LOCUS       sequence-325053-        5086 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Vector for recombinant protein expression in E. coli; 
            PreScission-cleavable N-terminal GST and TEV-cleavable C-terminal 
            6xHis.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5086)
  AUTHORS   Saltzman AL, Soo MW, Aram R, Lee JT
  TITLE     Multiple Histone Methyl-Lysine Readers Ensure Robust Development and
            Germline Immortality in Caenorhabditis elegans.
  JOURNAL   Genetics. 2018 Nov;210(3):907-923. doi: 10.1534/genetics.118.301518.
            Epub 2018 Sep 5.
  PUBMED    30185429
REFERENCE   2  (bases 1 to 5086)
  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: 
            "10.1534/genetics.118.301518"; journalName: "Genetics"; date: 
            "2018-11"; volume: "210"; issue: "3"; pages: "907-923"
COMMENT     Sequence Label: pALS343 (pGEX-2TP-cTevHis-Gibson/MCS)
FEATURES             Location/Qualifiers
     source          1..5086
                     /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             942..965
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     protein_bind    972..996
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(1003..1027)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             1029..1049
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             1056..1073
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1136..1158)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1303..1321)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1389..1493
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1494..1562,1563..2354)
                     /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(1712..1731)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2525..3113
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3014..3033
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3267..3284
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        3357..3434
                     /gene="lacI"
                     /label=lacI promoter
     CDS             3435..4517
                     /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(3454..3473)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(4566..4571,4572..4589,4590..4596)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4604..4620
                     /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     4628..4644
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4628..4644
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(4640..5086,1..81)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(4660..4677)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4660..4676)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4669..4691)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 agcttatcga 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 gatccccggg actggaagtt ctgttccagg
      961 ggcccggtgg tacaagtttg tacaaaaaag ccggcttggg gtacccagct ttcttgtaca
     1021 aagtggtgga gaacctgtac ttccaatccg ctgggcacca ccatcaccat cactgaggaa
     1081 ttcatcgtga ctgactgacg atctgcctcg cgcgtttcgg tgatgacggt gaaaacctct
     1141 gacacatgca gctcccggag acggtcacag cttgtctgta agcggatgcc gggagcagac
     1201 aagcccgtca gggcgcgtca gcgggtgttg gcgggtgtcg gggcgcagcc atgacccagt
     1261 cacgtagcga tagcggagtg tataattctt gaagacgaaa gggcctcgtg atacgcctat
     1321 ttttataggt taatgtcatg ataataatgg tttcttagac gtcaggtggc acttttcggg
     1381 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
     1441 tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta
     1501 ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg
     1561 ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg
     1621 gttacatcga actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac
     1681 gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtgttg
     1741 acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt
     1801 actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg
     1861 ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac
     1921 cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt
     1981 gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgcag
     2041 caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc
     2101 aacaattaat agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc
     2161 ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta
     2221 tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg
     2281 ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga
     2341 ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac
     2401 ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa
     2461 tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat
     2521 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
     2581 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg
     2641 gcttcagcag agcgcagata ccaaatactg tccttctagt gtagccgtag ttaggccacc
     2701 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
     2761 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
     2821 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
     2881 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
     2941 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
     3001 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
     3061 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
     3121 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc
     3181 ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg
     3241 ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc
     3301 tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcata aattccgaca
     3361 ccatcgaatg gcgcaaaacc tttcgcggta tggcatgata gcgcccggaa gagagtcaat
     3421 tcagggtggt gaatgtgaaa ccagtaacgt tatacgatgt cgcagagtat gccggtgtct
     3481 cttatcagac cgtttcccgc gtggtgaacc aggccagcca cgtttctgcg aaaacgcggg
     3541 aaaaagtgga agcggcgatg gcggagctga attacattcc caaccgcgtg gcacaacaac
     3601 tggcgggcaa acagtcgttg ctgattggcg ttgccacctc cagtctggcc ctgcacgcgc
     3661 cgtcgcaaat tgtcgcggcg attaaatctc gcgccgatca actgggtgcc agcgtggtgg
     3721 tgtcgatggt agaacgaagc ggcgtcgaag cctgtaaagc ggcggtgcac aatcttctcg
     3781 cgcaacgcgt cagtgggctg atcattaact atccgctgga tgaccaggat gccattgctg
     3841 tggaagctgc ctgcactaat gttccggcgt tatttcttga tgtctctgac cagacaccca
     3901 tcaacagtat tattttctcc catgaagacg gtacgcgact gggcgtggag catctggtcg
     3961 cattgggtca ccagcaaatc gcgctgttag cgggcccatt aagttctgtc tcggcgcgtc
     4021 tgcgtctggc tggctggcat aaatatctca ctcgcaatca aattcagccg atagcggaac
     4081 gggaaggcga ctggagtgcc atgtccggtt ttcaacaaac catgcaaatg ctgaatgagg
     4141 gcatcgttcc cactgcgatg ctggttgcca acgatcagat ggcgctgggc gcaatgcgcg
     4201 ccattaccga gtccgggctg cgcgttggtg cggatatctc ggtagtggga tacgacgata
     4261 ccgaagacag ctcatgttat atcccgccgt taaccaccat caaacaggat tttcgcctgc
     4321 tggggcaaac cagcgtggac cgcttgctgc aactctctca gggccaggcg gtgaagggca
     4381 atcagctgtt gcccgtctca ctggtgaaaa gaaaaaccac cctggcgccc aatacgcaaa
     4441 ccgcctctcc ccgcgcgttg gccgattcat taatgcagct ggcacgacag gtttcccgac
     4501 tggaaagcgg gcagtgagcg caacgcaatt aatgtaagtt agctcactca ttaggcaccc
     4561 caggctttac actttatgct tccggctcgt atgttgtgtg gaattgtgag cggataacaa
     4621 tttcacacag gaaacagcta tgaccatgat tacggattca ctggccgtcg ttttacaacg
     4681 tcgtgactgg gaaaaccctg gcgttaccca acttaatcgc cttgcagcac atcccccttt
     4741 cgccagctgg cgtaatagcg aagaggcccg caccgatcgc ccttcccaac agttgcgcag
     4801 cctgaatggc gaatggcgct ttgcctggtt tccggcacca gaagcggtgc cggaaagctg
     4861 gctggagtgc gatcttcctg aggccgatac tgtcgtcgtc ccctcaaact ggcagatgca
     4921 cggttacgat gcgcccatct acaccaacgt gacctatccc attacggtca atccgccgtt
     4981 tgttcccacg gagaatccga cgggttgtta ctcgctcaca tttaatgttg atgaaagctg
     5041 gctacaggaa ggccagacgc gaattatttt tgatggcgtt ggaatt
//
