LOCUS       sequence-490796-        7005 bp    DNA     circular SYN 27-OCT-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7005)
  AUTHORS   Hsieh AC, Bo R, Manola J, Vazquez F, Bare O, Khvorova A, Scaringe S,
            Sellers WR
  TITLE     A library of siRNA duplexes targeting the phosphoinositide 3-kinase 
            pathway: determinants of gene silencing for use in cell-based 
            screens.
  JOURNAL   Nucleic Acids Res. 2004  . 32(3):893-901.
  PUBMED    14769947
REFERENCE   2  (bases 1 to 7005)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Oct 27, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nucleic 
            Acids Res. 2004  . 32(3):893-901."
COMMENT     Sequence Label: 1477 pcDNA3 flag HA Akt1
FEATURES             Location/Qualifiers
     source          1..7005
                     /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"
     CDS             901..924
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             937..963
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YPYDVPDYA"
     CDS             937..963
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             973..2412
                     /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(2558..2576)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2559..2576)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2596..2613)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2602..2826
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2872..3300
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2959..2978)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3169..3190
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3309..3329)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3314..3643
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3494..3629
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3556..3575
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3710..4504
                     /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(3764..3783)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4374..4393
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4678..4799
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4715..4734)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4769..4788
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4848..4864)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4848..4864)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4861..4883)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4872..4888
                     /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(4896..4902,4903..4920,4921..4926))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4941..4962
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5079..5096)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5250..5838)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5330..5349)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6009..6800,6801..6869))
                     /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     6632..6651
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6870..6974)
                     /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 gcttaccatg
      901 gactacaagg acgacgatga caagctcgat ggaggatacc cctacgacgt gcccgactac
      961 gccagcggat ccatgagcga cgtggctatt gtgaaggagg gttggctgca caaacgaggg
     1021 gagtacatca agacctggcg gccacgctac ttcctcctca agaatgatgg caccttcatt
     1081 ggctacaagg agcggccgca ggatgtggac caacgtgagg ctcccctcaa caacttctct
     1141 gtggcgcagt gccagctgat gaagacggag cggccccggc ccaacacctt catcatccgc
     1201 tgcctgcagt ggaccactgt catcgaacgc accttccatg tggagactcc tgaggagcgg
     1261 gaggagtgga caaccgccat ccagactgtg gctgacggcc tcaagaagca ggaggaggag
     1321 gagatggact tccggtcggg ctcacccagt gacaactcag gggctgaaga gatggaggtg
     1381 tccctggcca agcccaagca ccgcgtgacc atgaacgagt ttgagtacct gaagctgctg
     1441 ggcaagggca ctttcggcaa ggtgatcctg gtgaaggaga aggccacagg ccgctactac
     1501 gccatgaaga tcctcaagaa ggaagtcatc gtggccaagg acgaggtggc ccacacactc
     1561 accgagaacc gcgtcctgca gaactccagg caccccttcc tcacagccct gaagtactct
     1621 ttccagaccc acgaccgcct ctgctttgtc atggagtacg ccaacggggg cgagctgttc
     1681 ttccacctgt cccgggaacg tgtgttctcc gaggaccggg cccgcttcta tggcgctgag
     1741 attgtgtcag ccctggacta cctgcactcg gagaagaacg tggtgtaccg ggacctcaag
     1801 ctggagaacc tcatgctgga caaggacggg cacattaaga tcacagactt cgggctgtgc
     1861 aaggagggga tcaaggacgg tgccaccatg aagacctttt gcggcacacc tgagtacctg
     1921 gcccccgagg tgctggagga caatgactac ggccgtgcag tggactggtg ggggctgggc
     1981 gtggtcatgt acgagatgat gtgcggtcgc ctgcccttct acaaccagga ccatgagaag
     2041 ctttttgagc tcatcctcat ggaggagatc cgcttcccgc gcacgcttgg tcccgaggcc
     2101 aagtccttgc tttcagggct gctcaagaag gaccccaagc agaggcttgg cgggggctcc
     2161 gaggacgcca aggagatcat gcagcatcgc ttctttgccg gtatcgtgtg gcagcacgtg
     2221 tacgagaaga agctcagccc acccttcaag ccccaggtca cgtcggagac tgacaccagg
     2281 tattttgatg aggagttcac ggcccagatg atcaccatca caccacctga ccaagatgac
     2341 agcatggagt gtgtggacag cgagcgcagg ccccacttcc cccagttctc ctactcggcc
     2401 agcggcacgg cctgaggcgg cggtggaatt cccaattgca cctgaaattg cactggaact
     2461 gctgatggct gcgaacttct tagattgtta aggtctccaa ttctgcagat atccatcaca
