LOCUS       sequence-166162-        6897 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6897)
  AUTHORS   Ramaswamy S, Nakamura N, Vazquez F, Batt DB, Perera S, Roberts TM, 
            Sellers WR
  TITLE     Regulation of G1 progression by the PTEN tumor suppressor protein is
            linked to inhibition of the phosphatidylinositol 3-kinase/Akt 
            pathway.
  JOURNAL   Proc Natl Acad Sci U S A. 1999 Mar 2. 96(5):2110-5.
  PUBMED    10051603
REFERENCE   2  (bases 1 to 6897)
  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"; journalName: "Proc Natl 
            Acad Sci U S A. 1999 Mar 2. 96(5):2110-5."
COMMENT     Sequence Label: 1013 pcDNA3 T7 Akt1 T308A S473A
FEATURES             Location/Qualifiers
     source          1..6897
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      895..904
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             937..2376
                     /codon_start=1
                     /gene="AKT1"
                     /product="human serine/threonine protein kinase activated 
                     by growth factors"
                     /label=Akt1
                     /translation="MSDVAIVKEGWLHKRGEYIKTWRPRYFLLKNDGTFIGYKERPQDV
                     DQREAPLNNFSVAQCQLMKTERPRPNTFIIRCLQWTTVIERTFHVETPEEREEWTTAIQ
                     TVADGLKKQEEEEMDFRSGSPSDNSGAEEMEVSLAKPKHRVTMNEFEYLKLLGKGTFGK
                     VILVKEKATGRYYAMKILKKEVIVAKDEVAHTLTENRVLQNSRHPFLTALKYSFQTHDR
                     LCFVMEYANGGELFFHLSRERVFSEDRARFYGAEIVSALDYLHSEKNVVYRDLKLENLM
                     LDKDGHIKITDFGLCKEGIKDGATMKAFCGTPEYLAPEVLEDNDYGRAVDWWGLGVVMY
                     EMMCGRLPFYNQDHEKLFELILMEEIRFPRTLGPEAKSLLSGLLKKDPKQRLGGGSEDA
                     KEIMQHRFFAGIVWQHVYEKKLSPPFKPQVTSETDTRYFDEEFTAQMITITPPDQDDSM
                     ECVDSERRPHFPQFAYSASGTA"
     promoter        complement(2450..2468)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2451..2468)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2488..2505)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2494..2718
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2764..3192
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2851..2870)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3061..3082
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3201..3221)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3206..3535
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3386..3521
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3448..3467
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3602..4396
                     /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(3656..3675)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4266..4285
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4570..4691
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4607..4626)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4661..4680
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4740..4756)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4740..4756)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4753..4775)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4764..4780
                     /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(4788..4794,4795..4812,4813..4818))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4833..4854
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4971..4988)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5142..5730)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5222..5241)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5901..6692,6693..6761))
                     /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     6524..6543
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6762..6866)
                     /gene="bla"
                     /label=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 ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gcttgccacc
      901 atggctagca tgactggtgg acagcaaatg ggatccatga gcgacgtggc tattgtgaag
      961 gagggttggc tgcacaaacg aggggagtac atcaagacct ggcggccacg ctacttcctc
     1021 ctcaagaatg atggcacctt cattggctac aaggagcggc cgcaggatgt ggaccaacgt
     1081 gaggctcccc tcaacaactt ctctgtggcg cagtgccagc tgatgaagac ggagcggccc
     1141 cggcccaaca ccttcatcat ccgctgcctg cagtggacca ctgtcatcga acgcaccttc
     1201 catgtggaga ctcctgagga gcgggaggag tggacaaccg ccatccagac tgtggctgac
     1261 ggcctcaaga agcaggagga ggaggagatg gacttccggt cgggctcacc cagtgacaac
     1321 tcaggggctg aagagatgga ggtgtccctg gccaagccca agcaccgcgt gaccatgaac
     1381 gagtttgagt acctgaagct gctgggcaag ggcactttcg gcaaggtgat cctggtgaag
     1441 gagaaggcca caggccgcta ctacgccatg aagatcctca agaaggaagt catcgtggcc
     1501 aaggacgagg tggcccacac actcaccgag aaccgcgtcc tgcagaactc caggcacccc
     1561 ttcctcacag ccctgaagta ctctttccag acccacgacc gcctctgctt tgtcatggag
     1621 tacgccaacg ggggcgagct gttcttccac ctgtcccggg aacgtgtgtt ctccgaggac
     1681 cgggcccgct tctatggcgc tgagattgtg tcagccctgg actacctgca ctcggagaag
     1741 aacgtggtgt accgggacct caagctggag aacctcatgc tggacaagga cgggcacatt
     1801 aagatcacag acttcgggct gtgcaaggag gggatcaagg acggtgccac catgaaggcc
     1861 ttttgcggca cacctgagta cctggccccc gaggtgctgg aggacaatga ctacggccgt
     1921 gcagtggact ggtgggggct gggcgtggtc atgtacgaga tgatgtgcgg tcgcctgccc
     1981 ttctacaacc aggaccatga gaagcttttt gagctcatcc tcatggagga gatccgcttc
     2041 ccgcgcacgc ttggtcccga ggccaagtcc ttgctttcag ggctgctcaa gaaggacccc
     2101 aagcagaggc ttggcggggg ctccgaggac gccaaggaga tcatgcagca tcgcttcttt
