LOCUS       sequence-288136-        9605 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 9605)
  AUTHORS   Magnifico A, Ettenberg S, Yang C, Mariano J, Tiwari S, Fang S, 
            Lipkowitz S, Weissman AM
  TITLE     WW domain HECT E3s target Cbl RING finger E3s for proteasomal 
            degradation.
  JOURNAL   J Biol Chem. 2003 Oct 31. 278(44):43169-77.
  PUBMED    12907674
REFERENCE   2  (bases 1 to 9605)
  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: "J Biol 
            Chem. 2003 Oct 31. 278(44):43169-77."
COMMENT     Sequence Label: pCINeo-myc-Itch (C822S)/2860
FEATURES             Location/Qualifiers
     source          1..9605
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        77..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..592
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     543..563
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     589..613
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     intron          753..885
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     868..887
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        930..948
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      980..989
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             989..1018
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     polyA_signal    5172..5293
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5183..5202)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5237..5256
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      5479..5934
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5566..5585)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5776..5797
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        6051..6408
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        6051..6242
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(6074..6094)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      6259..6394
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             6459..7253
                     /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(6513..6532)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     7123..7142
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    7317..7365
                     /note="synthetic polyadenylation signal"
     primer_bind     complement(7406..7425)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7525..7547
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(7585..7603)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        7671..7775
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7776..7844,7845..8636)
                     /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     complement(7994..8013)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      8807..9395
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     9296..9315
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ggcattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt ccgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttacggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta caccaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctgcgatcgc ccgccccgtt
      541 gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagag ctcgtttagt
      601 gaaccgtcag atcactagaa gctttattgc ggtagtttat cacagttaaa ttgctaacgc
      661 agtcagtgct tctgacacaa cagtctcgaa cttaagctgc agtgactctc ttaaggtagc
      721 cttgcagaag ttggtcgtga ggcactgggc aggtaagtat caaggttaca agacaggttt
      781 aaggagacca atagaaactg ggcttgtcga gacagagaag actcttgcgt ttctgatagg
      841 cacctattgg tcttactgac atccactttg cctttctctc cacaggtgtc cactcccagt
      901 tcaattacag ctcttaaggc tagagtactt aatacgactc actataggct agcgtttaaa
      961 cttaagcttg gtaccagccg ccaccatgga gcagaagctg atttccgagg aggacctgaa
     1021 cggtggtagt ctgaccatga aatctcaact tcagatcact gtcatctcag ccaaacttaa
     1081 agaaaataaa aagaattggt ttggaccaag tccttatgta gaagtcacag tagatggaca
     1141 gtcaaagaag acagaaaaat gcaataatac aaacagtccc aagtggaagc agcccctcac
     1201 agttattgtt acccctacga gtaaattatg ttttcgtgtg tggagtcacc agaccctgaa
     1261 gtctgatgtt ttattgggaa ctgctggatt agatatttat gaaacattaa agtcaaacaa
     1321 tatgaaactt gaagaagtag ttatgacttt gcagcttgta ggtgacaaag agccaacaga
     1381 gacgatggga gatttgtcag tttgtcttga tgggctgcaa gtagaagctg aggttgttac
     1441 taacggtgaa acgtcatgct ccgagagtac tacacagaat gatgatggct gcagaaccag
     1501 agatgataca agagtgagca caaatggatc agaggaccct gaggttgcag cgtcagggga
     1561 aaacaagagg gccaatggga acaattctcc gtccctttca aatggtggtt ttaagccttc
     1621 tagacctcct agaccttcga gaccacctcc acccacccca cgaagaccag cttctgtcaa
     1681 tggctcacca tccacgaatt ctgacagtga tggatctagt acaggctcct tgccaccaac
