LOCUS       sequence-161701-        6782 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Residues 1-359 of ALIX; Bacterial Expression Vector; N-term His-tag;
            Sundquist Lab Internal ID: WISP11-428.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6782)
  AUTHORS   McCullough J, Fisher RD, Whitby FG, Sundquist WI, Hill CP
  TITLE     ALIX-CHMP4 interactions in the human ESCRT pathway.
  JOURNAL   Proc Natl Acad Sci U S A. 2008 Jun 3;105(22):7687-91. doi: 
            10.1073/pnas.0801567105. Epub 2008 May 29.
  PUBMED    18511562
REFERENCE   2  (bases 1 to 6782)
  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"; doi: 
            "10.1073/pnas.0801567105"; journalName: "Proc Natl Acad Sci U S A"; 
            date: "2008-06-3- 3"; volume: "105"; issue: "22"; pages: "7687-91"
COMMENT     Sequence Label: pet16b ALIX Bro1 (1-359)
FEATURES             Location/Qualifiers
     source          1..6782
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    1..25
                     /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)."
     RBS             40..62
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             75..104
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             111..131
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     primer_bind     complement(1265..1283)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1279..1326
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(1501..1506,1507..1523,1524..1529))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(1556..1574)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1642..1746
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1747..1815,1816..2607)
                     /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     complement(1965..1984)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2778..3366
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3267..3286
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3520..3537
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3552..3691
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3617..3636)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3777..3799
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3793..3984)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     6339..6358
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6378..6455)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6661..6680
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     join(6764..6782,1)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        6764..6782
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 ggaattgtga gcggataaca attcccctct agaaataatt ttgtttaact ttaagaagga
       61 gatataccat gggccatcat catcatcatc atcatcatca tcatagcggc gaaaacctgt
      121 atttccaggg ccatatggcg acattcatct cggtgcagct gaaaaagacc tcagaggtgg
      181 acctggccaa gccgctggtg aagttcatcc agcagactta cccaagcggc ggggaagagc
      241 aggcccagta ctgccgcgcg gcggaggagc tcagcaagct gcgccgcgcc gcagtcggtc
      301 gtccgctgga caagcacgag ggcgcgctcg agacgctcct gagatattat gatcagattt
      361 gttctattga acccaaattc ccattttctg aaaatcagat ctgcttgaca tttacctgga
      421 aggatgcttt cgataaaggt tcactttttg gaggctctgt aaaactggct cttgcaagct
      481 taggatatga aaagagctgt gtgttgttca attgtgcagc cttagctagc caaattgcag
      541 cagaacagaa cttagacaac gatgaaggat tgaaaatcgc tgctaaacat taccagtttg
      601 ctagtggtgc ctttttacat attaaagaga cggttttatc tgccttaagt cgagagccga
      661 ccgtggacat atctccagat actgttggga ccctcagtct tattatgctg gcacaggctc
      721 aagaagtatt ttttttaaaa gccacaagag ataaaatgaa agatgccatc atagctaaat
      781 tggctaatca ggctgcagat tattttggtg atgctttcaa acagtgtcaa tacaaagata
      841 ctctccccaa ggaggtgttc cctgtcttgg ctgcaaagca ctgtatcatg caggccaatg
      901 ctgagtacca tcagtctatc ctggcaaaac agcagaagaa atttggagaa gaaattgcaa
      961 ggttacagca tgcagcagaa ctgattaaaa cagtggcatc tcgctatgat gaatatgtta
     1021 atgtgaagga tttttctgac aaaatcaatc gtgcccttgc tgcagcaaag aaggataatg
     1081 acttcattta tcatgatcga gttccagacc ttaaagatct agatcctatt ggcaaagcca
     1141 cacttgtgaa atctaccccg gtcaatgtac ccatcagtca gaaatttact gatctgtttg
     1201 agaagatggt ttaaggatcc ggctgctaac aaagcccgaa aggaagctga gttggctgct
     1261 gccaccgctg agcaataact agcataaccc cttggggcct ctaaacgggt cttgaggggt
     1321 tttttgctga aaggaggaac tatatccgga tatcccgcaa gaggcccggc agtaccggca
     1381 taaccaagcc tatgcctaca gcatccaggg tgacggtgcc gaggatgacg atgagcgcat
     1441 tgttagattt catacacggt gcctgactgc gttagcaatt taactgtgat aaactaccgc
     1501 attaaagctt atcgatgata agctgtcaaa catgagaatt cttgaagacg aaagggcctc
     1561 gtgatacgcc tatttttata ggttaatgtc atgataataa tggtttctta gacgtcaggt
     1621 ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca
     1681 aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg
     1741 aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc
     1801 cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg
     1861 ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt
     1921 cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta
     1981 ttatcccgtg ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat
     2041 gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga
     2101 gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca
     2161 acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact
     2221 cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc
     2281 acgatgcctg cagcaatggc aacaacgttg cgcaaactat taactggcga actacttact
