LOCUS       Exported                6866 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  To overexpress AKT1 in Mammalian Cells.
ACCESSION   .
VERSION     .
KEYWORDS    pcDNA3.1-HA AKT1
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6866)
  AUTHORS   Lee MS, Jeong MH, Lee HW, Han HJ, Ko A, Hewitt SM, Kim JH, Chun KH, 
            Chung JY, Lee C, Cho H, Song J
  TITLE     PI3K/AKT activation induces PTEN ubiquitination and destabilization 
            accelerating tumourigenesis.
  JOURNAL   Nat Commun. 2015 Jul 17;6:7769. doi: 10.1038/ncomms8769.
  PUBMED    26183061
REFERENCE   2  (bases 1 to 6866)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6866
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        13..392
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        393..596
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     547..567
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     641..660
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        641..659
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             688..714
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             727..2166
                     /codon_start=1
                     /gene="AKT1"
                     /product="human serine/threonine protein kinase activated 
                     by growth factors"
                     /label=Akt1
                     /translation="MSDVAIVKEGWLHKRGEYIKTWRPRYFLLKNDGTFIGYKERPQDV
                     DQREAPLNNFSVAQCQLMKTERPRPNTFIIRCLQWTTVIERTFHVETPEEREEWTTAIQ
                     TVADGLKKQEEEEMDFRSGSPSDNSGAEEMEVSLAKPKHRVTMNEFEYLKLLGKGTFGK
                     VILVKEKATGRYYAMKILKKEVIVAKDEVAHTLTENRVLQNSRHPFLTALKYSFQTHDR
                     LCFVMEYANGGELFFHLSRERVFSEDRARFYGAEIVSALDYLHSEKNVVYRDLKLENLM
                     LDKDGHIKITDFGLCKEGIKDGATMKTFCGTPEYLAPEVLEDNDYGRAVDWWGLGVVMY
                     EMMCGRLPFYNQDHEKLFELILMEEIRFPRTLGPEAKSLLSGLLKKDPKQRLGGGSEDA
                     KEIMQHRFFAGIVWQHVYEKKLSPPFKPQVTSETDTRYFDEEFTAQMITITPPDQDDSM
                     ECVDSERRPHFPQFSYSASGTA"
     promoter        complement(2197..2215)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2198..2215)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2235..2252)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2241..2465
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2511..2939
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2598..2617)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2808..2829
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2948..2968)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2953..3282
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3133..3268
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3195..3214
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3349..4143
                     /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(3403..3422)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4013..4032
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4317..4438
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4354..4373)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4408..4427
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4487..4503)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4487..4503)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4500..4522)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4511..4527
                     /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(4535..4565)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4580..4601
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4718..4735)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4889..5477)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4969..4988)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(5648..6508)
                     /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     6271..6290
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6509..6613)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6688..6707)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gatatacgcg ttgacattga ttattgacta gttattaata gtaatcaatt acggggtcat
       61 tagttcatag cccatatatg gagttccgcg ttacataact tacggtaaat ggcccgcctg
      121 gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt cccatagtaa
      181 cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa actgcccact
      241 tggcagtaca tcaagtgtat catatgccaa gtacgccccc tattgacgtc aatgacggta
      301 aatggcccgc ctggcattat gcccagtaca tgaccttatg ggactttcct acttggcagt
      361 acatctacgt attagtcatc gctattacca tggtgatgcg gttttggcag tacatcaatg
      421 ggcgtggata gcggtttgac tcacggggat ttccaagtct ccaccccatt gacgtcaatg
      481 ggagtttgtt ttggcaccaa aatcaacggg actttccaaa atgtcgtaac aactccgccc
      541 cattgacgca aatgggcggt aggcgtgtac ggtgggaggt ctatataagc agagctctct
      601 ggctaactag agaacccact gcttactggc ttatcgaaat taatacgact cactataggg
      661 agacccaagc ttggtaccat ggcctcctac ccttatgatg tgccagatta tgcctctccc
      721 gaattcatga gcgacgtggc tattgtgaag gagggttggc tgcacaaacg aggggagtac
      781 atcaagacct ggcggccacg ctacttcctc ctcaagaatg atggcacctt cattggctac
      841 aaggagcggc cgcaggatgt ggaccaacgt gaggctcccc tcaacaactt ctctgtggcg
      901 cagtgccagc tgatgaagac ggagcggccc cggcccaaca ccttcatcat ccgctgcctg
      961 cagtggacca ctgtcatcga acgcaccttc catgtggaga ctcctgagga gcgggaggag
     1021 tggacaaccg ccatccagac tgtggctgac ggcctcaaga agcaggagga ggaggagatg
     1081 gacttccggt cgggctcacc cagtgacaac tcaggggctg aagagatgga ggtgtccctg
     1141 gccaagccca agcaccgcgt gaccatgaac gagtttgagt acctgaagct gctgggcaag
     1201 ggcactttcg gcaaggtgat cctggtgaag gagaaggcca caggccgcta ctacgccatg
     1261 aagatcctca agaaggaagt catcgtagcc aaggacgaag tggcccacac actcaccgag
     1321 aaccgcgtcc tgcagaactc caggcacccc ttcctcacag ccctgaagta ctctttccag
     1381 acccacgacc gcctctgctt tgtcatggag tacgccaacg ggggcgagct gttcttccac
     1441 ctgtcccggg agcgtgtgtt ctccgaggac cgggcccgct tctatggcgc tgagattgtg
     1501 tcagccctgg actacctgca ctcggagaag aacgtggtgt accgggacct caagctggag
     1561 aacctcatgc tggacaagga cgggcacatt aagatcacag acttcgggct gtgcaaggag
