LOCUS       sequence-432993-        5659 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Empty 10xHis-SUMO (Smt3) vector for rapid cloning/DNA assembly via 
            SwaI restriction site for E. coli based protein expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5659)
  AUTHORS   Begley M, Aragon M, Baker RW
  TITLE     A structure-based mechanism for initiation of AP-3 coated vesicle 
            formation.
  JOURNAL   Proc Natl Acad Sci U S A. 2024 Dec 24;121(52):e2411974121. doi: 
            10.1073/pnas.2411974121. Epub 2024 Dec 20.
  PUBMED    39705307
REFERENCE   2  (bases 1 to 5659)
  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: "Proc Natl 
            Acad Sci U S A."; date: "2024-12-24"; volume: "121(52):e2411974121. 
            doi"; pages: " 10.1073/pnas.2411974121. Epub 2024 Dec 20"
COMMENT     Sequence Label: MZB139
FEATURES             Location/Qualifiers
     source          1..5659
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     106..125
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     209..228
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        209..227
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    228..252
                     /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             267..289
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             303..332
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             732..749
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(802..820)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      816..863
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      900..1355
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(987..1006)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1197..1218
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1447..2262)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(1447..2262)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     2170..2189
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2254..2273
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2384..2972
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2873..2892
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3126..3143
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3158..3297
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3223..3242)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3383..3405
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3399..3590)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4399..5481)
                     /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     5443..5462
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5482..5559)
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 caaggagatg gcgcccaaca gtcccccggc cacggggcct gccaccatac ccacgccgaa
       61 acaagcgctc atgagcccga agtggcgagc ccgatcttcc ccatcggtga tgtcggcgat
      121 ataggcgcca gcaaccgcac ctgtggcgcc ggtgatgccg gccacgatgc gtccggcgta
      181 gaggatcgag atctcgatcc cgcgaaatta atacgactca ctatagggga attgtgagcg
      241 gataacaatt cccctctaga aataattttg tttaacttta agaaggagat atacatatgg
      301 gccatcacca tcaccatcac catcaccatc acggcagcat gagcgatagc gaagtgaacc
      361 aggaagcgaa accggaagtg aaaccggaag tgaaaccgga aacccatatt aacctgaaag
      421 tgagcgatgg cagcagcgaa atttttttta aaattaaaaa aaccaccccg ctgcgtcgtc
      481 tgatggaagc gtttgcgaaa cgtcagggca aagaaatgga tagcctgcgt tttctgtatg
      541 atggcattcg tattcaggcg gatcagaccc cggaagatct ggatatggaa gataacgata
      601 ttattgaagc gcatcgtgaa cagattggcg gcatttaaat aataaggatc catgggtacc
      661 ctcgagtgaa gacaagctta ggtatttatt cggcgcaaag tgcgtcgggt gatgctgcca
      721 acttagtcga gcaccaccac caccaccact gagatccggc tgctaacaaa gcccgaaagg
      781 aagctgagtt ggctgctgcc accgctgagc aataactagc ataacccctt ggggcctcta
      841 aacgggtctt gaggggtttt ttgctgaaag gaggaactat atccggattg gcgaatggga
      901 cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc
      961 tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac
     1021 gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag
     1081 tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc
     1141 atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg
     1201 actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata
