LOCUS       Exported                7445 bp ds-DNA    circular SYN 21-MAR-2018
DEFINITION  .
ACCESSION   .
VERSION     .
KEYWORDS    pcDNA5FRTTO-Puro-eGFP-HEC1-9A
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7445)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported 5 Jul 2018 from SnapGene 2.3.2
            http://www.snapgene.com
COMMENT     This file is created by Vector NTI
            http://www.invitrogen.com/
FEATURES             Location/Qualifiers
     source          1..7445
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        235..590
                     /note="CMV enhancer"
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        591..958
                     /label="CMV promoter (with TetO's)"
                     /note="CMV promoter"
     primer_bind     769..789
                     /note="CMV forward primer"
     protein_bind    820..838
                     /gene="tetO"
                     /bound_moiety="tetracycline repressor TetR"
                     /note="tet operator"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    841..859
                     /gene="tetO"
                     /bound_moiety="tetracycline repressor TetR"
                     /note="tet operator"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     CDS             1080..1796
                     /codon_start=1
                     /product="enhanced GFP"
                     /note="EGFP"
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             1812..3737
                     /codon_start=1
                     /label="Hec1 9A"
                     /note="Hec1 9A"
                     /translation="MKRAAVSAGGAGRLAMQELRSQDVNKQGLYTPQTKEKPTFGKLAI
                     NKPASERKVALFGKRTAGHGSRNAQLGIFSSSEKIKDPRPLNDKAFIQQCIRQLCEFLT
                     GNGYAHNVSMKSLQAPSVKDFLKIFTFLYGFLCPSYELPDTEFEEEVPRIFKDLGYPFA
                     LSKSSMYTVGAPHTWPHIVAALVWLIDCIKIHTAMKESSPLFDDGQPWGEETEDGIMHN
                     KLFLDYTIKCYESFMSGADSFDEMNAELQSKLKDLFNVDAFKLESLEAKNRALNEQIAR
                     LEQEREKEPNRLESLRKLKASLQGDVQKYQAYMSNLESHSAILDQKLNGLNEEIARVEL
                     ECETIKQENTRLQNIIDNQKYSVADIERINHERNELQQTINKLTKDLEAEQQKLWNEEL
                     KYARGKEAIETQLAEYHKLARKLKLIPKGAENSKGYDFEIKFNPEAGADCLVKYRAQVY
                     VPLKELLNETEEEINKALNKKMGLEDTLEQLNAMITESKRSVRTLKEEVQKLDDLYQQK
                     IKEAEEEDEKCASELESLEKHKHLLESTVNQGLSEAMNELDAVQREYQLVVQTTTEERR
                     KVGNNLQRLLEMVATHVGSVEKHLEEQIAKVDREYEECMSEDLSENIKEIRDKYEKKAT
                     LIKSSEE"
     polyA_signal    3789..4013
                     /note="bGH poly(A) signal"
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    4296..4343
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FRT"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             4387..4983
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /note="PuroR"
                     /note="confers resistance to puromycin"
                     /translation="TEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIERV
                     TELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLAA
                     QQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETSA
                     PRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     polyA_signal    5119..5239
                     /note="SV40 poly(A) signal"
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5288..5304)
                     /note="M13 rev"
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    5312..5328
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="lac operator"
                     /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(5336..5366)
                     /note="lac promoter"
                     /note="promoter for the E. coli lac operon"
     protein_bind    5381..5402
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding site"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(5690..6278)
                     /direction=LEFT
                     /note="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(6449..7309)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /note="AmpR"
