LOCUS       sequence-159358-        5057 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Residues 205-224 of CHMP4B, Bacterial Expression Vector; 
            TEV-cleavabel GST tag; Sundquist Lab Internal ID: WISP06-201.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5057)
  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 5057)
  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: GST-CHMP4B (205-224) (pGEX2T(TEV))
FEATURES             Location/Qualifiers
     source          1..5057
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(183..188,189..204,205..211)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    219..235
                     /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)."
     CDS             258..911
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     869..891
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             918..935
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             939..959
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     primer_bind     complement(1107..1129)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1274..1292)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1360..1464
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1465..1533,1534..2325)
                     /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(1683..1702)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2496..3084
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2985..3004
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3238..3255
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        3328..3405
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     CDS             3406..4488
                     /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"
     primer_bind     complement(3425..3444)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     protein_bind    4501..4522
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4537..4542,4543..4560,4561..4567)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4575..4591
                     /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)."
     primer_bind     4599..4615
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4599..4615
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(4611..5057,1..81)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGITLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(4631..4648)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4631..4647)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4640..4662)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 acgttatcga ctgcacggtg caccaatgct tctggcgtca ggcagccatc ggaagctgtg
       61 gtatggctgt gcaggtcgta aatcactgca taattcgtgt cgctcaaggc gcactcccgt
      121 tctggataat gttttttgcg ccgacatcat aacggttctg gcaaatattc tgaaatgagc
      181 tgttgacaat taatcatcgg ctcgtataat gtgtggaatt gtgagcggat aacaatttca
      241 cacaggaaac agtattcatg tcccctatac taggttattg gaaaattaag ggccttgtgc
      301 aacccactcg acttcttttg gaatatcttg aagaaaaata tgaagagcat ttgtatgagc
      361 gcgatgaagg tgataaatgg cgaaacaaaa agtttgaatt gggtttggag tttcccaatc
      421 ttccttatta tattgatggt gatgttaaat taacacagtc tatggccatc atacgttata
      481 tagctgacaa gcacaacatg ttgggtggtt gtccaaaaga gcgtgcagag atttcaatgc
      541 ttgaaggagc ggttttggat attagatacg gtgtttcgag aattgcatat agtaaagact
      601 ttgaaactct caaagttgat tttcttagca agctacctga aatgctgaaa atgttcgaag
      661 atcgtttatg tcataaaaca tatttaaatg gtgatcatgt aacccatcct gacttcatgt
      721 tgtatgacgc tcttgatgtt gttttataca tggacccaat gtgcctggat gcgttcccaa
      781 aattagtttg ttttaaaaaa cgtattgaag ctatcccaca aattgataag tacttgaaat
      841 ccagcaagta tatagcatgg cctttgcagg gctggcaagc cacgtttggt ggtggcgacc
      901 atcctccaaa atcggatctg gttccgcgtg gatctccgga aaacctgtat ttccagggcc
      961 atatgaagaa gaaagaagag gaggacgacg acatgaagga attggagaac tgggctggct
     1021 ctatgtaagg atcctcgaga attctcgaga attcatcgtg actgactgac gatctgcctc
     1081 gcgcgtttcg gtgatgacgg tgaaaacctc tgacacatgc agctcccgga gacggtcaca
     1141 gcttgtctgt aagcggatgc cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt
     1201 ggcgggtgtc ggggcgcagc catgacccag tcacgtagcg atagcggagt gtataattct
     1261 tgaagacgaa agggcctcgt gatacgccta tttttatagg ttaatgtcat gataataatg
     1321 gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta
