LOCUS       sequence-307465-        5485 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses GST-SUMO tagged CTA2 Subunit with 11th Strand.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5485)
  AUTHORS   Luong P, Li Q, Chen PF, Wrighton PJ, Chang D, Dwyer S, Bayer MT, 
            Snapper SB, Hansen SH, Thiagarajah JT, Goessling W, Lencer WI
  TITLE     A quantitative single-cell assay for retrograde membrane traffic 
            enables rapid detection of defects in cellular organization.
  JOURNAL   Mol Biol Cell. 2019 Nov 27:mbcE19070375. doi: 
            10.1091/mbc.E19-07-0375.
  PUBMED    31774722
REFERENCE   2  (bases 1 to 5485)
  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: "Mol Biol 
            Cell. 2019 Nov 27:mbcE19070375. doi: 10.1091/mbc.E19-07-0375."
COMMENT     Sequence Label: CTA2-mNG211
FEATURES             Location/Qualifiers
     source          1..5485
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(526..531,532..549,550..556)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    564..580
                     /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     588..604
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     588..604
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             600..1127
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(620..637)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(620..636)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(629..651)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     promoter        join(1229..1234,1235..1250,1251..1257)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1265..1281
                     /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             1304..1957
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     1915..1937
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             1964..1981
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             1982..2002
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             2009..2302
                     /codon_start=1
                     /gene="S. cerevisiae SMT3 (truncated)"
                     /product="cleavable ubiquitin-like protein tag"
                     /label=SUMO
                     /translation="MSDSEVNQEAKPEVKPEVKPETHINLKVSDGSSEIFFKIKKTTPL
                     RRLMEAFAKRQGKEMDSLRFLYDGIRIQADQTPEDLDMEDNDIIEAHREQIGG"
     primer_bind     complement(2581..2603)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2748..2766)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2834..2938
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2939..3007,3008..3799)
                     /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(3157..3176)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3970..4558
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4459..4478
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4712..4729
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        4802..4879
                     /gene="lacI"
                     /label=lacI promoter
     CDS             join(4880..5485,1..477)
                     /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(4899..4918)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 aaatatctca ctcgcaatca aattcagccg atagcggaac gggaaggcga ctggagtgcc
       61 atgtccggtt ttcaacaaac catgcaaatg ctgaatgagg gcatcgttcc cactgcgatg
      121 ctggttgcca acgatcagat ggcgctgggc gcaatgcgcg ccattaccga gtccgggctg
      181 cgcgttggtg cggatatctc ggtagtggga tacgacgata ccgaagacag ctcatgttat
      241 atcccgccgt taaccaccat caaacaggat tttcgcctgc tggggcaaac cagcgtggac
      301 cgcttgctgc aactctctca gggccaggcg gtgaagggca atcagctgtt gcccgtctca
      361 ctggtgaaaa gaaaaaccac cctggcgccc aatacgcaaa ccgcctctcc ccgcgcgttg
      421 gccgattcat taatgcagct ggcacgacag gtttcccgac tggaaagcgg gcagtgagcg
      481 caacgcaatt aatgtaagtt agctcactca ttaggcaccc caggctttac actttatgct
      541 tccggctcgt atgttgtgtg gaattgtgag cggataacaa tttcacacag gaaacagcta
      601 tgaccatgat tacggattca ctggccgtcg ttttacaacg tcgtgactgg gaaaaccctg
      661 gcgttaccca acttaatcgc cttgcagcac atcccccttt cgccagctgg cgtaatagcg
      721 aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc gaatggcgct
      781 ttgcctggtt tccggcacca gaagcggtgc cggaaagctg gctggagtgc gatcttcctg
      841 aggccgatac tgtcgtcgtc ccctcaaact ggcagatgca cggttacgat gcgcccatct
      901 acaccaacgt gacctatccc attacggtca atccgccgtt tgttcccacg gagaatccga
      961 cgggttgtta ctcgctcaca tttaatgttg atgaaagctg gctacaggaa ggccagacgc
     1021 gaattatttt tgatggcgtt ggaattagct tatcgactgc acggtgcacc aatgcttctg
     1081 gcgtcaggca gccatcggaa gctgtggtat ggctgtgcag gtcgtaaatc actgcataat
     1141 tcgtgtcgct caaggcgcac tcccgttctg gataatgttt tttgcgccga catcataacg
     1201 gttctggcaa atattctgaa atgagctgtt gacaattaat catcggctcg tataatgtgt
     1261 ggaattgtga gcggataaca atttcacaca ggaaacagta ttcatgtccc ctatactagg
     1321 ttattggaaa attaagggcc ttgtgcaacc cactcgactt cttttggaat atcttgaaga