     2521 ctggcggccg ctcgagcatg catctagagg gccctattct atagtgtcac ctaaatgcta
     2581 gagctcgctg atcagcctcg actgtgcctt ctagttgcca gccatctgtt gtttgcccct
     2641 cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtcctttcc taataaaatg
     2701 aggaaattgc atcgcattgt ctgagtaggt gtcattctat tctggggggt ggggtggggc
     2761 aggacagcaa gggggaggat tgggaagaca atagcaggca tgctggggat gcggtgggct
     2821 ctatggcttc tgaggcggaa agaaccagct ggggctctag ggggtatccc cacgcgccct
     2881 gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg
     2941 ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg
     3001 gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac
     3061 ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct
     3121 gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt
     3181 tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta taagggattt
     3241 tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt
     3301 aattctgtgg aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc cagcaggcag
     3361 aagtatgcaa agcatgcatc tcaattagtc agcaaccagg tgtggaaagt ccccaggctc
     3421 cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca tagtcccgcc
     3481 cctaactccg cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg
     3541 ctgactaatt ttttttattt atgcagaggc cgaggccgcc tctgcctctg agctattcca
     3601 gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaaaagctcc cgggagcttg
     3661 tatatccatt ttcggatctg atcaagagac aggatgagga tcgtttcgca tgattgaaca
     3721 agatggattg cacgcaggtt ctccggccgc ttgggtggag aggctattcg gctatgactg
     3781 ggcacaacag acaatcggct gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg
     3841 cccggttctt tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc aggacgaggc
     3901 agcgcggcta tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt
     3961 cactgaagcg ggaagggact ggctgctatt gggcgaagtg ccggggcagg atctcctgtc
     4021 atctcacctt gctcctgccg agaaagtatc catcatggct gatgcaatgc ggcggctgca
     4081 tacgcttgat ccggctacct gcccattcga ccaccaagcg aaacatcgca tcgagcgagc
     4141 acgtactcgg atggaagccg gtcttgtcga tcaggatgat ctggacgaag agcatcaggg
     4201 gctcgcgcca gccgaactgt tcgccaggct caaggcgcgc atgcccgacg gcgaggatct
     4261 cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc
     4321 tggattcatc gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc
     4381 tacccgtgat attgctgaag agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta
     4441 cggtatcgcc gctcccgatt cgcagcgcat cgccttctat cgccttcttg acgagttctt
     4501 ctgagcggga ctctggggtt cgaaatgacc gaccaagcga cgcccaacct gccatcacga
     4561 gatttcgatt ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac
     4621 gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccccaac
     4681 ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat
     4741 aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat
     4801 catgtctgta taccgtcgac ctctagctag agcttggcgt aatcatggtc atagctgttt
     4861 cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag
     4921 tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg
     4981 cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg
     5041 gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc
     5101 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc
     5161 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg
     5221 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     5281 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     5341 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
     5401 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
     5461 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
     5521 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
     5581 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
     5641 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt
     5701 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
     5761 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
     5821 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg
     5881 aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag
     5941 atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg
     6001 tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt
     6061 tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca
     6121 tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca
     6181 gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc
     6241 tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt
     6301 ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg
     6361 gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc
     6421 aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg
     6481 ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga
     6541 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga
     6601 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta
     6661 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg
     6721 ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact
     6781 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata
     6841 agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt
     6901 tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa
     6961 ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtc
//
