LOCUS       sequence-235344-        7038 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of  truncated Glo3 (1-375) in E.Coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7038)
  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 7038)
  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-GAP-BoCCS
FEATURES             Location/Qualifiers
     source          1..7038
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(10..24,25..30,31..47,48..54)
                     /label=T5 promoter
                     /note="bacteriophage T5 promoter for E. coli RNA 
                     polymerase, with embedded lac operator"
     protein_bind    30..46
                     /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    62..78
                     /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             97..108
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             124..1221
                     /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     1195..1218
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             1237..1257
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             2443..2472
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     terminator      2495..2589
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             2633..3292
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFIMSKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(2699..2718)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      3357..3443
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(3536..4618)
                     /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     4580..4599
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(4619..4696)
                     /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(4780..4802)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4888..5028
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4943..4962
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(5043..5060)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5214..5802)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5294..5313)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5973..6764,6765..6833))
                     /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     6596..6615
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6834..6938)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7006..7024
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 ctcgagaaat cataaaaaat ttatttgctt tgtgagcgga taacaattat aatagattca
       61 attgtgagcg gataacaatt tcacacagaa ttcattaaag aggagaaatt aaccatgacc
      121 gggaaaactg aagaaggtaa actggtaatc tggattaacg gcgataaagg ctataacggt
      181 ctcgctgaag tcggtaagaa attcgagaaa gataccggaa ttaaagtcac cgttgagcat
      241 ccggataaac tggaagagaa attcccacag gttgcggcaa ctggcgatgg ccctgacatt
      301 atcttctggg cacacgaccg ctttggtggc tacgctcaat ctggcctgtt ggctgaaatc
      361 accccggaca aagcgttcca ggacaagctg tatccgttta cctgggatgc cgtacgttac
      421 aacggcaagc tgattgctta cccgatcgct gttgaagcgt tatcgctgat ttataacaaa
      481 gatctgctgc cgaacccgcc aaaaacctgg gaagagatcc cggcgctgga taaagaactg
      541 aaagcgaaag gtaagagcgc gctgatgttc aacctgcaag aaccgtactt cacctggccg
      601 ctgattgctg ctgacggggg ttatgcgttc aagtatgaaa acggcaagta cgacattaaa
      661 gacgtgggcg tggataacgc tggcgcgaaa gcgggtctga ccttcctggt tgacctgatt
      721 aaaaacaaac acatgaatgc agacactgat tactccatcg cagaagctgc ctttaataaa
      781 ggcgaaacag cgatgaccat caacggcccg tgggcatggt ccaacatcga caccagcaaa
      841 gtgaattatg gtgtaacggt actgccgacc ttcaagggtc aaccatccaa accgttcgtt
      901 ggcgtgctga gcgcaggtat taacgccgcc agtccgaaca aagagctggc aaaagagttc
      961 ctcgaaaact atctgctgac tgatgaaggt ctggaagcgg ttaataaaga caaaccgctg
     1021 ggtgccgtag cgctgaagtc ttacgaggaa gagttggcga aagatccacg tattgccgcc
     1081 acgatggaaa acgcccagaa aggtgaaatc atgccgaaca tcccgcagat gtccgctttc
     1141 tggtatgccg tgcgtactgc ggtgatcaac gccgccagcg gtcgtcagac tgtcgatgaa
     1201 gccctgaaag acgcgcagac taattcgagc tcgggtgaaa acctctactt ccaagggtcc
     1261 atgggtaccc ggggggaagg cggcggcggc tcgggcggcg gcggctcgat gagtaacgat
     1321 gaaggagaaa catttgccac ggaacagact actcaacaag tttttcagaa gctaggtagc
     1381 aacatggaaa atcgagtttg ttttgattgt ggtaacaaga atcctacttg gacttcggta
     1441 ccttttggtg tgatgctgtg tattcaatgc tctgctgtgc atagaaacat gggtgttcat
     1501 atcacctttg taaagtcttc tacattagat aaatggacaa ttaacaattt aagaaggttt
     1561 aagttgggtg ggaaccataa ggctcgtgat tttttcttaa agaacaatgg gaaacagcta
     1621 ctaaatacag caaatgttga cgcgaaaaca aaatatacga gccctgttgc taagaaatat
     1681 aagatacatc tcgacaagaa ggttcaaaag gatatggagt tatacccatc agaattagtc
     1741 ttgaacggtc aagattcttc ggactcaccg ttggacaccg attcagacgc ttccaggtcg
     1801 acttccaaag aaaatagtgt tgacgatttt ttctctaatt ggcaaaaacc aagctccaat
     1861 tccagttcta aactcaatgt aaatacaggc tctctcgccc caaaaaacaa caccacaggc
     1921 agcacaccca agaccactgt cactaaaacg agaagttcca tcttaacggc tagcagaaag
     1981 aaaccggtat taaactcgca agataaaaag aaacattcaa ttttgtcttc gtcaagaaag
     2041 cccactagac taaccgcaaa gaaagtcgac aaatcacagg cagaagactt gtttgatcaa
     2101 tttaaaaaag aggctcaaca agaaaaggag gacgagttca ctaattcgtc ttctagcacc
     2161 aaaattagac aaaatgatta cgattcccaa tttatgaata acagtaaagg caacaataat
     2221 aacagtatag atgatattaa tacccagcct gatgaattca atgatttttt aaatgacact
     2281 tcaaacagct tcgacacaac tagaaaagaa cagcaagata ctttaacacc caagtttgcc
     2341 aaattgggat ttggtatgac catgaatgac gccaatgatt tggcaaagca acaaaaagaa