     1381 aaaatatgaa gagcatttgt atgagcgcga tgaaggtgat aaatggcgaa acaaaaagtt
     1441 tgaattgggt ttggagtttc ccaatcttcc ttattatatt gatggtgatg ttaaattaac
     1501 acagtctatg gccatcatac gttatatagc tgacaagcac aacatgttgg gtggttgtcc
     1561 aaaagagcgt gcagagattt caatgcttga aggagcggtt ttggatatta gatacggtgt
     1621 ttcgagaatt gcatatagta aagactttga aactctcaaa gttgattttc ttagcaagct
     1681 acctgaaatg ctgaaaatgt tcgaagatcg tttatgtcat aaaacatatt taaatggtga
     1741 tcatgtaacc catcctgact tcatgttgta tgacgctctt gatgttgttt tatacatgga
     1801 cccaatgtgc ctggatgcgt tcccaaaatt agtttgtttt aaaaaacgta ttgaagctat
     1861 cccacaaatt gataagtact tgaaatccag caagtatata gcatggcctt tgcagggctg
     1921 gcaagccacg tttggtggtg gcgaccatcc tccaaaatcg gatctggttc cgcgtggatc
     1981 tgaaaacctg tattttcagg gcggatccat gtcggactca gaagtcaatc aagaagctaa
     2041 gccagaggtc aagccagaag tcaagcctga gactcacatc aatttaaagg tgtccgatgg
     2101 atcttcagag atcttcttca agatcaaaaa gaccactcct ttaagaaggc tgatggaagc
     2161 gttcgctaaa agacagggta aggaaatgga ctccttaaga ttcttgtacg acggtattag
     2221 aattcaagct gatcagaccc ctgaagattt ggacatggag gataacgata ttattgaggc
     2281 tcacagagaa cagattggtg gtaccgagct caacttcaag gagtggcaaa aggcctttac
     2341 cgatatgatg ggaggtggag gttcctcgat gagtaatact tgcgatgaaa aaacccaaag
     2401 tctaggtgta aaattccttg acgaatacca atctaaagtt aaaagacaaa tattttcagg
     2461 ctatcaatct gatattgata cacataatag aattaaggat gaattatgac tcgagctgca
     2521 gcggccgcat cgtgactgac tgacgatctg cctcgcgcgt ttcggtgatg acggtgaaaa
     2581 cctctgacac atgcagctcc cggagacggt cacagcttgt ctgtaagcgg atgccgggag
     2641 cagacaagcc cgtcagggcg cgtcagcggg tgttggcggg tgtcggggcg cagccatgac
     2701 ccagtcacgt agcgatagcg gagtgtataa ttcttgaaga cgaaagggcc tcgtgatacg
     2761 cctattttta taggttaatg tcatgataat aatggtttct tagacgtcag gtggcacttt
     2821 tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta
     2881 tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtat
     2941 gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt
     3001 ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg
     3061 agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga
     3121 agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg
     3181 tgttgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt
     3241 tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg
     3301 cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg
     3361 aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga
     3421 tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc
     3481 tgcagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc
     3541 ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc
     3601 ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc gtgggtctcg
     3661 cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac
     3721 gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc
     3781 actgattaag cattggtaac tgtcagacca agtttactca tatatacttt agattgattt
     3841 aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac
     3901 caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa
     3961 aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc
     4021 accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt
     4081 aactggcttc agcagagcgc agataccaaa tactgtcctt ctagtgtagc cgtagttagg
     4141 ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc
     4201 agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt
     4261 accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga
     4321 gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct
     4381 tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg
     4441 cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca
     4501 cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa
     4561 cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt
     4621 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga
     4681 taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga
     4741 gcgcctgatg cggtattttc tccttacgca tctgtgcggt atttcacacc gcataaattc
     4801 cgacaccatc gaatggcgca aaacctttcg cggtatggca tgatagcgcc cggaagagag
     4861 tcaattcagg gtggtgaatg tgaaaccagt aacgttatac gatgtcgcag agtatgccgg
     4921 tgtctcttat cagaccgttt cccgcgtggt gaaccaggcc agccacgttt ctgcgaaaac
     4981 gcgggaaaaa gtggaagcgg cgatggcgga gctgaattac attcccaacc gcgtggcaca
     5041 acaactggcg ggcaaacagt cgttgctgat tggcgttgcc acctccagtc tggccctgca
     5101 cgcgccgtcg caaattgtcg cggcgattaa atctcgcgcc gatcaactgg gtgccagcgt
     5161 ggtggtgtcg atggtagaac gaagcggcgt cgaagcctgt aaagcggcgg tgcacaatct
     5221 tctcgcgcaa cgcgtcagtg ggctgatcat taactatccg ctggatgacc aggatgccat
     5281 tgctgtggaa gctgcctgca ctaatgttcc ggcgttattt cttgatgtct ctgaccagac
     5341 acccatcaac agtattattt tctcccatga agacggtacg cgactgggcg tggagcatct
     5401 ggtcgcattg ggtcaccagc aaatcgcgct gttagcgggc ccattaagtt ctgtctcggc
     5461 gcgtctgcgt ctggctggct ggcat
//
