LOCUS       sequence-255872-        6811 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of truncated Glo3 (137-459) in E.Coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6811)
  AUTHORS   Arakel EC, Huranova M, Estrada AF, Rau EM, Spang A, Schwappach B
  TITLE     Dissection of GTPase-activating proteins reveals functional 
            asymmetry in the COPI coat of budding yeast.
  JOURNAL   J Cell Sci. 2019 Aug 29;132(16). pii: jcs.232124. doi: 
            10.1242/jcs.232124.
  PUBMED    31331965
REFERENCE   2  (bases 1 to 6811)
  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 Cell 
            Sci."; date: "2019-08-29"; volume: "132(16). pii: jcs.232124. doi"; 
            pages: " 10.1242/jcs.232124"
COMMENT     Sequence Label: A102-BoCCS-GRM
FEATURES             Location/Qualifiers
     source          1..6811
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(1..15,16..21,22..38,39..45)
                     /label=T5 promoter
                     /note="bacteriophage T5 promoter for E. coli RNA 
                     polymerase, with embedded lac operator"
     protein_bind    21..37
                     /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)."
     protein_bind    53..69
                     /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             88..99
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             115..1212
                     /codon_start=1
                     /gene="malE (mutated)"
                     /product="maltose binding protein from E. coli"
                     /label=MBP
                     /note="This version of the gene does not encode a signal 
                     sequence, so MBP will remain in the cytosol."
                     /translation="KTEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLEE
                     KFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKLI
                     AYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIAA
                     DGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGET
                     AMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFLE
                     NYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAFW
                     YAVRTAVINAASGRQTVDEALKDAQT"
     primer_bind     1186..1209
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             1228..1248
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             2203..2232
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     terminator      2255..2349
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     primer_bind     complement(2459..2478)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      3121..3207
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(3300..4382)
                     /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     4344..4363
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(4383..4460)
                     /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."
     primer_bind     complement(4544..4566)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4652..4792
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4707..4726
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4807..4824)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4978..5566)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5058..5077)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5737..6528,6529..6597))
                     /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     6360..6379
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6598..6702)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6770..6788
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcataaaaaa tttatttgct ttgtgagcgg ataacaatta taatagattc aattgtgagc
       61 ggataacaat ttcacacaga attcattaaa gaggagaaat taaccatgac cgggaaaact
      121 gaagaaggta aactggtaat ctggattaac ggcgataaag gctataacgg tctcgctgaa
      181 gtcggtaaga aattcgagaa agataccgga attaaagtca ccgttgagca tccggataaa
