LOCUS       sequence-328857-        5092 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  expresses NEDD4L WW2 in bacterial cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5092)
  AUTHORS   Rheinemann L, Thompson T, Mercenne G, Paine EL, Peterson FC, Volkman
            BF, Alam SL, Alian A, Sundquist WI
  TITLE     Interactions between AMOT PPxY Motifs and NEDD4L WW Domains Function
            in HIV-1 Release.
  JOURNAL   J Biol Chem. 2021 Jul 17:100975. doi: 10.1016/j.jbc.2021.100975.
  PUBMED    34284061
REFERENCE   2  (bases 1 to 5092)
  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. 2021 Jul 17:100975. doi: 10.1016/j.jbc.2021.100975."
COMMENT     Sequence Label: pGEX-PP-NEDD4L-WW2(385-418)
FEATURES             Location/Qualifiers
     source          1..5092
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     966..985
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(1005..1082)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     complement(1155..1172)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1326..1914)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1406..1425)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2085..2876,2877..2945))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     2708..2727
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2946..3050)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3118..3136
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     3281..3303
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3483..3506)
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             complement(3510..3527)
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             complement(3534..4187)
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     complement(3554..3576)
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     protein_bind    4210..4226
                     /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(4234..4240,4241..4256,4257..4262))
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     CDS             complement(4364..4891)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     4840..4862
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4854..4871
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4855..4871
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4887..4903)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4887..4903)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     protein_bind    4911..4927
                     /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(4935..4941,4942..4959,4960..4965))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     CDS             complement(join(5014..5092,1..1004))
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
ORIGIN
        1 tgcgtattgg gcgccagggt ggtttttctt ttcaccagtg agacgggcaa cagctgattg
       61 cccttcaccg cctggccctg agagagttgc agcaagcggt ccacgctggt ttgccccagc
      121 aggcgaaaat cctgtttgat ggtggttaac ggcgggatat aacatgagct gtcttcggta
      181 tcgtcgtatc ccactaccga gatatccgca ccaacgcgca gcccggactc ggtaatggcg
      241 cgcattgcgc ccagcgccat ctgatcgttg gcaaccagca tcgcagtggg aacgatgccc
      301 tcattcagca tttgcatggt ttgttgaaaa ccggacatgg cactccagtc gccttcccgt
      361 tccgctatcg gctgaatttg attgcgagtg agatatttat gccagccagc cagacgcaga
      421 cgcgccgaga cagaacttaa tgggcccgct aacagcgcga tttgctggtg acccaatgcg
      481 accagatgct ccacgcccag tcgcgtaccg tcttcatggg agaaaataat actgttgatg
      541 ggtgtctggt cagagacatc aagaaataac gccggaacat tagtgcaggc agcttccaca
      601 gcaatggcat cctggtcatc cagcggatag ttaatgatca gcccactgac gcgttgcgcg
      661 agaagattgt gcaccgccgc tttacaggct tcgacgccgc ttcgttctac catcgacacc
      721 accacgctgg cacccagttg atcggcgcga gatttaatcg ccgcgacaat ttgcgacggc
      781 gcgtgcaggg ccagactgga ggtggcaacg ccaatcagca acgactgttt gcccgccagt
      841 tgttgtgcca cgcggttggg aatgtaattc agctccgcca tcgccgcttc cactttttcc
      901 cgcgttttcg cagaaacgtg gctggcctgg ttcaccacgc gggaaacggt ctgataagag
      961 acaccggcat actctgcgac atcgtataac gttactggtt tcacattcac caccctgaat
     1021 tgactctctt ccgggcgcta tcatgccata ccgcgaaagg ttttgcgcca ttcgatggtg
     1081 tcggaattta tgcggtgtga aataccgcac agatgcgtaa ggagaaaata ccgcatcagg
     1141 cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg
     1201 gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga
     1261 aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
