LOCUS       sequence-372337-        7193 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express his-tagged NEDD4L W2-W3-W4-HECT (381) in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7193)
  AUTHORS   Wright KM, Nathan S, Jiang H, Xia W, Kim H, Chakouri N, Nwafor JN, 
            Fossier L, Srinivasan L, Chen Z, Boronina T, Post J, Paul S, Cole 
            RN, Ben-Johny M, Cole PA, Gabelli SB
  TITLE     NEDD4L intramolecular interactions regulate its auto and substrate 
            Na(V)1.5 ubiquitination.
  JOURNAL   J Biol Chem. 2024 Mar;300(3):105715. doi: 10.1016/j.jbc.2024.105715.
            Epub 2024 Feb 2.
  PUBMED    38309503
REFERENCE   2  (bases 1 to 7193)
  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."; date: "2024-03"; volume: "300(3):105715. doi"; pages: " 
            10.1016/j.jbc.2024.105715. Epub 2024 Feb 2"
COMMENT     Sequence Label: His-Precission-NEDD4L (381-975)_pcDNA3.1(+)
FEATURES             Location/Qualifiers
     source          1..7193
                     /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             907..924
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             928..951
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     primer_bind     complement(2784..2801)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2790..3014
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3060..3488
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3147..3166)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3357..3378
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3497..3517)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3502..3831
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3682..3817
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3744..3763
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3898..4692
                     /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(3952..3971)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4562..4581
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4866..4987
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4903..4922)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4957..4976
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5036..5052)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5036..5052)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5049..5071)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5060..5076
                     /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(5084..5090,5091..5108,5109..5114))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5129..5150
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5267..5284)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5438..6026)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5518..5537)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6197..6988,6989..7057))
                     /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     6820..6839
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7058..7162)
                     /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 atggggcacc accaccacca ccacggcctg gaggtgctgt tccagggccc agtacatacc
      961 acgccgggtc tgccttcagg ctgggaagaa agaaaagatg ctaaggggcg cacatactat
     1021 gtcaatcata acaatcgaac cacaacttgg actcgaccta tcatgcagct tgcagaagat
     1081 ggtgcgtccg gatcagccac aaacagtaac aaccatctaa tcgagcctca gatccgccgg
     1141 cctcgtagcc tcagctcgcc aacagtaact ttatctgccc cgctggaggg tgccaaggac
     1201 tcacccgtac gtcgggctgt gaaagacacc ctttccaacc cacagtcccc acagccatca
     1261 ccttacaact cccccaaacc acaacacaaa gtcacacaga gcttcttgcc acccggctgg
     1321 gaaatgagga tagcgccaaa cggccggccc ttcttcattg atcataacac aaagactaca
     1381 acctgggaag atccacgttt gaaatttcca gtacatatgc ggtcaaagac atctttaaac
     1441 cccaatgacc ttggccccct tcctcctggc tgggaagaaa gaattcactt ggatggccga
     1501 acgttttata ttgatcataa tagcaaaatt actcagtggg aagacccaag actgcagaac
     1561 ccagctatta ctggtccggc tgtcccttac tccagagaat ttaagcagaa atatgactac
     1621 ttcaggaaga aattaaagaa acctgctgat atccccaata ggtttgaaat gaaacttcac
     1681 agaaataaca tatttgaaga gtcctatcgg agaattatgt ccgtgaaaag accagatgtc
     1741 ctaaaagcta gactgtggat tgagtttgaa tcagagaaag gtcttgacta tgggggtgtg
     1801 gccagagaat ggttcttctt actgtccaaa gagatgttca acccctacta cggcctcttt
     1861 gagtactctg ccacggacaa ctacaccctt cagatcaacc ctaattcagg cctctgtaat
     1921 gaggatcatt tgtcctactt cacttttatt ggaagagttg ctggtctggc cgtatttcat
     1981 gggaagctct tagatggttt cttcattaga ccattttaca agatgatgtt gggaaagcag
     2041 ataaccctga atgacatgga atctgtggat agtgaatatt acaactcttt gaaatggatc
     2101 ctggagaatg accctactga gctggacctc atgttctgca tagacgaaga aaactttgga
     2161 cagacatatc aagtggattt gaagcccaat gggtcagaaa taatggtcac aaatgaaaac
     2221 aaaagggaat atatcgactt agtcatccag tggagatttg tgaacagggt ccagaagcag
     2281 atgaacgcct tcttggaggg attcacagaa ctacttccta ttgatttgat taaaattttt
     2341 gatgaaaatg agctggagtt gctcatgtgc ggcctcggtg atgtggatgt gaatgactgg
     2401 agacagcatt ctatttacaa gaacggctac tgcccaaacc accccgtcat tcagtggttc
     2461 tggaaggctg tgctactcat ggacgccgaa aagcgtatcc ggttactgca gtttgtcaca
     2521 gggacatcgc gagtacctat gaatggattt gccgaacttt atggttccaa tggtcctcag
     2581 ctatttacaa tagagcaatg gggcagtcct gagaaactgc ccagagctca cacatgcttt
     2641 aatcgccttg acttacctcc atatgaaacc tttgaagatt tacgagagaa acttctcatg
     2701 gccgtggaaa atgctcaagg atttgaaggg gtggattaag cggccgctcg agtctagagg
     2761 gcccgtttaa acccgctgat cagcctcgac tgtgccttct agttgccagc catctgttgt
     2821 ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta
     2881 ataaaatgag gaaattgcat cgcattgtct gagtaggtgt cattctattc tggggggtgg
     2941 ggtggggcag gacagcaagg gggaggattg ggaagacaat agcaggcatg ctggggatgc
     3001 ggtgggctct atggcttctg aggcggaaag aaccagctgg ggctctaggg ggtatcccca
     3061 cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc
     3121 tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac
     3181 gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag
     3241 tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc
     3301 atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg
     3361 actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata
     3421 agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa
     3481 cgcgaattaa ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca
     3541 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc
     3601 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata
     3661 gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg
     3721 ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc tgcctctgag
     3781 ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagctcccg
     3841 ggagcttgta tatccatttt cggatctgat caagagacag gatgaggatc gtttcgcatg
     3901 attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc
     3961 tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg
     4021 caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcag
     4081 gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc
     4141 gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat
     4201 ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg
     4261 cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc
     4321 gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag
     4381 catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgcgcat gcccgacggc
     4441 gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc
     4501 cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata
     4561 gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc
     4621 gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac
     4681 gagttcttct gagcgggact ctggggttcg aaatgaccga ccaagcgacg cccaacctgc
     4741 catcacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt
     4801 tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc
     4861 accccaactt gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt
     4921 tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg
     4981 tatcttatca tgtctgtata ccgtcgacct ctagctagag cttggcgtaa tcatggtcat
     5041 agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa
     5101 gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc
     5161 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc
     5221 aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact
     5281 cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac
     5341 ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa
     5401 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
     5461 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
     5521 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     5581 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
     5641 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     5701 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     5761 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     5821 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa
     5881 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     5941 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
     6001 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
     6061 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
     6121 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
     6181 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     6241 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
     6301 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
     6361 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
     6421 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
     6481 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
     6541 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
     6601 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
     6661 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
     6721 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
     6781 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
     6841 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
     6901 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
     6961 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     7021 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
     7081 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     7141 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac gtc
//
