LOCUS       sequence-372356-        7341 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express GST-tagged NEDD4L W2-W3-W4-HECT (349) in insect cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7341)
  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 7341)
  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: pFastBac1-NEDD4L_W2-W3-W4-HECT (349)
FEATURES             Location/Qualifiers
     source          1..7341
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      2..456
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(89..108)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     298..319
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        482..586
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(587..655,656..1447)
                     /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(805..824)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1618..2206
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2107..2126
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2360..2377
                     /label=L4440
                     /note="L4440 vector, forward primer"
     mobile_element  2511..2735
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7R
                     /note="mini-Tn7 element (right end of the Tn7 transposon)"
     CDS             complement(2802..3335)
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(join(3524..3529,3530..3546,3547..3552))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     promoter        3904..3995
                     /gene="polh from Autographa californica"
                     /label=polyhedrin promoter
                     /note="promoter for the baculovirus polyhedrin gene"
     primer_bind     3920..3937
                     /label=Polyhedrin-F
                     /note="Polyhedrin promoter, forward primer"
     CDS             4065..4718
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     4676..4698
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             4725..4748
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     polyA_signal    6831..6965
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6868..6887)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6922..6941
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     mobile_element  6994..7159
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7L
                     /note="mini-Tn7 element (left end of the Tn7 transposon)"
ORIGIN
        1 gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc
       61 gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc
      121 acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt
      181 agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg
      241 ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt cttaatagtg
      301 gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct tttgatttat
      361 aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta
      421 acgcgaattt taacaaaata ttaacgctta caatttaggt ggcacttttc ggggaaatgt
      481 gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag
      541 acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga gtattcaaca
      601 tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt ttgctcaccc
      661 agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag tgggttacat
      721 cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag aacgttttcc
      781 aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta ttgacgccgg
      841 gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtactcacc
      901 agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca gtgctgccat
      961 aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag gaccgaagga
     1021 gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc
     1081 ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg tagcaatggc
