LOCUS       sequence-255907-        6103 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of truncated Glo3 (373-459) in E.Coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6103)
  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 6103)
  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-GRM
FEATURES             Location/Qualifiers
     source          1..6103
                     /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             1495..1524
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     terminator      1547..1641
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     primer_bind     complement(1751..1770)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      2413..2499
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(2592..3674)
                     /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     3636..3655
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(3675..3752)
                     /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(3836..3858)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3944..4084
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3999..4018
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4099..4116)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4270..4858)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4350..4369)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5029..5820,5821..5889))
                     /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     5652..5671
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5890..5994)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6062..6080
                     /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 ggcggctcga gatatacggg cagaatcgcg
     1321 gaaaggtacg gcacacaaaa agctatatca tctgatcaat tatttggtcg tggctcattt
     1381 gatgaagcag caaacaggga agcgcatgac aagttgaaaa catttgacaa tgctacatca
     1441 atttcctctt cttcatactt tggagaagat aaagaagtgg atgaacgcgg atcccatcat
     1501 catcatcacc accaccatca ccactaagct taattagctg agcttggact cctgttgata
     1561 gatccagtaa tgacctcaga actccatctg gatttgttca gaacgctcgg ttgccgccgg
     1621 gcgtttttta ttggtgagaa tccaagctag cttggcgaga ttttcaggag ctaaggaagc
     1681 taaaatggag aaaaaaatca ctggatatac caccgttgat atatcccaat ggcatcgtaa
     1741 agaacatttt gaggcatttc agtcagttgc tcaatgtacc tataaccaga ccgttcagct
     1801 ggatattacg gcctttttaa agaccgtaaa gaaaaataag cacaagtttt atccggcctt
     1861 tattcacatt cttgcccgcc tgatgaatgc tcatccggaa tttcgtatgg caatgaaaga
     1921 cggtgagctg gtgatatggg atagtgttca cccttgttac accgttttcc atgagcaaac
     1981 tgaaacgttt tcatcgctct ggagtgaata ccacgacgat ttccggcagt ttctacacat
     2041 atattcgcaa gatgtggcgt gttacggtga aaacctggcc tatttcccta aagggtttat
     2101 tgagaatatg tttttcgtct cagccaatcc ctgggtgagt ttcaccagtt ttgatttaaa
     2161 cgtggccaat atggacaact tcttcgcccc cgttttcacc atgcatgggc aaatattata
     2221 cgcaaggcga caaggtgctg atgccgctgg cgattcaggt tcatcatgcc gtttgtgatg
     2281 gcttccatgt cggcagaatg cttaatgaat tacaacagta ctgcgatgag tggcagggcg
     2341 gggcgtaatt tttttaaggc agttattggt gcccttaaac gcctggggta atgactctct
     2401 agcttgaggc atcaaataaa acgaaaggct cagtcgaaag actgggcctt tcgttttatc
     2461 tgttgtttgt cggtgaacgc tctcctgagt aggacaaatc cgccctctag attacgtgca
     2521 gtcgatgata agctgtcaaa catgagaatt gtgcctaatg agtgagctaa cttacattaa
     2581 ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat
     2641 gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgccagggt ggtttttctt
     2701 ttcaccagtg agacgggcaa cagctgattg cccttcaccg cctggccctg agagagttgc
     2761 agcaagcggt ccacgctggt ttgccccagc aggcgaaaat cctgtttgat ggtggttaac
     2821 ggcgggatat aacatgagct gtcttcggta tcgtcgtatc ccactaccga gatatccgca
     2881 ccaacgcgca gcccggactc ggtaatggcg cgcattgcgc ccagcgccat ctgatcgttg
     2941 gcaaccagca tcgcagtggg aacgatgccc tcattcagca tttgcatggt ttgttgaaaa
     3001 ccggacatgg cactccagtc gccttcccgt tccgctatcg gctgaatttg attgcgagtg
     3061 agatatttat gccagccagc cagacgcaga cgcgccgaga cagaacttaa tgggcccgct
     3121 aacagcgcga tttgctggtg acccaatgcg accagatgct ccacgcccag tcgcgtaccg
     3181 tcttcatggg agaaaataat actgttgatg ggtgtctggt cagagacatc aagaaataac
     3241 gccggaacat tagtgcaggc agcttccaca gcaatggcat cctggtcatc cagcggatag
     3301 ttaatgatca gcccactgac gcgttgcgcg agaagattgt gcaccgccgc tttacaggct
     3361 tcgacgccgc ttcgttctac catcgacacc accacgctgg cacccagttg atcggcgcga
     3421 gatttaatcg ccgcgacaat ttgcgacggc gcgtgcaggg ccagactgga ggtggcaacg
     3481 ccaatcagca acgactgttt gcccgccagt tgttgtgcca cgcggttggg aatgtaattc
     3541 agctccgcca tcgccgcttc cactttttcc cgcgttttcg cagaaacgtg gctggcctgg
     3601 ttcaccacgc gggaaacggt ctgataagag acaccggcat actctgcgac atcgtataac
     3661 gttactggtt tcacattcac caccctgaat tgactctctt ccgggcgcta tcatgccata
     3721 ccgcgaaagg ttttgcacca ttcgatggtg tcggaatttc gggcagcgtt gggtcctggc
     3781 cacgggtgcg catgatctag agctgcctcg cgcgtttcgg tgatgacggt gaaaacctct
     3841 gacacatgca gctcccggag acggtcacag cttgtctgta agcggatgcc gggagcagac
     3901 aagcccgtca gggcgcgtca gcgggtgttg gcgggtgtcg gggcgcagcc atgacccagt
     3961 cacgtagcga tagcggagtg tatactggct taactatgcg gcatcagagc agattgtact
     4021 gagagtgcac catatgcggt gtgaaatacc gcacagatgc gtaaggagaa aataccgcat
     4081 caggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg
     4141 agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc
     4201 aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt
     4261 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag
     4321 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc
     4381 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc
     4441 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt
     4501 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt
     4561 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc
     4621 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa
     4681 gtggtggcct aactacggct acactagaag gacagtattt ggtatctgcg ctctgctgaa
     4741 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg
     4801 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga
     4861 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg
     4921 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg
     4981 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt
     5041 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact
     5101 ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat
     5161 gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg
     5221 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg
     5281 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat
     5341 tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc
     5401 ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt
     5461 cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc
     5521 agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga
     5581 gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc
     5641 gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa
     5701 acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta
     5761 acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg
     5821 agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg
     5881 aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat
     5941 gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt
     6001 tccccgaaaa gtgccacctg acgtctaaga aaccattatt atcatgacat taacctataa
     6061 aaataggcgt atcacgaggc cctttcgtct tcacctcgag aaa
//