      241 ctggaagaga aattcccaca ggttgcggca actggcgatg gccctgacat tatcttctgg
      301 gcacacgacc gctttggtgg ctacgctcaa tctggcctgt tggctgaaat caccccggac
      361 aaagcgttcc aggacaagct gtatccgttt acctgggatg ccgtacgtta caacggcaag
      421 ctgattgctt acccgatcgc tgttgaagcg ttatcgctga tttataacaa agatctgctg
      481 ccgaacccgc caaaaacctg ggaagagatc ccggcgctgg ataaagaact gaaagcgaaa
      541 ggtaagagcg cgctgatgtt caacctgcaa gaaccgtact tcacctggcc gctgattgct
      601 gctgacgggg gttatgcgtt caagtatgaa aacggcaagt acgacattaa agacgtgggc
      661 gtggataacg ctggcgcgaa agcgggtctg accttcctgg ttgacctgat taaaaacaaa
      721 cacatgaatg cagacactga ttactccatc gcagaagctg cctttaataa aggcgaaaca
      781 gcgatgacca tcaacggccc gtgggcatgg tccaacatcg acaccagcaa agtgaattat
      841 ggtgtaacgg tactgccgac cttcaagggt caaccatcca aaccgttcgt tggcgtgctg
      901 agcgcaggta ttaacgccgc cagtccgaac aaagagctgg caaaagagtt cctcgaaaac
      961 tatctgctga ctgatgaagg tctggaagcg gttaataaag acaaaccgct gggtgccgta
     1021 gcgctgaagt cttacgagga agagttggcg aaagatccac gtattgccgc cacgatggaa
     1081 aacgcccaga aaggtgaaat catgccgaac atcccgcaga tgtccgcttt ctggtatgcc
     1141 gtgcgtactg cggtgatcaa cgccgccagc ggtcgtcaga ctgtcgatga agccctgaaa
     1201 gacgcgcaga ctaattcgag ctcgggtgaa aacctctact tccaagggtc catgggtacc
     1261 cggggggaag gcggcggcgg ctcgggcggc ggcggctcgg agttataccc atcagaatta
     1321 gtcttgaacg gtcaagattc ttcggactca ccgttggaca ccgattcaga cgcttccagg
     1381 tcgacttcca aagaaaatag tgttgacgat tttttctcta attggcaaaa accaagctcc
     1441 aattccagtt ctaaactcaa tgtaaataca ggctctctcg ccccaaaaaa caacaccaca
     1501 ggcagcacac ccaagaccac tgtcactaaa acgagaagtt ccatcttaac ggctagcaga
     1561 aagaaaccgg tattaaactc gcaagataaa aagaaacatt caattttgtc ttcgtcaaga
     1621 aagcccacta gactaaccgc aaagaaagtc gacaaatcac aggcagaaga cttgtttgat
     1681 caatttaaaa aagaggctca acaagaaaag gaggacgagt tcactaattc gtcttctagc
     1741 accaaaatta gacaaaatga ttacgattcc caatttatga ataacagtaa aggcaacaat
     1801 aataacagta tagatgatat taatacccag cctgatgaat tcaatgattt tttaaatgac
     1861 acttcaaaca gcttcgacac aactagaaaa gaacagcaag atactttaac acccaagttt
     1921 gccaaattgg gatttggtat gaccatgaat gacgccaatg atttggcaaa gcaacaaaaa
     1981 gaatctcaga aaatagctca gggaccaaga tatacgggca gaatcgcgga aaggtacggc
     2041 acacaaaaag ctatatcatc tgatcaatta tttggtcgtg gctcatttga tgaagcagca
     2101 aacagggaag cgcatgacaa gttgaaaaca tttgacaatg ctacatcaat ttcctcttct
     2161 tcatactttg gagaagataa agaagtggat gaacgcggat cccatcatca tcatcaccac
     2221 caccatcacc actaagctta attagctgag cttggactcc tgttgataga tccagtaatg
     2281 acctcagaac tccatctgga tttgttcaga acgctcggtt gccgccgggc gttttttatt
     2341 ggtgagaatc caagctagct tggcgagatt ttcaggagct aaggaagcta aaatggagaa
     2401 aaaaatcact ggatatacca ccgttgatat atcccaatgg catcgtaaag aacattttga
     2461 ggcatttcag tcagttgctc aatgtaccta taaccagacc gttcagctgg atattacggc
     2521 ctttttaaag accgtaaaga aaaataagca caagttttat ccggccttta ttcacattct
     2581 tgcccgcctg atgaatgctc atccggaatt tcgtatggca atgaaagacg gtgagctggt
     2641 gatatgggat agtgttcacc cttgttacac cgttttccat gagcaaactg aaacgttttc
     2701 atcgctctgg agtgaatacc acgacgattt ccggcagttt ctacacatat attcgcaaga
     2761 tgtggcgtgt tacggtgaaa acctggccta tttccctaaa gggtttattg agaatatgtt
     2821 tttcgtctca gccaatccct gggtgagttt caccagtttt gatttaaacg tggccaatat
     2881 ggacaacttc ttcgcccccg ttttcaccat gcatgggcaa atattatacg caaggcgaca
     2941 aggtgctgat gccgctggcg attcaggttc atcatgccgt ttgtgatggc ttccatgtcg
     3001 gcagaatgct taatgaatta caacagtact gcgatgagtg gcagggcggg gcgtaatttt
     3061 tttaaggcag ttattggtgc ccttaaacgc ctggggtaat gactctctag cttgaggcat
     3121 caaataaaac gaaaggctca gtcgaaagac tgggcctttc gttttatctg ttgtttgtcg
     3181 gtgaacgctc tcctgagtag gacaaatccg ccctctagat tacgtgcagt cgatgataag
     3241 ctgtcaaaca tgagaattgt gcctaatgag tgagctaact tacattaatt gcgttgcgct
     3301 cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac
     3361 gcgcggggag aggcggtttg cgtattgggc gccagggtgg tttttctttt caccagtgag