     1741 aaatacaaat gtaaatacaa gtacatctga aggagcaaca tctggattaa taattcctct
     1801 tactatatct ggaggctcgg gccctaggcc tctgaatact gtaagccaag ctcccctacc
     1861 acctgggtgg gagcagagag tagaccagca tgggcgtgtt tactatgtag accatgttga
     1921 aaagcgaaca acatgggata gaccagaacc tctacctcct ggctgggaac ggcgtgtgga
     1981 caatatggga cgtatttatt atgttgatca tttcacaaga accacaacat ggcagaggcc
     2041 aaccttggaa tctgtccgga actatgaaca gtggcagcta cagcgtagtc agcttcaggg
     2101 agcaatgcag cagtttaacc agagattcat ctatgggaat caagatttgt ttgctacatc
     2161 acaaaacaaa gaatttgatc cgcttggccc gttgccccct ggatgggaga agagaactga
     2221 tagcaatggc agagtgtatt ttgtcaacca caacactcgg attactcagt gggaagaccc
     2281 cagaagccaa ggtcagttaa atgaaaagcc cttaccagaa ggctgggaaa tgagattcac
     2341 agtggatgga attccatatt ttgtggacca caatagaaga gcaactactt atatagatcc
     2401 acgaacggga aaatcagcct tagacaatgg gccccagata gcctatgtgc gggacttcaa
     2461 ggcaaaagtt cagtatttcc ggttctggtg ccagcaactg gccatgccac agcatataaa
     2521 gatcacagtg acaagaaaaa cattgtttga ggattccttt cagcagatca tgagcttcag
     2581 cccgcaagac ctgagaagac gtttgtgggt gattttccca ggagaagaag gtttagatta
     2641 tggaggtgta gcaagagaat ggttctttct tttgtcacat gaagtgttga acccaatgta
     2701 ttgcctgttt gaatatgcag ggaaggataa ctactgcctg cagataaacc ccgcttctta
     2761 catcaatcca gaccacctga aatactttcg ttttattggc agatttattg ccatggctct
     2821 gtttcatggg aaattcatag ataccggatt ttctttacca ttctataaac gtatcctgaa
     2881 taaaccagtt ggacttaagg atttagaatc tattgatcca gaattttata attcgctcat
     2941 ctgggttaaa gaaaacaaca tcgaagaatg tggtttggaa atgtacttct cagttgataa
     3001 agaaattcta ggtgaaatta agagtcatga tttgaaaccg aatggtggca atattcttgt
     3061 gacagaagaa aacaaggagg aatacatcag gatggtagct gagtggaggt tgtctcgagg
     3121 tgttgaagaa cagacacaag ctttctttga gggctttaat gaaattcttc ctcagcaata
     3181 tttgcaatat tttgatgcga aggaattaga ggttcttctg tgtgggatgc aggagattga
     3241 tttgaatgac tggcagagac atgccatcta ccgccactac accagaacaa gcaagcagat
     3301 catgtggttt tggcagtttg ttaaagaaat tgataatgag aagaggatga gacttctgca
     3361 gtttgttact ggaacctgcc gattgccagt gggaggattt gctgacctta tggggagtaa
     3421 tggaccacag aagttctgca tcgaaaaagt tggcaaagaa aattggttac ccagaagcca
     3481 tactagtttt aaccgcctgg acctcccacc ttacaagagc tatgagcaac tgaaggaaaa
     3541 gctgttattt gccattgaag aaactgaagg atttggacaa gagtaattgt ttagacctta
     3601 agccaccaga ggccagggac ttgtctgcag tgtctgtccc cttccttctc tgcatgttgc
     3661 acatcttatt gtaaacttgg actgtgacta gagagttacc tgagagtaag taaattaatg
     3721 ttatcgttga gatcttccag tcattctgga tttctactca acgtttccag aaagcagatc
     3781 tacaaactgt agtcagaaac ttaacatgag atttaacaca gcgaggaaat ttccctcgtc
     3841 ttattccact agattgttcc cttaacaaca gtttgctgta tgtccgaagt ctcacttggg
     3901 cataggtctc ttagtgggtg ttgtaaagga ttctgcaggg acatctcgga aacgcaatac
     3961 acaaggtctt cctgtaaagc ctcgctcaga ggtgcaagct taactcagtg tctgggcaca
     4021 tgagcagctg ctgcttgtgc tctccgctcg tcccagcctg gcctgactgt tgtttttcat
     4081 tcctggagca tgaatcagtg caccattctg atgctattcc tggaagtgat gcaggacttg
     4141 catttttcta caaagtagtt ttgtctattt gaattcaggg aaaacttggt ccccgccaac
     4201 atatgaccat acctctcttc attcagtttg taattgttct attgccttgc aaatactaaa
     4261 ctttacaaac agtatcagca ctgtggttcc ttcacgtttt tagaaattaa cctagaatta
     4321 aaataagttc agatgaattc atctaagatc attcttgatt tttttaaatc ctacacataa
     4381 attgaagctt tatggtgttg aagtaatcca acaccttagt tacatttcat ttttgctaaa
     4441 ctattcaagt tgcaaccatt tcctaacaca aatggaggtc agtggtgcag actctgcagt
     4501 cacaacagcg tccagcagca cacagctgcg ctgtcttcac actccactcc tgacacagga
     4561 gcagccgtgt ctaccctctt acatggtcct gatctacaat atttttctct cccctaaaaa
     4621 ttgtagattt cccttttctt tccatgactc ctttatctta ggctgctcct ttagtaactg
     4681 atgaatgcac tttgaccttc tctgtaggtc atttaccttg gcttagcatt agactgtgag
     4741 gatggttgct gccacagact ttatgggcag caagatgaaa cctcagccag gtgagtccct
     4801 attgtagcca aatcccaaaa ttagccttat ttctctaatg gggaccttga aatgtgttgg
     4861 tggcatagca taaagccaag aacaccgagt gtgacaaacc cctgtcccat aggaaagaaa
     4921 gtgtttctgt gtccctagct gctcgtcctg ttctcatcct gggacagttt gaactcgtgc
     4981 agctgtgctc cttaacactg ccatcctcag cagagacgtg acagacgggt tgcctcatga
     5041 aaaagtgatc agtgaaggga gagctcggct cctccccagc cctcttttta cccttatacc
     5101 acaagaaata catgtcatct ctagagcggc cgcttccctt tagtgagggt taatgcttcg
     5161 agcagacatg ataagataca ttgatgagtt tggacaaacc acaactagaa tgcagtgaaa
     5221 aaaatgcttt atttgtgaaa tttgtgatgc tattgcttta tttgtaacca ttataagctg
     5281 caataaacaa gttaacaaca acaattgcat tcattttatg tttcaggttc agggggagat
     5341 gtgggaggtt ttttaaagca agtaaaacct ctacaaatgt ggtaaaatcc gataaggatc
     5401 gatccgggct ggcgtaatag cgaagaggcc cgcaccgatc gcccttccca acagttgcgc
     5461 agcctgaatg gcgaatggac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg
     5521 ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct
     5581 tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc
     5641 ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg
     5701 atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt
     5761 ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg
     5821 tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc
     5881 tgatttaaca aaaatttaac gcgaatttta acaaaatatt aacgcttaca atttcctgat
     5941 gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcatacgcg gatctgcgca
     6001 gcaccatggc ctgaaataac ctctgaaaga ggaacttggt taggtacctt ctgaggcgga
     6061 aagaaccagc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca
     6121 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca
     6181 ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc
     6241 ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc
     6301 catggctgac taattttttt tatttatgca gaggccgagg ccgcctcggc ctctgagcta
     6361 ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa gcttgattct
     6421 tctgacacaa cagtctcgaa cttaaggcta gagccaccat gattgaacaa gatggattgc
     6481 acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga
     6541 caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt
     6601 ttgtcaagac cgacctgtcc ggtgccctga atgaactgca ggacgaggca gcgcggctat
     6661 cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg
     6721 gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg
     6781 ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc
     6841 cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga
     6901 tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag
     6961 ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc gtcgtgaccc
     7021 atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg
     7081 actgtggccg gctgggtgtg gcggaccgct atcaggacat agcgttggct acccgtgata
     7141 ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg
     7201 ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc tgagcgggac
     7261 tctggggttc gaaatgaccg accaagcgac gcccaacctg ccatcacgat ggccgcaata
     7321 aaatatcttt attttcatta catctgtgtg ttggtttttt gtgtgaatcg atagcgataa
     7381 ggatccgcgt atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagc
     7441 cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg
     7501 cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat
     7561 caccgaaacg cgcgagacga aagggcctcg tgatacgcct atttttatag gttaatgtca
     7621 tgataataat ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc
     7681 ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct
     7741 gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg
     7801 cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg
     7861 tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc
     7921 tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca
     7981 cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac
     8041 tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa
     8101 agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg
     8161 ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt
     8221 ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg
     8281 aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc
     8341 gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga
     8401 tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta
     8461 ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc
     8521 cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg
     8581 atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt
     8641 cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa
     8701 ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt
     8761 cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt
     8821 ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt
     8881 tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga
     8941 taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag
     9001 caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata
     9061 agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg
     9121 gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga
     9181 gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca
     9241 ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa
     9301 acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt
     9361 tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac
     9421 ggttcctggc cttttgctgg ccttttgctc acatggctcg acagatcttc aatattggcc
     9481 attagccata ttattcattg gttatatagc ataaatcaat attggctatt ggccattgca
     9541 tacgttgtat ctatatcata atatgtacat ttatattggc tcatgtccaa tatgaccgcc
     9601 atgtt
//
