LOCUS       sequence-401603-        8924 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 8924)
  AUTHORS   Hu J, Zacharek S, He YJ, Lee H, Shumway S, Duronio RJ, Xiong Y
  TITLE     WD40 protein FBW5 promotes ubiquitination of tumor suppressor TSC2 
            by DDB1-CUL4-ROC1 ligase.
  JOURNAL   Genes Dev. 2008 Apr 1. 22(7):866-71.
  PUBMED    18381890
REFERENCE   2  (bases 1 to 8924)
  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: "Genes Dev. 
            2008 Apr 1. 22(7):866-71."
COMMENT     Sequence Label: pcDNA3-HA3-TSC1
FEATURES             Location/Qualifiers
     source          1..8924
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..204
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     155..175
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     249..268
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        249..267
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             283..309
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             313..339
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             319..345
                     /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="YDVPDYASL"
     promoter        complement(3863..3881)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(3864..3881)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(3901..3918)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3907..4131
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4177..4605
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4264..4283)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4474..4495
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4614..4634)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4619..4948
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4799..4934
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4861..4880
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             5015..5809
                     /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(5069..5088)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5679..5698
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5983..6104
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6020..6039)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6074..6093
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6153..6169)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6153..6169)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6166..6188)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6177..6193
                     /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(6201..6207,6208..6225,6226..6231))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6246..6267
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6384..6401)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6555..7143)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6635..6654)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7314..8105,8106..8174))
                     /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     7937..7956
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8175..8279)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(8354..8373)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        8545..8924
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 gtgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt
       61 ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac
      121 tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg
      181 tgggaggtct atataagcag agctctctgg ctaactagag aacccactgc ttactggctt
      241 atcgaaatta atacgactca ctatagggag acccaagcta tgtacccata cgatgttcct
      301 gactatgcgg gctatcccta tgacgtcccg gactatgcaa gcttggtacc tatggcccaa
      361 caagcaaatg tcggggagct tcttgccatg ctggactccc ccatgctggg tgtgcgggac
      421 gacgtgacag ctgtctttaa agagaacctc aattctgacc gtggccctat gcttgtaaac
      481 accttggtgg attattacct ggaaaccagc tctcagccgg cattgcacat cctgaccacc
      541 ttgcaagagc cacatgacaa gcacctcttg gacaggatta acgaatatgt gggcaaagcc
      601 gccactcgtt tatccatcct ctcgttactg ggtcatgtca taagactgca gccatcttgg
      661 aagcataagc tctctcaagc acctcttttg ccttctttac taaaatgtct caagatggac
      721 actgacgtcg ttgtcctcac aacaggcgtc ttggtgttga taaccatgct accaatgatt
      781 ccacagtctg ggaaacagca tcttcttgat ttctttgaca tttttggccg tctgtcatca
      841 tggtgcctga agaaaccagg ccacgtggcg gaagtctatc tcgtccatct ccatgccagt
      901 gtgtacgcac tctttcatcg cctttatgga atgtaccctt gcaacttcgt ctcctttttg
      961 cgttctcatt acagtatgaa agaaaacctg gagacttttg aagaagtggt caagccaatg
     1021 atggagcatg tgcgaattca tccggaatta gtgactggat ccaaggacca tgaactggac
     1081 cctcgaaggt ggaagagatt agaaactcat gatgttgtga tcgagtgtgc caaaatctct