     2341 ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt
     2401 ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt
     2461 gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt
     2521 atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata
     2581 ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag
     2641 attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat
     2701 ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa
     2761 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
     2821 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
     2881 ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgtccttct agtgtagccg
     2941 tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc
     3001 ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga
     3061 cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc
     3121 agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc
     3181 gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca
     3241 ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg
     3301 tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta
     3361 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct
     3421 cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag
     3481 tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa
     3541 gcggaagagc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc
     3601 aatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag tatacactcc
     3661 gctatcgcta cgtgactggg tcatggctgc gccccgacac ccgccaacac ccgctgacgc
     3721 gccctgacgg gcttgtctgc tcccggcatc cgcttacaga caagctgtga ccgtctccgg
     3781 gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgaggc agctgcggta
     3841 aagctcatca gcgtggtcgt gaagcgattc acagatgtct gcctgttcat ccgcgtccag
     3901 ctcgttgagt ttctccagaa gcgttaatgt ctggcttctg ataaagcggg ccatgttaag
     3961 ggcggttttt tcctgtttgg tcactgatgc ctccgtgtaa gggggatttc tgttcatggg
     4021 ggtaatgata ccgatgaaac gagagaggat gctcacgata cgggttactg atgatgaaca
     4081 tgcccggtta ctggaacgtt gtgagggtaa acaactggcg gtatggatgc ggcgggacca
     4141 gagaaaaatc actcagggtc aatgccagcg cttcgttaat acagatgtag gtgttccaca
     4201 gggtagccag cagcatcctg cgatgcagat ccggaacata atggtgcagg gcgctgactt
     4261 ccgcgtttcc agactttacg aaacacggaa accgaagacc attcatgttg ttgctcaggt
     4321 cgcagacgtt ttgcagcagc agtcgcttca cgttcgctcg cgtatcggtg attcattctg
     4381 ctaaccagta aggcaacccc gccagcctag ccgggtcctc aacgacagga gcacgatcat
     4441 gcgcacccgt ggccaggacc caacgctgcc cgagatgcgc cgcgtgcggc tgctggagat
     4501 ggcggacgcg atggatatgt tctgccaagg gttggtttgc gcattcacag ttctccgcaa
     4561 gaattgattg gctccaattc ttggagtggt gaatccgtta gcgaggtgcc gccggcttcc
     4621 attcaggtcg aggtggcccg gctccatgca ccgcgacgca acgcggggag gcagacaagg
     4681 tatagggcgg cgcctacaat ccatgccaac ccgttccatg tgctcgccga ggcggcataa
     4741 atcgccgtga cgatcagcgg tccaatgatc gaagttaggc tggtaagagc cgcgagcgat
     4801 ccttgaagct gtccctgatg gtcgtcatct acctgcctgg acagcatggc ctgcaacgcg
     4861 ggcatcccga tgccgccgga agcgagaaga atcataatgg ggaaggccat ccagcctcgc
     4921 gtcgcgaacg ccagcaagac gtagcccagc gcgtcggccg ccatgccggc gataatggcc
     4981 tgcttctcgc cgaaacgttt ggtggcggga ccagtgacga aggcttgagc gagggcgtgc
     5041 aagattccga ataccgcaag cgacaggccg atcatcgtcg cgctccagcg aaagcggtcc
     5101 tcgccgaaaa tgacccagag cgctgccggc acctgtccta cgagttgcat gataaagaag
     5161 acagtcataa gtgcggcgac gatagtcatg ccccgcgccc accggaagga gctgactggg
     5221 ttgaaggctc tcaagggcat cggtcgagat cccggtgcct aatgagtgag ctaacttaca
     5281 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa actgtcgtgc cagctgcatt
     5341 aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgccag ggtggttttt
     5401 cttttcacca gtgagacggg caacagctga ttgcccttca ccgcctggcc ctgagagagt
     5461 tgcagcaagc ggtccacgct ggtttgcccc agcaggcgaa aatcctgttt gatggtggtt
     5521 aacggcggga tataacatga gctgtcttcg gtatcgtcgt atcccactac cgagatatcc
     5581 gcaccaacgc gcagcccgga ctcggtaatg gcgcgcattg cgcccagcgc catctgatcg
     5641 ttggcaacca gcatcgcagt gggaacgatg ccctcattca gcatttgcat ggtttgttga
     5701 aaaccggaca tggcactcca gtcgccttcc cgttccgcta tcggctgaat ttgattgcga
     5761 gtgagatatt tatgccagcc agccagacgc agacgcgccg agacagaact taatgggccc
     5821 gctaacagcg cgatttgctg gtgacccaat gcgaccagat gctccacgcc cagtcgcgta
     5881 ccgtcttcat gggagaaaat aatactgttg atgggtgtct ggtcagagac atcaagaaat
     5941 aacgccggaa cattagtgca ggcagcttcc acagcaatgg catcctggtc atccagcgga
     6001 tagttaatga tcagcccact gacgcgttgc gcgagaagat tgtgcaccgc cgctttacag
     6061 gcttcgacgc cgcttcgttc taccatcgac accaccacgc tggcacccag ttgatcggcg
     6121 cgagatttaa tcgccgcgac aatttgcgac ggcgcgtgca gggccagact ggaggtggca
     6181 acgccaatca gcaacgactg tttgcccgcc agttgttgtg ccacgcggtt gggaatgtaa
     6241 ttcagctccg ccatcgccgc ttccactttt tcccgcgttt tcgcagaaac gtggctggcc
     6301 tggttcacca cgcgggaaac ggtctgataa gagacaccgg catactctgc gacatcgtat
     6361 aacgttactg gtttcacatt caccaccctg aattgactct cttccgggcg ctatcatgcc
     6421 ataccgcgaa aggttttgcg ccattcgatg gtgtccggga tctcgacgct ctcccttatg
     6481 cgactcctgc attaggaagc agcccagtag taggttgagg ccgttgagca ccgccgccgc
     6541 aaggaatggt gcatgcaagg agatggcgcc caacagtccc ccggccacgg ggcctgccac
     6601 catacccacg ccgaaacaag cgctcatgag cccgaagtgg cgagcccgat cttccccatc
     6661 ggtgatgtcg gcgatatagg cgccagcaac cgcacctgtg gcgccggtga tgccggccac
     6721 gatgcgtccg gcgtagagga tcgagatctc gatcccgcga aattaatacg actcactata
     6781 gg
//