     3421 acgggcaaca gctgattgcc cttcaccgcc tggccctgag agagttgcag caagcggtcc
     3481 acgctggttt gccccagcag gcgaaaatcc tgtttgatgg tggttaacgg cgggatataa
     3541 catgagctgt cttcggtatc gtcgtatccc actaccgaga tatccgcacc aacgcgcagc
     3601 ccggactcgg taatggcgcg cattgcgccc agcgccatct gatcgttggc aaccagcatc
     3661 gcagtgggaa cgatgccctc attcagcatt tgcatggttt gttgaaaacc ggacatggca
     3721 ctccagtcgc cttcccgttc cgctatcggc tgaatttgat tgcgagtgag atatttatgc
     3781 cagccagcca gacgcagacg cgccgagaca gaacttaatg ggcccgctaa cagcgcgatt
     3841 tgctggtgac ccaatgcgac cagatgctcc acgcccagtc gcgtaccgtc ttcatgggag
     3901 aaaataatac tgttgatggg tgtctggtca gagacatcaa gaaataacgc cggaacatta
     3961 gtgcaggcag cttccacagc aatggcatcc tggtcatcca gcggatagtt aatgatcagc
     4021 ccactgacgc gttgcgcgag aagattgtgc accgccgctt tacaggcttc gacgccgctt
     4081 cgttctacca tcgacaccac cacgctggca cccagttgat cggcgcgaga tttaatcgcc
     4141 gcgacaattt gcgacggcgc gtgcagggcc agactggagg tggcaacgcc aatcagcaac
     4201 gactgtttgc ccgccagttg ttgtgccacg cggttgggaa tgtaattcag ctccgccatc
     4261 gccgcttcca ctttttcccg cgttttcgca gaaacgtggc tggcctggtt caccacgcgg
     4321 gaaacggtct gataagagac accggcatac tctgcgacat cgtataacgt tactggtttc
     4381 acattcacca ccctgaattg actctcttcc gggcgctatc atgccatacc gcgaaaggtt
     4441 ttgcaccatt cgatggtgtc ggaatttcgg gcagcgttgg gtcctggcca cgggtgcgca
     4501 tgatctagag ctgcctcgcg cgtttcggtg atgacggtga aaacctctga cacatgcagc
     4561 tcccggagac ggtcacagct tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg
     4621 gcgcgtcagc gggtgttggc gggtgtcggg gcgcagccat gacccagtca cgtagcgata
     4681 gcggagtgta tactggctta actatgcggc atcagagcag attgtactga gagtgcacca
     4741 tatgcggtgt gaaataccgc acagatgcgt aaggagaaaa taccgcatca ggcgctcttc
     4801 cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc
     4861 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat
     4921 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
     4981 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
     5041 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     5101 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     5161 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     5221 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     5281 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     5341 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     5401 ctacggctac actagaagga cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     5461 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     5521 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     5581 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     5641 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc
     5701 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc
     5761 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta
     5821 gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga
     5881 cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg
     5941 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc
     6001 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat
     6061 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag
     6121 gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat
     6181 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa
     6241 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa
     6301 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga
     6361 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg
     6421 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc
     6481 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
     6541 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact
     6601 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat
     6661 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt
     6721 gccacctgac gtctaagaaa ccattattat catgacatta acctataaaa ataggcgtat
     6781 cacgaggccc tttcgtcttc acctcgagaa a
//