     1621 gggatcaagg acggtgccac catgaagacc ttttgcggca cacctgagta cctggccccc
     1681 gaggtgctgg aggacaatga ctacggccgt gcagtggact ggtgggggct gggcgtggtc
     1741 atgtacgaga tgatgtgcgg tcgcctgccc ttctacaacc aggaccatga gaagcttttt
     1801 gagctcatcc tcatggagga gatccgcttc ccgcgcacgc ttggtcccga ggccaagtcc
     1861 ttgctttcag ggctgctcaa gaaggacccc aagcagaggc ttggcggggg ctccgaggac
     1921 gccaaggaga tcatgcagca tcgcttcttt gccggtatcg tgtggcagca cgtgtacgag
     1981 aagaagctca gcccaccctt caagccccag gtcacgtcgg agactgacac caggtatttt
     2041 gatgaggagt tcacggccca gatgatcacc atcacaccac ctgaccaaga tgacagcatg
     2101 gagtgtgtgg acagcgagcg caggccccac ttcccccagt tctcctactc ggccagcggc
     2161 acggcctgac tcgagcatgc atctagaggg ccctattcta tagtgtcacc taaatgctag
     2221 agctcgctga tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc
     2281 ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga
     2341 ggaaattgca tcgcattgtc tgagtaggtg tcattctatt ctggggggtg gggtggggca
     2401 ggacagcaag ggggaggatt gggaagacaa tagcaggcat gctggggatg cggtgggctc
     2461 tatggcttct gaggcggaaa gaaccagctg gggctctagg gggtatcccc acgcgccctg
     2521 tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc
     2581 cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg
     2641 ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg
     2701 gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg
     2761 atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt
     2821 ccaaactgga acaacactca accctatctc ggtctattct tttgatttat aagggatttt
     2881 gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaatta
     2941 attctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga
     3001 agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc
     3061 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc
     3121 ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc
     3181 tgactaattt tttttattta tgcagaggcc gaggccgcct ctgcctctga gctattccag
     3241 aagtagtgag gaggcttttt tggaggccta ggcttttgca aaaagctccc gggagcttgt
     3301 atatccattt tcggatctga tcaagagaca ggatgaggat cgtttcgcat gattgaacaa
     3361 gatggattgc acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg
     3421 gcacaacaga caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc
     3481 ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga atgaactgca ggacgaggca
     3541 gcgcggctat cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc
     3601 actgaagcgg gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca
     3661 tctcaccttg ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat
     3721 acgcttgatc cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca
     3781 cgtactcgga tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg
     3841 ctcgcgccag ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc
     3901 gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct
     3961 ggattcatcg actgtggccg gctgggtgtg gcggaccgct atcaggacat agcgttggct
     4021 acccgtgata ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac
     4081 ggtatcgccg ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc
     4141 tgagcgggac tctggggttc gaaatgaccg accaagcgac gcccaacctg ccatcacgag
     4201 atttcgattc caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg
     4261 ccggctggat gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccccaact
     4321 tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata
     4381 aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc
     4441 atgtctgtat accgtcgacc tctagctaga gcttggcgta atcatggtca tagctgtttc
     4501 ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt
     4561 gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc
     4621 ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg
     4681 ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct
     4741 cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca
     4801 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga
     4861 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc
     4921 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
     4981 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
     5041 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt
     5101 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
     5161 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
     5221 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
     5281 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg
     5341 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
     5401 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
     5461 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga
     5521 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga
     5581 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt
     5641 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
     5701 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
     5761 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag
     5821 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
     5881 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
     5941 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg
     6001 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca
     6061 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
     6121 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
     6181 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
     6241 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa
     6301 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt
     6361 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
     6421 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
     6481 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt
     6541 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
     6601 taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtcgacgga tcgggagatc
     6661 tcccgatccc ctatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca
     6721 gtatctgctc cctgcttgtg tgttggaggt cgctgagtag tgcgcgagca aaatttaagc
     6781 tacaacaagg caaggcttga ccgacaattg catgaagaat ctgcttaggg ttaggcgttt
     6841 tgcgctgctt cgcgatgtac gggcca
//