     1321 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
     1381 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
     1441 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
     1501 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
     1561 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
     1621 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
     1681 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
     1741 tggcctaact acggctacac tagaaggaca gtatttggta tctgcgctct gctgaagcca
     1801 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     1861 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     1921 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     1981 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     2041 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
     2101 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     2161 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     2221 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     2281 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     2341 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     2401 gcaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     2461 cgatcaaggc gagttacatg atctcccatg ttgtgcaaaa aagcggttag ctccttcggt
     2521 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     2581 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     2641 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     2701 acacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     2761 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     2821 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     2881 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     2941 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     3001 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc
     3061 cgaaaagtgc cacctgacgt ctaagaaacc attattatca tgacattaac ctataaaaat
     3121 aggcgtatca cgaggccctt tcgtcttcaa gaattataca ctccgctatc gctacgtgac
     3181 tgggtcatgg ctgcgccccg acacccgcca acacccgctg acgcgccctg acgggcttgt
     3241 ctgctcccgg catccgctta cagacaagct gtgaccgtct ccgggagctg catgtgtcag
     3301 aggttttcac cgtcatcacc gaaacgcgcg aggcagatcg tcagtcagtc acgatgaatt
     3361 ctcgaggatc cttacatgat aggtcgagtc caagttgtgg ttcgattgtt atgattgaca
     3421 tagtatgtgc gccccttagc atcttttctt tcttcccagc ctgaaggcag acccggcata
     3481 tggggtccct gaaagaggac ttcaagcgga gatccacgcg gaaccagatc cgattttgga
     3541 ggatggtcgc caccaccaaa cgtggcttgc cagccctgca aaggccatgc tatatacttg
     3601 ctggatttca agtacttatc aatttgtggg atagcttcaa tacgtttttt aaaacaaact
     3661 aattttggga acgcatccag gcacattggg tccatgtata aaacaacatc aagagcgtca
     3721 tacaacatga agtcaggatg ggttacatga tcaccattta aatatgtttt atgacataaa
     3781 cgatcttcga acattttcag catttcaggt agcttgctaa gaaaatcaac tttgagagtt
     3841 tcaaagtctt tactatatgc aattctcgaa acaccgtatc taatatccaa aaccgctcct
     3901 tcaagcattg aaatctctgc acgctctttt ggacaaccac ccaacatgtt gtgcttgtca
     3961 gctatataac gtatgatggc catagactgt gttaatttaa catcaccatc aatataataa
     4021 ggaagattgg gaaactccaa acccaattca aactttttgt ttcgccattt atcaccttca
     4081 tcgcgctcat acaaatgctc ttcatatttt tcttcaagat attccaaaag aagtcgagtg
     4141 ggttgcacaa ggcccttaat tttccaataa cctagtatag gggacatgaa tactgtttcc
     4201 tgtgtgaaat tgttatccgc tcacaattcc acacattata cgagccgatg attaattgtc
     4261 aacagctcat ttcagaatat ttgccagaac cgttatgatg tcggcgcaaa aaacattatc
     4321 cagaacggga gtgcgccttg agcgacacga attatgcagt gatttacgac ctgcacagcc
     4381 ataccacagc ttccgatggc tgcctgacgc cagaagcatt ggtgcaccgt gcagtcgata
     4441 agctaattcc aacgccatca aaaataattc gcgtctggcc ttcctgtagc cagctttcat
     4501 caacattaaa tgtgagcgag taacaacccg tcggattctc cgtgggaaca aacggcggat
     4561 tgaccgtaat gggataggtc acgttggtgt agatgggcgc atcgtaaccg tgcatctgcc
     4621 agtttgaggg gacgacgaca gtatcggcct caggaagatc gcactccagc cagctttccg
     4681 gcaccgcttc tggtgccgga aaccaggcaa agcgccattc gccattcagg ctgcgcaact
     4741 gttgggaagg gcgatcggtg cgggcctctt cgctattacg ccagctggcg aaagggggat
     4801 gtgctgcaag gcgattaagt tgggtaacgc cagggttttc ccagtcacga cgttgtaaaa
     4861 cgacggccag tgaatccgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg
     4921 ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa
     4981 tgagtgagct aacttacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac
     5041 ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tt
//
