LOCUS       V035365                 8526 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V035365
VERSION     V035365
KEYWORDS    .
SOURCE      .
  ORGANISM  .
            .
FEATURES             Location/Qualifiers
     promoter        1..1179
                     /note="strong constitutive promoter for human elongation
                     factor EF-1α"
                     /label="EF-1α promoter"
     promoter        1196..1214
                     /note="promoter for bacteriophage T7 RNA polymerase"
                     /label="T7 promoter"
     protein_bind    1293..1317
                     /bound_moiety="BP Clonase™"
                     /gene="mutant version of&nbsp;attB"
                     /note="recombination site for the Gateway® BP reaction"
                     /label="attB1"
     regulatory      1342..1351
                     /note="vertebrate consensus sequence for strong initiation
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
                     /label="Kozak sequence"
     CDS             2773..2793
                     /codon_start=1
                     /product="N-myristoylation signal from Src kinase (Pellman
                     et al., 1985;&nbsp;Kaplan et al., 1988)"
                     /transl_table=1
                     /translation="MGSSKSK"
                     /label="myr"
     CDS             2806..4245
                     /label="Akt1(NM_001331107)"
                     /note="Akt1(NM_001331107)"
                     /gene="Akt1"
     CDS             4246..4272
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /transl_table=1
                     /translation="YPYDVPDYA"
                     /label="HA"
     protein_bind    complement(4292..4316)
                     /bound_moiety="BP Clonase™"
                     /gene="mutant version of&nbsp;attB"
                     /note="recombination site for the Gateway® BP reaction"
                     /label="attB2"
     CDS             4369..4410
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /transl_table=1
                     /translation="GKPIPNPLLGLDST"
                     /label="V5 tag"
     polyA_signal    4454..4678
                     /note="bovine growth hormone polyadenylation signal"
                     /label="bGH poly(A) signal"
     protein_bind    4748..4781
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core
                     sequence (ATGTATGC) (Shaw et al., 2021)."
                     /label="loxP"
     primer_bind     complement(5239..5255)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 fwd"
     promoter        5729..5833
                     /gene="bla"
                     /label="AmpR promoter"
     CDS             5834..6694
                     /codon_start=1
                     /gene="bla"
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /product="β-lactamase"
                     /transl_table=1
                     /translation="MSIQHFRVALIPFFAAFCLPVFA,HPETLVKVKDAEDQLGARVGY
                     IELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEY
                     SPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDR
                     WEPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRS
                     ALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGA
                     SLIKHW*"
                     /label="AmpR"
     rep_origin      6865..7453
                     /direction=RIGHT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     protein_bind    7741..7762
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     promoter        7777..7807
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    7815..7831
                     /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-β-D-thiogalactopyranoside (IPTG)."
                     /label="lac operator"
     primer_bind     7839..7855
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    8353..8386
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core
                     sequence (ATGTATGC) (Shaw et al., 2021)."
                     /label="loxP"
ORIGIN
        1 ggctccggtg cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg
       61 ggaggggtcg gcaattgaac cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt
      121 gatgtcgtgt actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca
      181 gtagtcgccg tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggtaagtgcc
      241 gtgtgtggtt cccgcgggcc tggcctcttt acgggttatg gcccttgcgt gccttgaatt
      301 acttccacct ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg
      361 gagagttcga ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg
      421 cctgggcgct ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct
      481 ttcgataagt ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg
      541 caagatagtc ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc
      601 gcgggcggcg acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga
      661 gcgcggccac cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct
      721 ggcctcgcgc cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca
      781 gttgcgtgag cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg
      841 acgcggcgct cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg
      901 tcctcagccg tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat
      961 tagttctcga gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg
     1021 gagtttcccc acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa
     1081 ttctccttgg aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca
     1141 gtggttcaaa gtttttttct tccatttcag gtgtcgtgag gaattagctt ggtactaata
     1201 cgactcacta tagggagacc caagctggct aggtaagctt ggtaccgagc tcggatccac
     1261 tagtccagtg tggtggaatt ctgcagatat caacaagttt gtacaaaaaa gcaggctccg
     1321 cggccgcccc cttcaccatg ggccaccatg gggctgtacg cggccgccgc ggcggtgctg
     1381 gccggcgtgg aaagccgcca gggctccctc aaggggttgg tgtattccag caacttccag
     1441 aacctgaagc agttgtacgc tctggtgtgc gagacgcagc gctactcggc cgtgctggac
     1501 gctgtcatcg ccagtgctgg gctccttcgc gccgagaaga agctgagacc gcaccttgcc
     1561 aaggtgctag tgtatgaatt attgctgggg aaggggtttc gagggggtgg aggccgatgg
     1621 aaggctctgc tgggccggca ccaggcaagg ctcaaggctg agctggcccg actcaaggtt
     1681 catcggggtg tgagccgaaa tgaggacctg ttgcaagaga gctccagacc tggccaagcc
     1741 taccaggtac cacggttcgt gcgtgtgaac acgctcaaga cccgccctga ggatgcaatc
     1801 gattatttca agagacaagg tttctcctac cagggtcggg cttccagcct ggaagacttg
     1861 cgagctctca aggggcagca ttttcttttg gatcccttgc tgcctgagct tcttgtattt
     1921 cctgcacaga cagatctgca cgagcatcca ctctaccgtg ctggccatct catcctgcag
     1981 gacaaggcca gctgcctccc agccatgctg ctgtcccccc cgccaggctc ccatgtcatt
     2041 gatgcctgtg ctgcccctgg taacaagacc agctacattg cagcccttct aaagaaccag
     2101 gggaagatct ttgctttcga ccaggacgct aagcggctgg cagccatggc gacgttggtg
     2161 gctcgggctg gggtctcttg ctgcgagtta gcagagaagg acttcctgac agtctctccc
     2221 tctgaccagc gttacagcca ggtccagtac atcttgctgg atccttcttg tagtggctct
     2281 ggcatgctga gccggcagct ggaggaacat ggggaaggca cacctagcaa ggagcgcttg
     2341 caggccttgg ccgggttcca gcagcgggct ctgtgccatg cacttaggtt cccctctctg
     2401 cagcgcctag tctactccac gtgttccctt tgccaagaag agaatgaaga cgtggtccag
     2461 gaggctctac agcacaactc ggggacattc aggttggctc ctgtcctacc cacatggcct
     2521 caccgaggtc tgagtacctt tccagggtct gaacactgcc tccgggcttc ccctgagacc
     2581 acgctcactg gtggcttctt cattgctgtg tttgaacgtg cagaggtggt accaacccca
     2641 gccccacaga ccgatgcaat ggatccagaa cctctcagcc aagtcccaaa gagaaagaga
     2701 agacgcaaag cagcagttgg tgccagcatg cagccaagca caggcggcgg agggagtgga
     2761 ggagggggaa gcatggggag cagcaagagc aagcccaagt ctagaatgaa cgacgtagcc
     2821 attgtgaagg agggctggct gcacaaacga ggggaatata ttaaaacctg gcggccacgc
     2881 tacttcctcc tcaagaacga tggcaccttt attggctaca aggaacggcc tcaggatgtg
     2941 gatcagcgag agtccccact caacaacttc tcagtggcac aatgccagct gatgaagaca
     3001 gagcggccaa ggcccaacac ctttatcatc cgctgcctgc agtggaccac agtcattgag
     3061 cgcaccttcc atgtggaaac gcctgaggag cgggaagaat gggccaccgc cattcagact
     3121 gtggccgatg gactcaagag gcaggaagaa gagacgatgg acttccgatc aggctcaccc
     3181 agtgacaact caggggctga agagatggag gtgtccctgg ccaagcccaa gcaccgtgtg
     3241 accatgaacg agtttgagta cctgaaacta ctgggcaagg gcacctttgg gaaagtgatt
     3301 ctggtgaaag agaaggccac aggccgctac tatgccatga agatcctcaa gaaggaggtc
     3361 atcgtcgcca aggatgaggt tgcccacacg cttactgaga accgtgtcct gcagaactct
     3421 aggcatccct tccttacggc cctcaagtac tcattccaga cccacgaccg cctctgcttt
     3481 gtcatggagt atgccaacgg gggcgagctc ttcttccacc tgtctcgaga gcgcgtgttc
     3541 tccgaggacc gggcccgctt ctatggtgcg gagattgtgt ctgccctgga ctacttgcac
     3601 tccgagaaga acgtggtgta ccgggacctg aagctggaga acctcatgct ggacaaggac
     3661 gggcacatca agataacgga cttcgggctg tgcaaggagg ggatcaagga tggtgccact
     3721 atgaagacat tctgcggaac gccggagtac ctggcccctg aggtgctgga ggacaacgac
     3781 tacggccgtg cagtggactg gtgggggctg ggcgtggtca tgtatgagat gatgtgtggc
     3841 cgcctgccct tctacaacca ggaccacgag aagctgttcg agctgatcct catggaggag
     3901 atccgcttcc cgcgcacact cggccctgag gccaagtccc tgctctccgg gctgctcaag
     3961 aaggacccta cacagaggct cggtgggggc tctgaggatg ccaaggagat catgcagcac
     4021 cggttctttg ccaacatcgt gtggcaggat gtgtatgaga agaagctgag cccacctttc
     4081 aagccccagg tcacctctga gactgacacc aggtatttcg atgaggagtt cacagctcag
     4141 atgatcacca tcacgccgcc tgatcaagat gacagcatgg agtgtgtgga cagtgagcgg
     4201 aggccgcact tcccccagtt ctcctactca gccagtggca cagcctaccc atacgacgtc
     4261 ccagactacg cttgaaaggg tgggcgcgcc gacccagctt tcttgtacaa agtggttgat
     4321 atccagcaca gtggcggccg ctcgagtcta gagggcccgc ggttcgaagg taagcctatc
     4381 cctaaccctc tcctcggtct cgattctacg cgtaccggtt agtaatgagt ttaaacccgc
     4441 tgatcagcct cgactgtgcc ttctagttgc cagccatctg ttgtttgccc ctcccccgtg