     2401 tctcagaaaa tagctcaggg accaagatat acgcgcggat cccatcatca tcatcaccac
     2461 caccatcacc actaagctta attagctgag cttggactcc tgttgataga tccagtaatg
     2521 acctcagaac tccatctgga tttgttcaga acgctcggtt gccgccgggc gttttttatt
     2581 ggtgagaatc caagctagct tggcgagatt ttcaggagct aaggaagcta aaatggagaa
     2641 aaaaatcact ggatatacca ccgttgatat atcccaatgg catcgtaaag aacattttga
     2701 ggcatttcag tcagttgctc aatgtaccta taaccagacc gttcagctgg atattacggc
     2761 ctttttaaag accgtaaaga aaaataagca caagttttat ccggccttta ttcacattct
     2821 tgcccgcctg atgaatgctc atccggaatt tcgtatggca atgaaagacg gtgagctggt
     2881 gatatgggat agtgttcacc cttgttacac cgttttccat gagcaaactg aaacgttttc
     2941 atcgctctgg agtgaatacc acgacgattt ccggcagttt ctacacatat attcgcaaga
     3001 tgtggcgtgt tacggtgaaa acctggccta tttccctaaa gggtttattg agaatatgtt
     3061 tttcgtctca gccaatccct gggtgagttt caccagtttt gatttaaacg tggccaatat
     3121 ggacaacttc ttcgcccccg ttttcatcat gagcaaatat tatacgcaag gcgacaaggt
     3181 gctgatgccg ctggcgattc aggttcatca tgccgtttgt gatggcttcc atgtcggcag
     3241 aatgcttaat gaattacaac agtactgcga tgagtggcag ggcggggcgt aattttttta
     3301 aggcagttat tggtgccctt aaacgcctgg ggtaatgact ctctagcttg aggcatcaaa
     3361 taaaacgaaa ggctcagtcg aaagactggg cctttcgttt tatctgttgt ttgtcggtga
     3421 acgctctcct gagtaggaca aatccgccct ctagattacg tgcagtcgat gataagctgt
     3481 caaacatgag aattgtgcct aatgagtgag ctaacttaca ttaattgcgt tgcgctcact
     3541 gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc
     3601 ggggagaggc ggtttgcgta ttgggcgcca gggtggtttt tcttttcacc agtgagacgg
     3661 gcaacagctg attgcccttc accgcctggc cctgagagag ttgcagcaag cggtccacgc
     3721 tggtttgccc cagcaggcga aaatcctgtt tgatggtggt taacggcggg atataacatg
     3781 agctgtcttc ggtatcgtcg tatcccacta ccgagatatc cgcaccaacg cgcagcccgg
     3841 actcggtaat ggcgcgcatt gcgcccagcg ccatctgatc gttggcaacc agcatcgcag
     3901 tgggaacgat gccctcattc agcatttgca tggtttgttg aaaaccggac atggcactcc
     3961 agtcgccttc ccgttccgct atcggctgaa tttgattgcg agtgagatat ttatgccagc
     4021 cagccagacg cagacgcgcc gagacagaac ttaatgggcc cgctaacagc gcgatttgct
     4081 ggtgacccaa tgcgaccaga tgctccacgc ccagtcgcgt accgtcttca tgggagaaaa
     4141 taatactgtt gatgggtgtc tggtcagaga catcaagaaa taacgccgga acattagtgc
     4201 aggcagcttc cacagcaatg gcatcctggt catccagcgg atagttaatg atcagcccac
     4261 tgacgcgttg cgcgagaaga ttgtgcaccg ccgctttaca ggcttcgacg ccgcttcgtt
     4321 ctaccatcga caccaccacg ctggcaccca gttgatcggc gcgagattta atcgccgcga
     4381 caatttgcga cggcgcgtgc agggccagac tggaggtggc aacgccaatc agcaacgact
     4441 gtttgcccgc cagttgttgt gccacgcggt tgggaatgta attcagctcc gccatcgccg
     4501 cttccacttt ttcccgcgtt ttcgcagaaa cgtggctggc ctggttcacc acgcgggaaa
     4561 cggtctgata agagacaccg gcatactctg cgacatcgta taacgttact ggtttcacat
     4621 tcaccaccct gaattgactc tcttccgggc gctatcatgc cataccgcga aaggttttgc
     4681 accattcgat ggtgtcggaa tttcgggcag cgttgggtcc tggccacggg tgcgcatgat
     4741 ctagagctgc ctcgcgcgtt tcggtgatga cggtgaaaac ctctgacaca tgcagctccc
     4801 ggagacggtc acagcttgtc tgtaagcgga tgccgggagc agacaagccc gtcagggcgc
     4861 gtcagcgggt gttggcgggt gtcggggcgc agccatgacc cagtcacgta gcgatagcgg
     4921 agtgtatact ggcttaacta tgcggcatca gagcagattg tactgagagt gcaccatatg
     4981 cggtgtgaaa taccgcacag atgcgtaagg agaaaatacc gcatcaggcg ctcttccgct
     5041 tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac
     5101 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga
     5161 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
     5221 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
     5281 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
     5341 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
     5401 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
     5461 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
     5521 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
     5581 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
     5641 ggctacacta gaaggacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
     5701 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
     5761 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
     5821 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga
     5881 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
     5941 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
     6001 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
     6061 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca
     6121 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
     6181 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
     6241 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg
     6301 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
     6361 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
     6421 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
     6481 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
     6541 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
     6601 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
     6661 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
     6721 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
     6781 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
     6841 ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt
     6901 gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca
     6961 cctgacgtct aagaaaccat tattatcatg acattaacct ataaaaatag gcgtatcacg
     7021 aggccctttc gtcttcac
//