     2161 gccggtatcg tgtggcagca cgtgtacgag aagaagctca gcccaccctt caagccccag
     2221 gtcacgtcgg agactgacac caggtatttt gatgaggagt tcacggccca gatgatcacc
     2281 atcacaccac ctgaccaaga tgacagcatg gagtgtgtgg acagcgagcg caggccccac
     2341 ttcccccagt tcgcctactc ggccagcggc acggcctgag gcggcggtgg aattctgcag
     2401 atatccatca cactggcggc cgctcgagca tgcatctaga gggccctatt ctatagtgtc
     2461 acctaaatgc tagagctcgc tgatcagcct cgactgtgcc ttctagttgc cagccatctg
     2521 ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc actgtccttt
     2581 cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct attctggggg
     2641 gtggggtggg gcaggacagc aagggggagg attgggaaga caatagcagg catgctgggg
     2701 atgcggtggg ctctatggct tctgaggcgg aaagaaccag ctggggctct agggggtatc
     2761 cccacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga
     2821 ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg
     2881 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat
     2941 ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg
     3001 ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata
     3061 gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt
     3121 tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat
     3181 ttaacgcgaa ttaattctgt ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc
     3241 cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca ggtgtggaaa
     3301 gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac
     3361 catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc
     3421 tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg cctctgcctc
     3481 tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt gcaaaaagct
     3541 cccgggagct tgtatatcca ttttcggatc tgatcaagag acaggatgag gatcgtttcg
     3601 catgattgaa caagatggat tgcacgcagg ttctccggcc gcttgggtgg agaggctatt
     3661 cggctatgac tgggcacaac agacaatcgg ctgctctgat gccgccgtgt tccggctgtc
     3721 agcgcagggg cgcccggttc tttttgtcaa gaccgacctg tccggtgccc tgaatgaact
     3781 gcaggacgag gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt gcgcagctgt
     3841 gctcgacgtt gtcactgaag cgggaaggga ctggctgcta ttgggcgaag tgccggggca
     3901 ggatctcctg tcatctcacc ttgctcctgc cgagaaagta tccatcatgg ctgatgcaat
     3961 gcggcggctg catacgcttg atccggctac ctgcccattc gaccaccaag cgaaacatcg
     4021 catcgagcga gcacgtactc ggatggaagc cggtcttgtc gatcaggatg atctggacga
     4081 agagcatcag gggctcgcgc cagccgaact gttcgccagg ctcaaggcgc gcatgcccga
     4141 cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca tggtggaaaa
     4201 tggccgcttt tctggattca tcgactgtgg ccggctgggt gtggcggacc gctatcagga
     4261 catagcgttg gctacccgtg atattgctga agagcttggc ggcgaatggg ctgaccgctt
     4321 cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc atcgccttct atcgccttct
     4381 tgacgagttc ttctgagcgg gactctgggg ttcgaaatga ccgaccaagc gacgcccaac
     4441 ctgccatcac gagatttcga ttccaccgcc gccttctatg aaaggttggg cttcggaatc
     4501 gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct ggagttcttc
     4561 gcccacccca acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca
     4621 aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc
     4681 aatgtatctt atcatgtctg tataccgtcg acctctagct agagcttggc gtaatcatgg
     4741 tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc
     4801 ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg
     4861 ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc
     4921 ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact
     4981 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta
     5041 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag
     5101 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
     5161 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
     5221 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg
     5281 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc
     5341 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac
     5401 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
     5461 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
     5521 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
     5581 agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
     5641 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag
     5701 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct
     5761 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg
     5821 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat
     5881 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc
     5941 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg
     6001 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct
     6061 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca
     6121 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg
     6181 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg
     6241 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc
     6301 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag
     6361 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg
     6421 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag
     6481 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat
     6541 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg
     6601 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca
     6661 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca
     6721 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat
     6781 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag
     6841 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgtc
//