     1141 ctggatccca cagaagcctc atatgaagat ggctattctg tgtctcacca aatctcagcc
     1201 cgctttcctc atcgttcagc cgatgtcacc accagccctt atgctgacac acagaatagc
     1261 tatgggtgtg ctacttctac cccttactcc acgtctcggc tgatgttgtt aaatatgcca
     1321 gggcagctac ctcagactct gagttcccca tcgacacggc tgataactga accaccacaa
     1381 gctactcttt ggagcccatc tatggtttgt ggtatgacca ctcctccaac ttctcctgga
     1441 aatgtcccac ctgatctgtc acacccttac agtaaagtct ttggtacaac tgcaggtgga
     1501 aaaggaactc ctctgggaac cccagcaacc tctcctcctc cagccccact ctgtcattcg
     1561 gatgactacg tgcacatttc actcccccag gccacagtca caccccccag gaaggaagag
     1621 agaatggatt ctgcaagacc atgtctacac agacaacacc atcttctgaa tgacagagga
     1681 tcagaagagc cacctggcag caaaggttct gtcactctaa gtgatcttcc agggttttta
     1741 ggtgatctgg cctctgaaga agatagtatt gaaaaagata aagaagaagc tgcaatatct
     1801 agagaacttt ctgagatcac cacagcagag gcagagcctg tggttcctcg aggaggcttt
     1861 gactctccct tttaccgaga cagtctccca ggttctcagc ggaagaccca ctcggcagcc
     1921 tccagttctc agggcgccag cgtgaaccct gagcctttac actcctccct ggacaagctt
     1981 gggcctgaca caccaaagca agcctttact cccatagacc tgccctgcgg cagtgctgat
     2041 gaaagccctg cgggagacag ggaatgccag acttctttgg agaccagtat cttcactccc
     2101 agtccttgta aaattccacc tccgacgaga gtgggctttg gaagcgggca gcctcccccg
     2161 tatgatcatc tttttgaggt ggcattgcca aagacagccc atcattttgt catcaggaag
     2221 actgaggagc tgttaaagaa agcaaaagga aacacagagg aagatggtgt gccctctacc
     2281 tccccaatgg aagtgctgga cagactgata cagcagggag cagacgcgca cagcaaggag
     2341 ctgaacaagt tgcctttacc cagcaagtct gtcgactgga cccactttgg aggctctcct
     2401 ccttcagatg agatccgcac cctccgagac cagttgcttt tactgcacaa ccagttactc
     2461 tatgagcgtt ttaagaggca gcagcatgcc ctccggaaca ggcggctcct ccgcaaggtg
     2521 atcaaagcag cagctctgga ggaacataat gctgccatga aagatcagtt gaagttacaa
     2581 gagaaggaca tccagatgtg gaaggttagt ctgcagaaag aacaagctag atacaatcag
     2641 ctccaggagc agcgtgacac tatggtaacc aagctccaca gccagatcag acagctgcag
     2701 catgaccgag aggaattcta caaccagagc caggaattac agacgaagct ggaggactgc
     2761 aggaacatga ttgcggagct gcggatagaa ctgaagaagg ccaacaacaa ggtgtgtcac
     2821 actgagctgc tgctcagtca ggtttcccaa aagctctcaa acagtgagtc ggtccagcag
     2881 cagatggagt tcttgaacag gcagctgttg gttcttgggg aggtcaacga gctctatttg
     2941 gaacaactgc agaacaagca ctcagatacc acaaaggaag tagaaatgat gaaagccgcc
     3001 tatcggaaag agctagaaaa aaacagaagc catgttctcc agcagactca gaggcttgat
     3061 acctcccaaa aacggatttt ggaactggaa tctcacctgg ccaagaaaga ccaccttctt
     3121 ttggaacaga agaaatatct agaggatgtc aaactccagg caagaggaca gctgcaggcc
     3181 gcagagagca ggtatgaggc tcagaaaagg ataacccagg tgtttgaatt ggagatctta
     3241 gatttatatg gcaggttgga gaaagatggc ctcctgaaaa aacttgaaga agaaaaagca
     3301 gaagcagctg aagcagcaga agaaaggctt gactgttgta atgacgggtg ctcagattcc
     3361 atggtagggc acaatgaaga ggcatctggc cacaacggtg agaccaagac ccccaggccc
     3421 agcagcgccc ggggcagtag tggaagcaga ggtggtggag gcagcagcag cagcagcagc
     3481 gagctttcta ccccagagaa acccccacac cagagggcag gcccattcag cagtcggtgg
     3541 gagacgacta tgggagaagc gtctgccagc atccccacca ctgtgggctc acttcccagt
     3601 tcaaaaagct tcctgggtat gaaggctcga gagttatttc gtaataagag cgagagccag
     3661 tgtgatgagg acggcatgac cagtagcctt tctgagagcc taaagacaga actgggcaaa
     3721 gacttgggtg tggaagccaa gattcccctg aacctagatg gccctcaccc gtctcccccg
     3781 accccggaca gtgttggaca gctacatatc atggactaca atgagactca tcatgaacac
     3841 agctaagact agagggccct attctatagt gtcacctaaa tgctagagct cgctgatcag
     3901 cctcgactgt gccttctagt tgccagccat ctgttgtttg cccctccccc gtgccttcct
     3961 tgaccctgga aggtgccact cccactgtcc tttcctaata aaatgaggaa attgcatcgc
     4021 attgtctgag taggtgtcat tctattctgg ggggtggggt ggggcaggac agcaaggggg
     4081 aggattggga agacaatagc aggcatgctg gggatgcggt gggctctatg gcttctgagg
     4141 cggaaagaac cagctggggc tctagggggt atccccacgc gccctgtagc ggcgcattaa
     4201 gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc
     4261 ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag
     4321 ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca
     4381 aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc
     4441 gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa
     4501 cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct
     4561 attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattaattc tgtggaatgt
     4621 gtgtcagtta gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat
     4681 gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag
     4741 tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat
     4801 cccgccccta actccgccca gttccgccca ttctccgccc catggctgac taattttttt
     4861 tatttatgca gaggccgagg ccgcctctgc ctctgagcta ttccagaagt agtgaggagg
