LOCUS       sequence-255892-        7042 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 7042)
  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 7042)
  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..7042
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     RBS             1..12
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             28..1125
                     /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     1099..1122
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             1141..1161
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             2347..2376
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     terminator      2399..2493
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     primer_bind     complement(2603..2622)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      3265..3351
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(3444..4526)
                     /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     4488..4507
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(4527..4604)
                     /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(4688..4710)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4796..4936
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4851..4870
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4951..4968)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5122..5710)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5202..5221)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5881..6672,6673..6741))
                     /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     6504..6523
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6742..6846)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6914..6932
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        join(6956..6970,6971..6976,6977..6993,6994..7000)
                     /label=T5 promoter
                     /note="bacteriophage T5 promoter for E. coli RNA 
                     polymerase, with embedded lac operator"
     protein_bind    6976..6992
                     /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    7008..7024
                     /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)."
ORIGIN
        1 aaagaggaga aattaaccat gaccgggaaa actgaagaag gtaaactggt aatctggatt
       61 aacggcgata aaggctataa cggtctcgct gaagtcggta agaaattcga gaaagatacc
      121 ggaattaaag tcaccgttga gcatccggat aaactggaag agaaattccc acaggttgcg
      181 gcaactggcg atggccctga cattatcttc tgggcacacg accgctttgg tggctacgct
      241 caatctggcc tgttggctga aatcaccccg gacaaagcgt tccaggacaa gctgtatccg
      301 tttacctggg atgccgtacg ttacaacggc aagctgattg cttacccgat cgctgttgaa
      361 gcgttatcgc tgatttataa caaagatctg ctgccgaacc cgccaaaaac ctgggaagag
      421 atcccggcgc tggataaaga actgaaagcg aaaggtaaga gcgcgctgat gttcaacctg
      481 caagaaccgt acttcacctg gccgctgatt gctgctgacg ggggttatgc gttcaagtat
      541 gaaaacggca agtacgacat taaagacgtg ggcgtggata acgctggcgc gaaagcgggt
      601 ctgaccttcc tggttgacct gattaaaaac aaacacatga atgcagacac tgattactcc
      661 atcgcagaag ctgcctttaa taaaggcgaa acagcgatga ccatcaacgg cccgtgggca
      721 tggtccaaca tcgacaccag caaagtgaat tatggtgtaa cggtactgcc gaccttcaag
      781 ggtcaaccat ccaaaccgtt cgttggcgtg ctgagcgcag gtattaacgc cgccagtccg
      841 aacaaagagc tggcaaaaga gttcctcgaa aactatctgc tgactgatga aggtctggaa
      901 gcggttaata aagacaaacc gctgggtgcc gtagcgctga agtcttacga ggaagagttg
      961 gcgaaagatc cacgtattgc cgccacgatg gaaaacgccc agaaaggtga aatcatgccg
     1021 aacatcccgc agatgtccgc tttctggtat gccgtgcgta ctgcggtgat caacgccgcc
     1081 agcggtcgtc agactgtcga tgaagccctg aaagacgcgc agactaattc gagctcgggt
     1141 gaaaacctct acttccaagg gtccatgggt acccgggggg aaggcggcgg cggctcgggc
     1201 ggcggcggct cgatgagtaa cgatgaagga gaaacatttg ccacggaaca gactactcaa
     1261 caagtctttc agaagctagg tagcaacatg gaaaatcgag tttgttttga ttgtggtaac
     1321 aagaatccta cttggacttc ggtacctttt ggtgtgatgc tgtgtattca atgctctgct
     1381 gtgcatagaa acatgggtgt tcatatcacc tttgtaaagt cttctacatt agataaatgg
     1441 acaattaaca atttaagaag gtttaagttg ggcgggaacc ataaggctcg tgattttttc
     1501 ttaaagaaca atgggaaaca gctactaaat acagcaaatg ttgacgcgaa aacaaaatat
     1561 acgagccctg ttgctaagaa atataagata catctcgaca agaaggttca aaaggatatg
     1621 gagttatacc catcagaatt agtcttgaac ggtcaagatt cttcggactc accgttggac
     1681 accgattcag acgcttccag gtcgacttcc aaagaaaata gtgttgacga ttttttctct
     1741 aattggcaaa aaccaagctc caattccagt tctaaactca atgtaaatac aggctctctc
     1801 gccccaaaaa acaacaccac aggcagcaca cccaagacca ctgtcactaa aacgagaagt
     1861 tccatcttaa cggctagcag aaagaaaccg gtattaaact cgcaagataa aaagaaacat
     1921 tcaattttgt cttcgtcaag aaagcccact agactaaccg caaagaaagt cgacaaatca
     1981 caggcagaag acttgtttga tcaatttaaa aaagaggctc aacaagaaaa ggaggacgag
     2041 ttcactaatt cgtcttctag caccaaaatt agacaaaatg attacgattc ccaatttatg
     2101 aataacagta aaggcaacaa taataacagt atagatgata ttaataccca gcctgatgaa
     2161 ttcaatgatt ttttaaatga cacttcaaac agcttcgaca caactagaaa agaacagcaa
     2221 gatactttaa cacccaagtt tgccaaattg ggatttggta tgaccatgaa tgacgccaat
     2281 gatttggcaa agcaacaaaa agaatctcag aaaatagctc agggaccaag atatacgcgc
     2341 ggatcccatc atcatcatca ccaccaccat caccactaag cttaattagc tgagcttgga
     2401 ctcctgttga tagatccagt aatgacctca gaactccatc tggatttgtt cagaacgctc
     2461 ggttgccgcc gggcgttttt tattggtgag aatccaagct agcttggcga gattttcagg