     1141 aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc ggcaacaatt
     1201 aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc
     1261 tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg gtatcattgc
     1321 agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga cggggagtca
     1381 ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca
     1441 ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa aacttcattt
     1501 ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta
     1561 acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg
     1621 agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc
     1681 ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag
     1741 cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa
     1801 gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc
     1861 cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc
     1921 gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta
     1981 caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag
     2041 aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct
     2101 tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga
     2161 gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc
     2221 ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt
     2281 atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata ccgctcgccg
     2341 cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc gcctgatgcg
     2401 gtattttctc cttacgcatc tgtgcggtat ttcacaccgc atagaccagc cgcgtaacct
     2461 ggcaaaatcg gttacggttg agtaataaat ggatgccctg cgtaagcggg tgtgggcgga
     2521 caataaagtc ttaaactgaa caaaatagat ctaaactatg acaataaagt cttaaactag
     2581 acagaatagt tgtaaactga aatcagtcca gttatgctgt gaaaaagcat actggacttt
     2641 tgttatggct aaagcaaact cttcattttc tgaagtgcaa attgcccgtc gtattaaaga
     2701 ggggcgtggc caagggcatg gtaaagacta tattcgcggc gttgtgacaa tttaccgaac
     2761 aactccgcgg ccgggaagcc gatctcggct tgaacgaatt gttaggtggc ggtacttggg
     2821 tcgatatcaa agtgcatcac ttcttcccgt atgcccaact ttgtatagag agccactgcg
     2881 ggatcgtcac cgtaatctgc ttgcacgtag atcacataag caccaagcgc gttggcctca
     2941 tgcttgagga gattgatgag cgcggtggca atgccctgcc tccggtgctc gccggagact
     3001 gcgagatcat agatatagat ctcactacgc ggctgctcaa acttgggcag aacgtaagcc
     3061 gcgagagcgc caacaaccgc ttcttggtcg aaggcagcaa gcgcgatgaa tgtcttacta
     3121 cggagcaagt tcccgaggta atcggagtcc ggctgatgtt gggagtaggt ggctacgtct
     3181 ccgaactcac gaccgaaaag atcaagagca gcccgcatgg atttgacttg gtcagggccg
     3241 agcctacatg tgcgaatgat gcccatactt gagccaccta actttgtttt agggcgactg
     3301 ccctgctgcg taacatcgtt gctgctgcgt aacatcgttg ctgctccata acatcaaaca
     3361 tcgacccacg gcgtaacgcg cttgctgctt ggatgcccga ggcatagact gtacaaaaaa
     3421 acagtcataa caagccatga aaaccgccac tgcgccgtta ccaccgctgc gttcggtcaa
     3481 ggttctggac cagttgcgtg agcgcatacg ctacttgcat tacagtttac gaaccgaaca
     3541 ggcttatgtc aactgggttc gtgccttcat ccgtttccac ggtgtgcgtc acccggcaac
     3601 cttgggcagc agcgaagtcg aggcatttct gtcctggctg gcgaacgagc gcaaggtttc
     3661 ggtctccacg catcgtcagg cattggcggc cttgctgttc ttctacggca aggtgctgtg
     3721 cacggatctg ccctggcttc aggagatcgg aagacctcgg ccgtcgcggc gcttgccggt
     3781 ggtgctgacc ccggatgaag tggttcgcat cctcggtttt ctggaaggcg agcatcgttt
     3841 gttcgcccag gactctagct atagttctag tggttggcta cgtatactcc ggaatattaa
     3901 tagatcatgg agataattaa aatgataacc atctcgcaaa taaataagta ttttactgtt
     3961 ttcgtaacag ttttgtaata aaaaaaccta taaatattcc ggattattca taccgtccca
     4021 ccatcgggcg cggatcccgg tccgaagcgc gcggaattca aaggatgtcc cctatactag
     4081 gttattggaa aattaagggc cttgtgcaac ccactcgact tcttttggaa tatcttgaag
     4141 aaaaatatga agagcatttg tatgagcgcg atgaaggtga taaatggcga aacaaaaagt
     4201 ttgaattggg tttggagttt cccaatcttc cttattatat tgatggtgat gttaaattaa