     4921 cttttttgga ggcctaggct tttgcaaaaa gctcccggga gcttgtatat ccattttcgg
     4981 atctgatcaa gagacaggat gaggatcgtt tcgcatgatt gaacaagatg gattgcacgc
     5041 aggttctccg gccgcttggg tggagaggct attcggctat gactgggcac aacagacaat
     5101 cggctgctct gatgccgccg tgttccggct gtcagcgcag gggcgcccgg ttctttttgt
     5161 caagaccgac ctgtccggtg ccctgaatga actgcaggac gaggcagcgc ggctatcgtg
     5221 gctggccacg acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag
     5281 ggactggctg ctattgggcg aagtgccggg gcaggatctc ctgtcatctc accttgctcc
     5341 tgccgagaaa gtatccatca tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc
     5401 tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga
     5461 agccggtctt gtcgatcagg atgatctgga cgaagagcat caggggctcg cgccagccga
     5521 actgttcgcc aggctcaagg cgcgcatgcc cgacggcgag gatctcgtcg tgacccatgg
     5581 cgatgcctgc ttgccgaata tcatggtgga aaatggccgc ttttctggat tcatcgactg
     5641 tggccggctg ggtgtggcgg accgctatca ggacatagcg ttggctaccc gtgatattgc
     5701 tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg ctttacggta tcgccgctcc
     5761 cgattcgcag cgcatcgcct tctatcgcct tcttgacgag ttcttctgag cgggactctg
     5821 gggttcgaaa tgaccgacca agcgacgccc aacctgccat cacgagattt cgattccacc
     5881 gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc
     5941 ctccagcgcg gggatctcat gctggagttc ttcgcccacc ccaacttgtt tattgcagct
     6001 tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc atttttttca
     6061 ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctgtataccg
     6121 tcgacctcta gctagagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt
     6181 tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt
     6241 gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg
     6301 ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg
     6361 cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg
     6421 cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat
     6481 aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc
     6541 gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc
     6601 tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga
     6661 agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt
     6721 ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg
     6781 taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc
     6841 gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg
     6901 gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc
     6961 ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg
     7021 ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc
     7081 gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct
     7141 caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt
     7201 taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa
     7261 aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa
     7321 tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc
     7381 tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct
     7441 gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca
     7501 gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt
     7561 aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt
     7621 gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc
     7681 ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc
     7741 tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt
     7801 atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact
     7861 ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc
     7921 ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt
     7981 ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg
     8041 atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct
     8101 gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa
     8161 tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt
     8221 ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc
     8281 acatttcccc gaaaagtgcc acctgacgtc gacggatcgg gagatctccc gatcccctat
     8341 ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagtat ctgctccctg
     8401 cttgtgtgtt ggaggtcgct gagtagtgcg cgagcaaaat ttaagctaca acaaggcaag
     8461 gcttgaccga caattgcatg aagaatctgc ttagggttag gcgttttgcg ctgcttcgcg
     8521 atgtacgggc cagatatacg cgttgacatt gattattgac tagttattaa tagtaatcaa
     8581 ttacggggtc attagttcat agcccatata tggagttccg cgttacataa cttacggtaa
     8641 atggcccgcc tggctgaccg cccaacgacc cccgcccatt gacgtcaata atgacgtatg
     8701 ttcccatagt aacgccaata gggactttcc attgacgtca atgggtggag tatttacggt
     8761 aaactgccca cttggcagta catcaagtgt atcatatgcc aagtacgccc cctattgacg
     8821 tcaatgacgg taaatggccc gcctggcatt atgcccagta catgacctta tgggactttc
     8881 ctacttggca gtacatctac gtattagtca tcgctattac catg
//
