaagcgac gcccaacctg ccatcacgag atttcgattc caccgccgcc
     3481 ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat gatcctccag
     3541 cgcggggatc tcatgctgga gttcttcgcc caccccaact tgtttattgc agcttataat
     3601 ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat
     3661 tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctgtat accgtcgacc
     3721 tctagctaga gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg
     3781 ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa
     3841 tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac
     3901 ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt
     3961 gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga
     4021 gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca
     4081 ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg
     4141 ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt
     4201 cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc
     4261 ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct
     4321 tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc
     4381 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta
     4441 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca
     4501 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag
     4561 tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag
     4621 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt
     4681 agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa
     4741 gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg
     4801 attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga
     4861 agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta
     4921 atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc
     4981 cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg
     5041 ataccgcgag acccacgctc accggctcca gatttatcag caataaacca gccagccgga
     5101 agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt
     5161 tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt
     5221 gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc
     5281 caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc
     5341 ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca
     5401 gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag
     5461 tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg
     5521 tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa
     5581 cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa
     5641 cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga
     5701 gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga
     5761 atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg
     5821 agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt
     5881 ccccgaaaag tgccacctga cgtcgacgga tcgggagatc tcccgatccc ctatggtgca
     5941 ctctcagtac aatctgctct gatgccgcat agttaagcca gtatctgctc cctgcttgtg
     6001 tgttggaggt cgctgagtag tgcgcgagca aaatttaagc tacaacaagg caaggcttga
     6061 ccgacaattg catgaagaat ctgcttaggg ttaggcgttt tgcgctgctt cgcgatgtac
     6121 gggccagata ta
//