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(7310..7414)
                     /gene="bla"
                     /note="AmpR promoter"
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc
      841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga
      901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga
      961 ctctagcgtt taaacttaag cttggtaccg agctcggatc cactagtcca gtgtggtgga
     1021 attctgcaga tatccagcac agtggcggcc gctcgagtct agcgctaccg gtcgccacca
     1081 tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc catcctggtc gagctggacg
     1141 gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg
     1201 gcaagctgac cctgaagttc atctgcacca ccggcaagct gcccgtgccc tggcccaccc
     1261 tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg ctaccccgac cacatgaagc
     1321 agcacgactt cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc accatcttct
     1381 tcaaggacga cggcaactac aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg
     1441 tgaaccgcat cgagctgaag ggcatcgact tcaaggagga cggcaacatc ctggggcaca
     1501 agctggagta caactacaac agccacaacg tctatatcat ggccgacaag cagaagaacg
     1561 gcatcaaggt gaacttcaag atccgccaca acatcgagga cggcagcgtg cagctcgccg
     1621 accactacca gcagaacacc cccatcggcg acggccccgt gctgctgccc gacaaccact
     1681 acctgagcac ccagtccgcc ctgagcaaag accccaacga gaagcgcgat cacatggtcc
     1741 tgctggagtt cgtgaccgcc gccgggatca ctctcggcat ggacgagctg tacaagtccg
     1801 gactcagatc catgaagcgc gctgcagttt ccgccggtgg tgctggccgc ctcgccatgc
     1861 aggagttaag atcccaggat gtaaataaac aaggcctcta tacccctcaa accaaagaga
     1921 aaccaacctt tggaaagttg gctataaaca aaccggcatc tgaaagaaaa gtcgcgctat
     1981 ttggcaaaag aactgctgga catggatccc ggaatgctca acttggtata ttttccagtt
     2041 ctgagaaaat caaggacccg agaccactta atgacaaagc attcattcag cagtgtattc
     2101 gacaactctg tgagtttctt acaggaaatg gttatgcaca taatgtgtcc atgaaatctc
     2161 tacaagctcc ctctgttaaa gacttcctga agatcttcac atttctttat ggcttcctgt
     2221 gcccctcata cgaacttcct gacacagagt ttgaagaaga ggttccaaga atctttaaag
     2281 accttgggta tccttttgca ctatccaaaa gctccatgta cacagtgggg gctcctcata
     2341 catggcctca cattgtggca gccttagttt ggctaataga ctgcatcaag atacatactg
     2401 ccatgaaaga aagctcacct ttatttgatg atgggcagcc ttggggagaa gaaactgaag
     2461 atggaattat gcataataag ttgtttttgg actacaccat aaaatgctat gagagtttta
     2521 tgagtggtgc cgacagcttt gatgagatga atgcagagct gcagtcaaaa ctgaaggatt
     2581 tatttaatgt ggatgctttt aagctggaat cattagaagc aaaaaacaga gcattgaatg
     2641 aacagattgc aagattggaa caagaaagag aaaaagagcc gaatcgtcta gagtcgttga
     2701 gaaaactgaa ggcttcctta caaggagatg ttcaaaagta tcaggcatac atgagcaatt
     2761 tggagtctca ttcagccatt cttgaccaga aattaaatgg tctcaatgag gaaattgcta
     2821 gagtagaact agaatgtgaa acaataaaac aggagaacac tcgactacag aatatcattg
     2881 acaaccagaa gtactcagtt gcagacattg agcgaataaa tcatgaaaga aatgaattgc
     2941 agcagactat taataaatta accaaggacc tggaagctga acaacagaag ttgtggaatg
     3001 aggagttaaa atatgccaga ggcaaagaag cgattgaaac acaattagca gagtatcaca
     3061 aattggctag aaaattaaaa cttattccta aaggtgctga gaattccaaa ggttatgact
     3121 ttgaaattaa gtttaatccc gaggctggtg ccgactgcct tgtcaaatac agggctcaag
     3181 tttatgtacc tcttaaggaa ctcctgaatg aaactgaaga agaaattaat aaagccctaa
     3241 ataaaaaaat gggtttggag gatactttag aacaattgaa tgcaatgata acagaaagca
     3301 agagaagtgt gagaactctg aaagaagaag ttcagaaact ggatgatctt taccaacaaa
     3361 aaattaagga agcagaggaa gaggatgaaa aatgtgccag tgagcttgag tccttggaga
     3421 aacacaagca cctgctagaa agtactgtta accaggggct cagtgaagct atgaatgaat
     3481 tagatgctgt tcagcgggaa taccaactag ttgtgcaaac cacgactgaa gaaagacgaa
     3541 aagtgggaaa taacttgcaa cgtctgttag agatggttgc tacacatgtt gggtctgtag
     3601 agaaacatct tgaggagcag attgctaaag ttgatagaga atatgaagaa tgcatgtcag
     3661 aagatctctc ggaaaatatt aaagagatta gagataagta tgagaagaaa gctactctaa
     3721 ttaagtcttc tgaagaatga ctcgaggacg gtaccgcggg cccgtttaaa cccgctgatc
     3781 agcctcgact gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc
     3841 cttgaccctg gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc
     3901 gcattgtctg agtaggtgtc attctattct ggggggtggg gtggggcagg acagcaaggg
     3961 ggaggattgg gaagacaata gcaggcatgc tggggatgcg gtgggctcta tggcttctga
     4021 ggcggaaaga accagctggg gctctagggg gtatccccac gcgccctgta gcggcgcatt
     4081 aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc
     4141 gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca
     4201 agctctaaat cgggggtccc tttagggttc cgatttagtg ctttacggca cctcgacccc
     4261 aaaaaacttg attagggtga tggttcacgt acctagaagt tcctattccg aagttcctat
     4321 tctctagaaa gtataggaac ttccttggcc aaaaagcctg aactcaccgc gacgtgctgg
     4381 cgcgccaccg agtacaagcc cacggtgcgc ctcgccaccc gcgacgacgt cccccgggcc
     4441 gtacgcaccc tcgccgccgc gttcgccgac taccccgcca cgcgccacac cgtcgacccg
     4501 gaccgccaca tcgagcgggt caccgagctg caagaactct tcctcacgcg cgtcgggctc
     4561 gacatcggca aggtgtgggt cgcggacgac ggcgccgcgg tggcggtctg gaccacgccg
     4621 gagagcgtcg aagcgggggc ggtgttcgcc gagatcggcc cgcgcatggc cgagttgagc
     4681 ggttcccggc tggccgcgca gcaacagatg gaaggcctcc tggcgccgca ccggcccaag
     4741 gagcccgcgt ggttcctggc caccgtcggc gtctcgcccg accaccaggg caagggtctg
     4801 ggcagcgccg tcgtgctccc cggagtggag gcggccgagc gcgccggggt gcccgccttc
     4861 ctggagacct ccgcgccccg caacctcccc ttctacgagc ggctcggctt caccgtcacc
     4921 gccgacgtcg aggtgcccga aggaccgcgc acctggtgca tgacccgcaa gcccggtgcc
     4981 tgacctaggc acgtactacg agatttcgat tccaccgccg ccttctatga aaggttgggc
     5041 ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg
     5101 gagttcttcg cccaccccaa cttgtttatt gcagcttaaa tggttacaaa taaagcaata
     5161 gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca
     5221 aactcatcaa tgtatcttat catgtctgta taccgtcgac ctctagctag agcttggcgt
     5281 aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca
     5341 tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat
     5401 taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt
     5461 aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct
     5521 cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa
     5581 aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa
     5641 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
     5701 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
     5761 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
     5821 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
     5881 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
     5941 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
     6001 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
     6061 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     6121 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     6181 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
     6241 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
     6301 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
     6361 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
     6421 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
     6481 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
     6541 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct
     6601 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
     6661 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
     6721 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt
     6781 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
     6841 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
     6901 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
     6961 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
     7021 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg
     7081 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
     7141 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
     7201 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
     7261 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt
     7321 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
     7381 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
     7441 acgtc
//
