LOCUS       sequence-329324-        8687 bp    DNA     circular SYN 15-APR-2025
DEFINITION  expresses AMOT with inactivating mutation in third PPxY motif in 
            mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8687)
  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 8687)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 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: pcDNA3HA-AMOT p130∆PPxY3
FEATURES             Location/Qualifiers
     source          1..8687
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(48..67)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        239..618
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        619..822
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     773..793
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     867..886
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        867..885
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             908..934
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             938..964
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             2123..2143
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=7xHis
                     /translation="HHHHHHH"
     CDS             3673..3690
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     promoter        complement(4244..4262)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(4245..4262)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     polyA_signal    4288..4512
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4558..4986
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4645..4664)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4855..4876
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4995..5015)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5000..5329
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5180..5315
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5242..5261
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             5396..6190
                     /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(5450..5469)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6060..6079
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    6364..6485
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6401..6420)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6455..6474
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6534..6550)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6534..6550)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6547..6569)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6558..6574
                     /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(6582..6588,6589..6606,6607..6612))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6627..6648
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6765..6782)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6936..7524)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(7016..7035)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7695..8486,8487..8555))
                     /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     8318..8337
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8556..8660)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cgtcgacgga tcgggagatc tcccgatccc ctatggtgca ctctcagtac aatctgctct
       61 gatgccgcat agttaagcca gtatctgctc cctgcttgtg tgttggaggt cgctgagtag
      121 tgcgcgagca aaatttaagc tacaacaagg caaggcttga ccgacaattg catgaagaat
      181 ctgcttaggg ttaggcgttt tgcgctgctt cgcgatgtac gggccagata tacgcgttga
      241 cattgattat tgactagtta ttaatagtaa tcaattacgg ggtcattagt tcatagccca
      301 tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac
      361 gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact
      421 ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa
      481 gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg
      541 cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta
      601 gtcatcgcta ttaccatggt gatgcggttt tggcagtaca tcaatgggcg tggatagcgg
      661 tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag tttgttttgg
      721 caccaaaatc aacgggactt tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg
      781 ggcggtaggc gtgtacggtg ggaggtctat ataagcagag ctctctggct aactagagaa
      841 cccactgctt actggcttat cgaaattaat acgactcact atagggagac ccaagcttgc
      901 catggcctac ccatacgatg ttcctgacta tgcgggctat ccctatgacg tcccggacta
      961 tgcaggatcc atgagaaatt ctgacgaaca gccaagtgga gggaccacgg tattgcagcg
     1021 tttgctacaa gagcagcttc gctatggcaa tcctagtgag aatcgcagcc tgcttgccat