     4501 ccttccttga ccctggaagg tgccactccc actgtccttt cctaataaaa tgaggaaatt
     4561 gcatcgcatt gtctgagtag gtgtcattct attctggggg gtggggtggg gcaggacagc
     4621 aagggggagg attgggaaga caatagcagg catgctgggg atgcggtggg ctctatggct
     4681 tctgaggcgg aaagaaccag ctggggctct agggggtatc cccacgcgcc ctgtagcggc
     4741 gcattaaata acttcgtata atgtatgcta tacgaagtta tatcgtatgc atgctttgca
     4801 tacttctgcc tgctggggag cctggggact ttccacaccc taactgacac acattccaca
     4861 gaattccttg aaatacatcc acaggtacac ctccaattga ctcaaatgat gtcaattagt
     4921 ctatcagaag cttctaaagc catgacatca ttttctggaa ttttccaagc tgtttaaagg
     4981 cacagtcaac ttagtgtatg taaacttctg acccactgga attgtgatac agtgaattat
     5041 aagtgaaata atctgtctgt aaacaattgt tggaaaaatg acttgtgtca tgcacaaagt
     5101 agatgtccta actgacttgc caaaactatt gtttgttaac aagaaatttg tggagtagtt
     5161 gaaaaacgag ttttaatgac tccaacttaa gtgtatgtaa acttccgact tcaactgtag
     5221 ggtaccgagc tcgaattcac tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg
     5281 cgttacccaa cttaatcgcc ttgcagcaca tccccctttc gccagctggc gtaatagcga
     5341 agaggcccgc accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg aatggcgcct
     5401 gatgcggtat tttctcctta cgcatctgtg cggtatttca caccgcatat ggtgcactct
     5461 cagtacaatc tgctctgatg ccgcatagtt aagccagccc cgacacccgc caacacccgc
     5521 tgacgcgccc tgacgggctt gtctgctccc ggcatccgct tacagacaag ctgtgaccgt
     5581 ctccgggagc tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg cgagacgaaa
     5641 gggcctcgtg atacgcctat ttttataggt taatgtcatg ataataatgg tttcttagac
     5701 gtcaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat
     5761 acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg
     5821 aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc
     5881 attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga
     5941 tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga
     6001 gagttttcgc cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg
     6061 cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc
     6121 tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac
     6181 agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact
     6241 tctgacaacg atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca
     6301 tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg
     6361 tgacaccacg atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact
     6421 acttactcta gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg
     6481 accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg
     6541 tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat
     6601 cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc
     6661 tgagataggt gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat
     6721 actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt
     6781 tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc
     6841 cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt
     6901 gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac
     6961 tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt
     7021 gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct
     7081 gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga
     7141 ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac
     7201 acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg
     7261 agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt
     7321 cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc
     7381 tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg
     7441 gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc
     7501 ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc
     7561 ctttgagtga gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag
     7621 cgaggaagcg gaagagcgcc caatacgcaa accgcctctc cccgcgcgtt ggccgattca
     7681 ttaatgcagc tggcacgaca ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat
     7741 taatgtgagt tagctcactc attaggcacc ccaggcttta cactttatgc ttccggctcg
     7801 tatgttgtgt ggaattgtga gcggataaca atttcacaca ggaaacagct atgaccatga
     7861 ttacgccaag cttgcatgcc tgcaggtcga ctctagacct atacagttga agtcggaagt
     7921 ttacatacac ttaagttgga gtcattaaaa ctcgtttttc aactactcca caaatttctt
     7981 gttaacaaac aatagttttg gcaagtcagt taggacatct actttgtgca tgacacaagt
     8041 catttttcca acaattgttt acagacagat tatttcactt ataattcact gtatcacaat
     8101 tccagtgggt cagaagttta catacactaa gttgactgtg cctttaaaca gcttggaaaa
     8161 ttccagaaaa tgatgtcatg gctttagaag cttctgatag actaattgac atcatttgag
     8221 tcaattggag gtgtacctgt ggatgtattt caaggggatc cagacatgat aagatacatt
     8281 gatgagtttg gacaaaccac aactagaatg cagcggccgc tagctcgaga tcttaattaa
     8341 actagtatcg atataacttc gtataatgta tgctatacga agttatatgt acgggccaga
     8401 tatacgcgtt gacattgatt attgactagg cttttgcaaa aagctttgca aagatggata
     8461 aagttttaaa cagagaggaa tctttgcagc taatggacct tctaggtctt gaaaggagtg
     8521 ggaatt
//