     4261 cacagtctat ggccatcata cgttatatag ctgacaagca caacatgttg ggtggttgtc
     4321 caaaagagcg tgcagagatt tcaatgcttg aaggagcggt tttggatatt agatacggtg
     4381 tttcgagaat tgcatatagt aaagactttg aaactctcaa agttgatttt cttagcaagc
     4441 tacctgaaat gctgaaaatg ttcgaagatc gtttatgtca taaaacatat ttaaatggtg
     4501 atcatgtaac ccatcctgac ttcatgttgt atgacgctct tgatgttgtt ttatacatgg
     4561 acccaatgtg cctggatgcg ttcccaaaat tagtttgttt taaaaaacgt attgaagcta
     4621 tcccacaaat tgataagtac ttgaaatcca gcaagtatat agcatggcct ttgcagggct
     4681 ggcaagccac gtttggtggt ggcgaccatc ctccaaaatc ggatctggaa gttctgttcc
     4741 aggggcccct gggagtacgc gccggctccg aacagggcca tctaccaccg cccagtgccc
     4801 cagctgggag agcgcgttca tcaactgtca cgggtggtga ggaaccaacg ccatcagtgg
     4861 cctatgtaca tactacgccg ggtctgcctt caggctggga agaaagaaaa gatgctaagg
     4921 ggcgcacata ctatgtcaat cataacaatc gaaccacaac ttggactcga cctatcatgc
     4981 agcttgcaga agatggtgcg tccggatcag ccacaaacag taacaaccat ctaatcgagc
     5041 ctcagatccg ccggcctcgt agcctcagct cgccaacagt aactttatct gccccgctgg
     5101 agggtgccaa ggactcaccc gtacgtcggg ctgtgaaaga caccctttcc aacccacagt
     5161 ccccacagcc atcaccttac aactccccca aaccacaaca caaagtcaca cagagcttct
     5221 tgccacccgg ctgggaaatg aggatagcgc caaacggccg gcccttcttc attgatcata
     5281 acacaaagac tacaacctgg gaagatccac gtttgaaatt tccagtacat atgcggtcaa
     5341 agacatcttt aaaccccaat gaccttggcc cccttcctcc tggctgggaa gaaagaattc
     5401 acttggatgg ccgaacgttt tatattgatc ataatagcaa aattactcag tgggaagacc
     5461 caagactgca gaacccagct attactggtc cggctgtccc ttactccaga gaatttaagc
     5521 agaaatatga ctacttcagg aagaaattaa agaaacctgc tgatatcccc aataggtttg
     5581 aaatgaaact tcacagaaat aacatatttg aagagtccta tcggagaatt atgtccgtga
     5641 aaagaccaga tgtcctaaaa gctagactgt ggattgagtt tgaatcagag aaaggtcttg
     5701 actatggggg tgtggccaga gaatggttct tcttactgtc caaagagatg ttcaacccct
     5761 actacggcct ctttgagtac tctgccacgg acaactacac ccttcagatc aaccctaatt
     5821 caggcctctg taatgaggat catttgtcct acttcacttt tattggaaga gttgctggtc
     5881 tggccgtatt tcatgggaag ctcttagatg gtttcttcat tagaccattt tacaagatga
     5941 tgttgggaaa gcagataacc ctgaatgaca tggaatctgt ggatagtgaa tattacaact
     6001 ctttgaaatg gatcctggag aatgacccta ctgagctgga cctcatgttc tgcatagacg
     6061 aagaaaactt tggacagaca tatcaagtgg atttgaagcc caatgggtca gaaataatgg
     6121 tcacaaatga aaacaaaagg gaatatatcg acttagtcat ccagtggaga tttgtgaaca
     6181 gggtccagaa gcagatgaac gccttcttgg agggattcac agaactactt cctattgatt
     6241 tgattaaaat ttttgatgaa aatgagctgg agttgctcat gtgcggcctc ggtgatgtgg
     6301 atgtgaatga ctggagacag cattctattt acaagaacgg ctactgccca aaccaccccg
     6361 tcattcagtg gttctggaag gctgtgctac tcatggacgc cgaaaagcgt atccggttac
     6421 tgcagtttgt cacagggaca tcgcgagtac ctatgaatgg atttgccgaa ctttatggtt
     6481 ccaatggtcc tcagctattt acaatagagc aatggggcag tcctgagaaa ctgcccagag
     6541 ctcacacatg ctttaatcgc cttgacttac ctccatatga aacctttgaa gatttacgag
     6601 agaaacttct catggccgtg gaaaatgctc aaggatttga aggggtggat taagcggccg
     6661 ctttcgaatc tagagcctgc agtctcgagg catgcggtac caagcttgtc gagaagtact
     6721 agaggatcat aatcagccat accacatttg tagaggtttt acttgcttta aaaaacctcc
     6781 cacacctccc cctgaacctg aaacataaaa tgaatgcaat tgttgttgtt aacttgttta
     6841 ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat
     6901 ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct
     6961 ggatctgatc actgcttgag cctaggagat ccgaaccaga taagtgaaat ctagttccaa
     7021 actattttgt catttttaat tttcgtatta gcttacgacg ctacacccag ttcccatcta
     7081 ttttgtcact cttccctaaa taatccttaa aaactccatt tccacccctc ccagttccca
     7141 actattttgt ccgcccacag cggggcattt ttcttcctgt tatgttttta atcaaacatc
     7201 ctgccaactc catgtgacaa accgtcatct tcggctactt tttctctgtc acagaatgaa
     7261 aatttttctg tcatctcttc gttattaatg tttgtaattg actgaatatc aacgcttatt
     7321 tgcagcctga atggcgaatg g
//