     1081 acaccagcaa gccacaggga atggccctcc tttccccagt ggcagtggga acccgggccc
     1141 tcagagtgat gtgttgagtc cccaagacca ccaccaacag cttgtggctc atgctgctcg
     1201 acaagaaccc caggggcagg aaatccagtc agaaaacctc atcatggaga agcagctgtc
     1261 tcctcgaatg caaaataatg aagaactccc gacctatgaa gaagccaagg tccagtccca
     1321 gtactttcgg ggccaacagc atgccagtgt tggagctgcc ttctatgtca ctggagtcac
     1381 caaccagaag atgaggactg agggacgccc atcagttcag cggctcaatc ctggaaagat
     1441 gcaccaagat gagggactca gagaccttaa gcaagggcat gtccgttcct tgagtgaacg
     1501 actaatgcag atgtcactgg ccaccagtgg agttaaggcc catccacctg ttaccagtgc
     1561 tcccctctcc ccaccacaac ccaatgacct ctacaagaat cccacaagtt ccagtgaatt
     1621 ctacaaggcc caagggccac ttcctaacca gcatagcctg aagggcatgg aacaccgagg
     1681 ccccccacca gaatatccct tcaagggcat gccaccccaa tctgtagtgt gcaagcccca
     1741 agagccaggg cacttctata gtgagcatcg cctgaaccag ccagggagaa cagaggggca
     1801 actgatgagg tatcagcatg ccgctgagta tggagcagcc aggccagcgc aggacatctc
     1861 attgccattg tcagccagga actcgcagcc tcacagccct acttcttctc tgacctctgg
     1921 ggggtccttg cccttgctac aatctccacc atccactaga ttgtctcctg cccgacaccc
     1981 cttggtccca aaccagggag accattcagc tcacctgcct aggccgcagc agcatttcct
     2041 tcctaatcag gctcaccagg gggatcatta ccgtctctcc caacctggcc tgagtcagca
     2101 gcagcagcaa cagcagcagc agcaccatca tcaccatcac caccaacaac agcagcagca
     2161 gcagccacag cagcagccag gagaagccta ttcagctatg cctcgggctc agccatcctc
     2221 tgcttcttat cagccagtgc cagcagaccc ttttgccatt gtttccagag cccagcagat
     2281 ggttgagatc ctctcagacg agaaccggaa cttgaggcaa gagttggaag gatgctatga
     2341 gaaggtggca agactgcaga aggtggagac agaaatccag cgcgtctcgg aggcatatga
     2401 gaacctcgtg aagtcatcct ccaaaagaga ggccctagag aaagccatga gaaacaagct
     2461 agagggcgag attcggagga tgcatgattt caacagggat ctgagagagc gtctagagac
     2521 tgccaacaag cagcttgcag agaaggaata tgaggggtca gaggacacca gaaaaaccat
     2581 ctcgcagctc tttgcaaaaa ataaagaaag ccagcgtgag aaggagaagc tggaagcgga
     2641 gctggccact gcccgttcta ccaatgagga ccaaagacga cacatcgaaa tccgagatca
     2701 ggccctgagt aatgcccagg ccaaggtggt aaagctggaa gaagagctga aaaagaagca
     2761 agtgtacgtt gacaaggtgg agaagatgca gcaggccctt gtacagctcc aggcagcatg
     2821 tgaaaaacgt gagcagctag agcaccgtct ccggacacga ctggagaggg aactggaatc
     2881 cctgagaatc cagcagcgtc agggcaactg tcagcccacc aacgtttcag aatacaatgc
     2941 tgctgcactg atggagctcc ttcgggagaa agaggagagg attctggctc tggaagctga
     3001 tatgacaaag tgggagcaga aatatttgga ggagaatgtg atgagacatt ttgctctgga
     3061 tgctgctgca actgtggctg ctcagaggga cacaacagtc atcagtcact ctcctaacac
     3121 cagctatgac acagctctag aagctcgcat ccagaaagag gaggaagaaa tcttgatggc
     3181 caataagcgt tgccttgaca tggagggcag gattaagacc ctccatgccc agattattga
     3241 gaaggatgcc atgatcaaag tactccagca gcgttcccgg aaggagccga gcaagacaga
     3301 gcagctgtcg tgcatgcggc cagcgaagtc tctgatgtcc atttccaatg ctggatcagg
     3361 cttgctctcc cactcatcca ccctgactgg ctcccccatc atggaagaaa agcgagacga
     3421 caagagctgg aaggggagcc taggcattct cctgggtgga gactaccgtg ctgaatatgt
     3481 cccttccaca ccctcgcctg tgccaccctc gactcccctg ctctcggctc actccaagac
     3541 agacagccga gactgcagta cccaaactga acgtgggacg gaatcgaaca aaactgcagc
     3601 tgttgctccc atctctgttc ctgctccagt tgctgctgcc gccactgctg ccgccatcac
     3661 tgccactgct gccaccatca ccaccaccat ggtagctgct gctccagttg ctgttgctgc
     3721 tgctgctgct ccagctgctg ctgctgcccc gtctccagcc actgccgctg ctactgctgc
     3781 tgctgtttct ccagctgctg ctggtcagat tccagctgct gcctctgttg cctcagctgc
     3841 tgccgttgct ccttctgctg ctgctgctgc tgctgttcag gttgctccag ctgctccggc
     3901 tccagttcca gctccggctc tggttccggt cccagctcca gcagcggctc aggcttctgc
     3961 tcctgctcag actcaggcac caacttcagc tccggctgtg gctccaactc cagctccaac
     4021 tccaactcca gctgtggctc aggctgaggt tcctgcaagt ccagctaccg gtcctggacc
     4081 acatcgtttg tctataccaa gtttgacctg caatccagac aaaacagatg ggcctgtgtt
     4141 ccactccaat actctggaaa gaaaaactcc cattcagatc ctgggacaag agcctgatgc
     4201 agagatggtg gaatatctca tctaactcga gggggggccc tattctatag tgtcacctaa
     4261 atgctagagc tcgctgatca gcctcgactg tgccttctag ttgccagcca tctgttgttt
     4321 gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc ctttcctaat
     4381 aaaatgagga aattgcatcg cattgtctga gtaggtgtca ttctattctg gggggtgggg
     4441 tggggcagga cagcaagggg gaggattggg aagacaatag caggcatgct ggggatgcgg
     4501 tgggctctat ggcttctgag gcggaaagaa ccagctgggg ctctaggggg tatccccacg
     4561 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
     4621 cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
