LOCUS       sequence-194447-        8333 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 8333)
  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 8333)
  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: pcDNA-HA-Nedd4/2795
FEATURES             Location/Qualifiers
     source          1..8333
                     /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      934..943
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             946..972
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     primer_bind     complement(3924..3941)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3930..4154
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4200..4628
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4287..4306)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4497..4518
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4637..4657)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4642..4971
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4822..4957
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4884..4903
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             5038..5832
                     /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(5092..5111)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5702..5721
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    6006..6127
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6043..6062)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6097..6116
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6176..6192)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6176..6192)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6189..6211)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6200..6216
                     /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(6224..6230,6231..6248,6249..6254))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6269..6290
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6407..6424)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6578..7166)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6658..6677)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7337..8128,8129..8197))
                     /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     7960..7979
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8198..8302)
                     /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 gctggctagc
      901 gtttaaacgg gccctctaga ctcgagcggc cgcgccacca tggcctaccc atacgacgtc
      961 ccagactacg ctgagctcgg gagtggaatc cttaccagcg tgcagacaaa aactatcaaa
     1021 aagtctttga atccaaaatg gaatgaagaa atactgttca gggtccttcc acagcgacac
     1081 cgcattcttt tcgaagtgtt tgatgaaaat cgtttgacaa gagatgattt cctaggtcaa
     1141 gtggatgtcc ctctctatcc tttaccgact gaaaacccaa gaatggagag accatataca
     1201 tttaaggatt ttgttcttca tccaagaagt cacaaatcaa gagttaaagg ttatctgaga
     1261 ttaaaaatga cttatttacc taaaaatggc tcagaagatg aaaatgcaga ccaggctgag
     1321 gagttagagc ctggctgggt tgttttggac caaccagatg ctgccactca tttgccgcat
     1381 ccaccagaac cctctcccct acctccagga tgggaagaga ggcaggatgt ccttggaagg
     1441 acctactacg taaaccatga atctagaaga acacagtgga aaaggccaag ccctgacgat
     1501 gacctcacgg atgaagacaa tgatgatatg cagctgcaag cgcagcgagc attcaccacc
     1561 aggcggcaga tatcggagga tgtggatggc cctgacaacc gggagtcccc tgagaattgg
     1621 gaaatcgtac gagaagatga aaacaccgag tatagtggtc aggctgtcca gtcacctcca
     1681 tcgggtcaca ttgatgtgca gactcacctt gcagaagagt ttaataccag acttgccgtg
     1741 tgtggaaatc cagccaccag ccagccggtt accagctcaa atcattccag cagaggaggc
     1801 agcttgcaga cctgtatctt tgaggaacag cctacacttc ctgtgctttt gcctacttca
     1861 tctggattgc caccaggttg ggaagaaaaa caagatgaca gaggaagatc atactatgta
     1921 gaccacaact ctaaaaccac cacatggtcc aagcccacca tgcaggatga tccaagatcg
     1981 aaaatccctg ctcatctgag aggaaagact gactccaatg acctgggacc cttacctcca
     2041 ggctgggaag aaagaaccca cacagatggg cgagtcttct tcataaacca caatataaag
     2101 aagacccagt gggaagatcc tcgcctgcag aacgtggcaa tcactggacc agcagtgccc
     2161 tactccagag attacaagag aaagtacgag ttcttcagaa ggaagctcaa gaagcagact
     2221 gacattccaa acaaatttga aatgaagctt cgccgcgcaa acattctgga ggattcttac
     2281 cggaggatta tgggtgtgaa gagagctgac ttgctcaagg ccagactctg gattgagttt
     2341 gatggtgaaa agggccttga ctatggaggg gttgccagag agtggttctt cctcatctcg
     2401 aaggaaatgt tcaaccctta ctacggcctg tttgaatatt ctgctacgga taattacacc
     2461 ctacagataa atcctaactc gggcttgtgt aatgaagatc acctctcata cttcaagttc
     2521 attggccgtg tggctgggat ggcagtttat catggcaagc tgttggatgg gtttttcatc
     2581 cgtccgtttt acaagatgat gcttcagaaa ctgataacac tgcacgacat ggagtccgtg
     2641 gatagtgaat attacagttc tctgcgatgg attcttgaaa atgacccgac ggagctggac
     2701 ctgagattta tcatagatga agaacttttt ggacagacac atcagcacga actgaaaacc
     2761 ggaggatcag agattgttgt caccaataag aacaaaaagg agtatatcta ccttgtaata
     2821 caatggcgat ttgtgaaccg tatccagaag caaatggcag cttttaaaga gggatttttt
     2881 gaactgatac cacaggatct catcaagata tttgatgaaa atgagctaga gcttctcatg
     2941 tgtggtctgg gagatgtgaa tgtgaacgac tggcgggaac acacaaaata caaaaatggc
     3001 tacagcatga accaccaggt catccactgg ttctggaagg ctgtttggat gatggattcg
     3061 gaaaaaagaa tacgcttact tcagtttgtc actggcacat cccgtgtgcc gatgaatggg
     3121 tttgctgaac tctatggctc gaatggacca caatccttca cagtggaaca atggggcacc
     3181 cctgataagc tgccaagagc acacacctgc ttcaatcgcc tggacctgcc accctacgaa
     3241 tcctttgacg aactctggga taaacttcag atggcaattg agaacacaca gggctttgat
     3301 ggcgttgatt agattacaaa aaaaaacaag ctgtggtgtc ttgtacacca tagtttctaa
     3361 gcaaaagcaa aaacaaaatc acattttaac attttccaga tagtttctaa aagatggtca