     1381 tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt
     1441 caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc
     1501 ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa
     1561 gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt
     1621 aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt
     1681 ctgctatgtg gcgcggtatt atcccgtgtt gacgccgggc aagagcaact cggtcgccgc
     1741 atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg
     1801 gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg
     1861 gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac
     1921 atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca
     1981 aacgacgagc gtgacaccac gatgcctgca gcaatggcaa caacgttgcg caaactatta
     2041 actggcgaac tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat
     2101 aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa
     2161 tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag
     2221 ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat
     2281 agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt
     2341 tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg
     2401 aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga
     2461 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     2521 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     2581 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
     2641 gtccttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
     2701 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
     2761 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
     2821 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
     2881 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
     2941 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
     3001 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
     3061 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
     3121 ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac
     3181 cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc
     3241 gagtcagtga gcgaggaagc ggaagagcgc ctgatgcggt attttctcct tacgcatctg
     3301 tgcggtattt cacaccgcat aaattccgac accatcgaat ggtgcaaaac ctttcgcggt
     3361 atggcatgat agcgcccgga agagagtcaa ttcagggtgg tgaatgtgaa accagtaacg
     3421 ttatacgatg tcgcagagta tgccggtgtc tcttatcaga ccgtttcccg cgtggtgaac
     3481 caggccagcc acgtttctgc gaaaacgcgg gaaaaagtgg aagcggcgat ggcggagctg
     3541 aattacattc ccaaccgcgt ggcacaacaa ctggcgggca aacagtcgtt gctgattggc
     3601 gttgccacct ccagtctggc cctgcacgcg ccgtcgcaaa ttgtcgcggc gattaaatct
     3661 cgcgccgatc aactgggtgc cagcgtggtg gtgtcgatgg tagaacgaag cggcgtcgaa
     3721 gcctgtaaag cggcggtgca caatcttctc gcgcaacgcg tcagtgggct gatcattaac
     3781 tatccgctgg atgaccagga tgccattgct gtggaagctg cctgcactaa tgttccggcg
     3841 ttatttcttg atgtctctga ccagacaccc atcaacagta ttattttctc ccatgaagac
     3901 ggtacgcgac tgggcgtgga gcatctggtc gcattgggtc accagcaaat cgcgctgtta
     3961 gcgggcccat taagttctgt ctcggcgcgt ctgcgtctgg ctggctggca taaatatctc
     4021 actcgcaatc aaattcagcc gatagcggaa cgggaaggcg actggagtgc catgtccggt
     4081 tttcaacaaa ccatgcaaat gctgaatgag ggcatcgttc ccactgcgat gctggttgcc
     4141 aacgatcaga tggcgctggg cgcaatgcgc gccattaccg agtccgggct gcgcgttggt
     4201 gcggatatct cggtagtggg atacgacgat accgaagaca gctcatgtta tatcccgccg
     4261 ttaaccacca tcaaacagga ttttcgcctg ctggggcaaa ccagcgtgga ccgcttgctg
     4321 caactctctc agggccaggc ggtgaagggc aatcagctgt tgcccgtctc actggtgaaa
     4381 agaaaaacca ccctggcgcc caatacgcaa accgcctctc cccgcgcgtt ggccgattca
     4441 ttaatgcagc tggcacgaca ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat
     4501 taatgtgagt tagctcactc attaggcacc ccaggcttta cactttatgc ttccggctcg
     4561 tatgttgtgt ggaattgtga gcggataaca atttcacaca ggaaacagct atgaccatga
     4621 ttacggattc actggccgtc gttttacaac gtcgtgactg ggaaaaccct ggcgttaccc
     4681 aacttaatcg ccttgcagca catccccctt tcgccagctg gcgtaatagc gaagaggccc
     4741 gcaccgatcg cccttcccaa cagttgcgca gcctgaatgg cgaatggcgc tttgcctggt
     4801 ttccggcacc agaagcggtg ccggaaagct ggctggagtg cgatcttcct gaggccgata
     4861 ctgtcgtcgt cccctcaaac tggcagatgc acggttacga tgcgcccatc tacaccaacg
     4921 taacctatcc cattacggtc aatccgccgt ttgttcccac ggagaatccg acgggttgtt
     4981 actcgctcac atttaatgtt gatgaaagct ggctacagga aggccagacg cgaattattt
     5041 ttgatggcgt tggaatt
//