     2521 agctaaggaa gctaaaatgg agaaaaaaat cactggatat accaccgttg atatatccca
     2581 atggcatcgt aaagaacatt ttgaggcatt tcagtcagtt gctcaatgta cctataacca
     2641 gaccgttcag ctggatatta cggccttttt aaagaccgta aagaaaaata agcacaagtt
     2701 ttatccggcc tttattcaca ttcttgcccg cctgatgaat gctcatccgg aatttcgtat
     2761 ggcaatgaaa gacggtgagc tggtgatatg ggatagtgtt cacccttgtt acaccgtttt
     2821 ccatgagcaa actgaaacgt tttcatcgct ctggagtgaa taccacgacg atttccggca
     2881 gtttctacac atatattcgc aagatgtggc gtgttacggt gaaaacctgg cctatttccc
     2941 taaagggttt attgagaata tgtttttcgt ctcagccaat ccctgggtga gtttcaccag
     3001 ttttgattta aacgtggcca atatggacaa cttcttcgcc cccgttttca ccatgcatgg
     3061 gcaaatatta tacgcaaggc gacaaggtgc tgatgccgct ggcgattcag gttcatcatg
     3121 ccgtttgtga tggcttccat gtcggcagaa tgcttaatga attacaacag tactgcgatg
     3181 agtggcaggg cggggcgtaa tttttttaag gcagttattg gtgcccttaa acgcctgggg
     3241 taatgactct ctagcttgag gcatcaaata aaacgaaagg ctcagtcgaa agactgggcc
     3301 tttcgtttta tctgttgttt gtcggtgaac gctctcctga gtaggacaaa tccgccctct
     3361 agattacgtg cagtcgatga taagctgtca aacatgagaa ttgtgcctaa tgagtgagct
     3421 aacttacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc
     3481 agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgccagg
     3541 gtggtttttc ttttcaccag tgagacgggc aacagctgat tgcccttcac cgcctggccc
     3601 tgagagagtt gcagcaagcg gtccacgctg gtttgcccca gcaggcgaaa atcctgtttg
     3661 atggtggtta acggcgggat ataacatgag ctgtcttcgg tatcgtcgta tcccactacc
     3721 gagatatccg caccaacgcg cagcccggac tcggtaatgg cgcgcattgc gcccagcgcc
     3781 atctgatcgt tggcaaccag catcgcagtg ggaacgatgc cctcattcag catttgcatg
     3841 gtttgttgaa aaccggacat ggcactccag tcgccttccc gttccgctat cggctgaatt
     3901 tgattgcgag tgagatattt atgccagcca gccagacgca gacgcgccga gacagaactt
     3961 aatgggcccg ctaacagcgc gatttgctgg tgacccaatg cgaccagatg ctccacgccc
     4021 agtcgcgtac cgtcttcatg ggagaaaata atactgttga tgggtgtctg gtcagagaca
     4081 tcaagaaata acgccggaac attagtgcag gcagcttcca cagcaatggc atcctggtca
     4141 tccagcggat agttaatgat cagcccactg acgcgttgcg cgagaagatt gtgcaccgcc
     4201 gctttacagg cttcgacgcc gcttcgttct accatcgaca ccaccacgct ggcacccagt
     4261 tgatcggcgc gagatttaat cgccgcgaca atttgcgacg gcgcgtgcag ggccagactg
     4321 gaggtggcaa cgccaatcag caacgactgt ttgcccgcca gttgttgtgc cacgcggttg
     4381 ggaatgtaat tcagctccgc catcgccgct tccacttttt cccgcgtttt cgcagaaacg
     4441 tggctggcct ggttcaccac gcgggaaacg gtctgataag agacaccggc atactctgcg
     4501 acatcgtata acgttactgg tttcacattc accaccctga attgactctc ttccgggcgc
     4561 tatcatgcca taccgcgaaa ggttttgcac cattcgatgg tgtcggaatt tcgggcagcg
     4621 ttgggtcctg gccacgggtg cgcatgatct agagctgcct cgcgcgtttc ggtgatgacg
     4681 gtgaaaacct ctgacacatg cagctcccgg agacggtcac agcttgtctg taagcggatg
     4741 ccgggagcag acaagcccgt cagggcgcgt cagcgggtgt tggcgggtgt cggggcgcag
     4801 ccatgaccca gtcacgtagc gatagcggag tgtatactgg cttaactatg cggcatcaga
     4861 gcagattgta ctgagagtgc accatatgcg gtgtgaaata ccgcacagat gcgtaaggag
     4921 aaaataccgc atcaggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt
     4981 tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc
     5041 aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa
     5101 aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa
     5161 tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc
     5221 ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc
     5281 cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag
     5341 ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga
     5401 ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc
     5461 gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac
     5521 agagttcttg aagtggtggc ctaactacgg ctacactaga aggacagtat ttggtatctg
     5581 cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca
     5641 aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa
     5701 aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa
     5761 ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt
     5821 aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag
     5881 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
     5941 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc
     6001 cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa
     6061 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca
     6121 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa
     6181 cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt
     6241 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc
     6301 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact
     6361 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc
     6421 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg
     6481 ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct
     6541 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc
     6601 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag
     6661 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac
     6721 acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg
     6781 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
     6841 tccgcgcaca tttccccgaa aagtgccacc tgacgtctaa gaaaccatta ttatcatgac
     6901 attaacctat aaaaataggc gtatcacgag gccctttcgt cttcacctcg agaaatcata
     6961 aaaaatttat ttgctttgtg agcggataac aattataata gattcaattg tgagcggata
     7021 acaatttcac acagaattca tt
//