     3421 ctggctcttc ccaccgagac cagaggacag tttatccatg atttagccac caccatgctg
     3481 acctgttcac ttgtccagtt actttgtcca gttgttgggg ttcatgtgta cggcaggatg
     3541 tgagtcctgt acgcctgagt tcttacccta tctcatcttg ctaggcacat ccttctgaaa
     3601 ctactgagat tccaactgtg aaatatctgt aatgagtagc tggtgggaaa aatttgatgc
     3661 cttttaaaga tgaaatttgg acaaagaaag tctttaatga ttagtatact gcgacatact
     3721 tagtaccaca tgtcgctact tattctgtag acctgcctgg tgtaccttat cgcctagaac
     3781 cttgaagccg ctttggaaga tgtatcacac gtgtttttag tcacttaagt cactagtgtt
     3841 taaaaaaaaa aaaaaggaat tccaccacac tggactagtg gatccgagct cggtaccaag
     3901 cttaagttta aaccgctgat cagcctcgac tgtgccttct agttgccagc catctgttgt
     3961 ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta
     4021 ataaaatgag gaaattgcat cgcattgtct gagtaggtgt cattctattc tggggggtgg
     4081 ggtggggcag gacagcaagg gggaggattg ggaagacaat agcaggcatg ctggggatgc
     4141 ggtgggctct atggcttctg aggcggaaag aaccagctgg ggctctaggg ggtatcccca
     4201 cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc
     4261 tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac
     4321 gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag
     4381 tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc
     4441 atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg
     4501 actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata
     4561 agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa
     4621 cgcgaattaa ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca
     4681 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc
     4741 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata
     4801 gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg
     4861 ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc tgcctctgag
     4921 ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagctcccg
     4981 ggagcttgta tatccatttt cggatctgat caagagacag gatgaggatc gtttcgcatg
     5041 attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc
     5101 tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg
     5161 caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcag
     5221 gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc
     5281 gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat
     5341 ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg
     5401 cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc
     5461 gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag
     5521 catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgcgcat gcccgacggc
     5581 gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc
     5641 cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata
     5701 gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc
     5761 gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac
     5821 gagttcttct gagcgggact ctggggttcg aaatgaccga ccaagcgacg cccaacctgc
     5881 catcacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt
     5941 tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc
     6001 accccaactt gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt
     6061 tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg
     6121 tatcttatca tgtctgtata ccgtcgacct ctagctagag cttggcgtaa tcatggtcat
     6181 agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa
     6241 gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc
     6301 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc
     6361 aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact
     6421 cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac
     6481 ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa
     6541 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
     6601 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
     6661 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     6721 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
     6781 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     6841 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     6901 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     6961 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa
     7021 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     7081 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
     7141 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
     7201 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
     7261 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
     7321 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     7381 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
     7441 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
     7501 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
     7561 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
     7621 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
     7681 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
     7741 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
     7801 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
     7861 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
     7921 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
     7981 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
     8041 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
     8101 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     8161 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
     8221 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     8281 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac gtc
//