     4681 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
     4741 ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat
     4801 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
     4861 tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag
     4921 ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     4981 cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc
     5041 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc
     5101 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt
     5161 cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc
     5221 ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctctg cctctgagct
     5281 attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa agctcccggg
     5341 agcttgtata tccattttcg gatctgatca agagacagga tgaggatcgt ttcgcatgat
     5401 tgaacaagat ggattgcacg caggttctcc ggccgcttgg gtggagaggc tattcggcta
     5461 tgactgggca caacagacaa tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca
     5521 ggggcgcccg gttctttttg tcaagaccga cctgtccggt gccctgaatg aactgcagga
     5581 cgaggcagcg cggctatcgt ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga
     5641 cgttgtcact gaagcgggaa gggactggct gctattgggc gaagtgccgg ggcaggatct
     5701 cctgtcatct caccttgctc ctgccgagaa agtatccatc atggctgatg caatgcggcg
     5761 gctgcatacg cttgatccgg ctacctgccc attcgaccac caagcgaaac atcgcatcga
     5821 gcgagcacgt actcggatgg aagccggtct tgtcgatcag gatgatctgg acgaagagca
     5881 tcaggggctc gcgccagccg aactgttcgc caggctcaag gcgcgcatgc ccgacggcga
     5941 ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat atcatggtgg aaaatggccg
     6001 cttttctgga ttcatcgact gtggccggct gggtgtggcg gaccgctatc aggacatagc
     6061 gttggctacc cgtgatattg ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt
     6121 gctttacggt atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga
     6181 gttcttctga gcgggactct ggggttcgaa atgaccgacc aagcgacgcc caacctgcca
     6241 tcacgagatt tcgattccac cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc
     6301 cgggacgccg gctggatgat cctccagcgc ggggatctca tgctggagtt cttcgcccac
     6361 cccaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc
     6421 acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta
     6481 tcttatcatg tctgtatacc gtcgacctct agctagagct tggcgtaatc atggtcatag
     6541 ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc
     6601 ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc
     6661 tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa
     6721 cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg
     6781 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
     6841 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     6901 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     6961 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     7021 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     7081 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     7141 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     7201 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     7261 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     7321 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
     7381 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     7441 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     7501 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     7561 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     7621 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     7681 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     7741 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     7801 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
     7861 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     7921 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     7981 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
     8041 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     8101 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     8161 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     8221 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     8281 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
     8341 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     8401 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     8461 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     8521 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     8581 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     8641 aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctga
//