     1261 agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa
     1321 cgcgaatttt aacaaaatat taacgcttac aatttaggtg gcacttttcg gggaaatgtg
     1381 cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaat
     1441 taattcttag aaaaactcat cgagcatcaa atgaaactgc aatttattca tatcaggatt
     1501 atcaatacca tatttttgaa aaagccgttt ctgtaatgaa ggagaaaact caccgaggca
     1561 gttccatagg atggcaagat cctggtatcg gtctgcgatt ccgactcgtc caacatcaat
     1621 acaacctatt aatttcccct cgtcaaaaat aaggttatca agtgagaaat caccatgagt
     1681 gacgactgaa tccggtgaga atggcaaaag tttatgcatt tctttccaga cttgttcaac
     1741 aggccagcca ttacgctcgt catcaaaatc actcgcatca accaaaccgt tattcattcg
     1801 tgattgcgcc tgagcgagac gaaatacgcg atcgctgtta aaaggacaat tacaaacagg
     1861 aatcgaatgc aaccggcgca ggaacactgc cagcgcatca acaatatttt cacctgaatc
     1921 aggatattct tctaatacct ggaatgctgt tttcccgggg atcgcagtgg tgagtaacca
     1981 tgcatcatca ggagtacgga taaaatgctt gatggtcgga agaggcataa attccgtcag
     2041 ccagtttagt ctgaccatct catctgtaac atcattggca acgctacctt tgccatgttt
     2101 cagaaacaac tctggcgcat cgggcttccc atacaatcga tagattgtcg cacctgattg
     2161 cccgacatta tcgcgagccc atttataccc atataaatca gcatccatgt tggaatttaa
     2221 tcgcggccta gagcaagacg tttcccgttg aatatggctc ataacacccc ttgtattact
     2281 gtttatgtaa gcagacagtt ttattgttca tgaccaaaat cccttaacgt gagttttcgt
     2341 tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc
     2401 tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc
     2461 cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac
     2521 caaatactgt ccttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac
     2581 cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt
     2641 cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct
     2701 gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat
     2761 acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt
     2821 atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg
     2881 cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt
     2941 gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt
     3001 tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg
     3061 tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg
     3121 agcgcagcga gtcagtgagc gaggaagcgg aagagcgcct gatgcggtat tttctcctta
     3181 cgcatctgtg cggtatttca caccgcaatg gtgcactctc agtacaatct gctctgatgc
     3241 cgcatagtta agccagtata cactccgcta tcgctacgtg actgggtcat ggctgcgccc
     3301 cgacacccgc caacacccgc tgacgcgccc tgacgggctt gtctgctccc ggcatccgct
     3361 tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc accgtcatca
     3421 ccgaaacgcg cgaggcagct gcggtaaagc tcatcagcgt ggtcgtgaag cgattcacag
     3481 atgtctgcct gttcatccgc gtccagctcg ttgagtttct ccagaagcgt taatgtctgg
     3541 cttctgataa agcgggccat gttaagggcg gttttttcct gtttggtcac tgatgcctcc
     3601 gtgtaagggg gatttctgtt catgggggta atgataccga tgaaacgaga gaggatgctc
     3661 acgatacggg ttactgatga tgaacatgcc cggttactgg aacgttgtga gggtaaacaa
     3721 ctggcggtat ggatgcggcg ggaccagaga aaaatcactc agggtcaatg ccagcgcttc
     3781 gttaatacag atgtaggtgt tccacagggt agccagcagc atcctgcgat gcagatccgg
     3841 aacataatgg tgcagggcgc tgacttccgc gtttccagac tttacgaaac acggaaaccg
     3901 aagaccattc atgttgttgc tcaggtcgca gacgttttgc agcagcagtc gcttcacgtt
     3961 cgctcgcgta tcggtgattc attctgctaa ccagtaaggc aaccccgcca gcctagccgg
     4021 gtcctcaacg acaggagcac gatcatgcgc acccgtgggg ccgccatgcc ggcgataatg
     4081 gcctgcttct cgccgaaacg tttggtggcg ggaccagtga cgaaggcttg agcgagggcg
     4141 tgcaagattc cgaataccgc aagcgacagg ccgatcatcg tcgcgctcca gcgaaagcgg
     4201 tcctcgccga aaatgaccca gagcgctgcc ggcacctgtc ctacgagttg catgataaag
     4261 aagacagtca taagtgcggc gacgatagtc atgccccgcg cccaccggaa ggagctgact
     4321 gggttgaagg ctctcaaggg catcggtcga gatcccggtg cctaatgagt gagctaactt
     4381 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg
     4441 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ccagggtggt
     4501 ttttcttttc accagtgaga cgggcaacag ctgattgccc ttcaccgcct ggccctgaga
     4561 gagttgcagc aagcggtcca cgctggtttg ccccagcagg cgaaaatcct gtttgatggt
     4621 ggttaacggc gggatataac atgagctgtc ttcggtatcg tcgtatccca ctaccgagat
     4681 atccgcacca acgcgcagcc cggactcggt aatggcgcgc attgcgccca gcgccatctg
     4741 atcgttggca accagcatcg cagtgggaac gatgccctca ttcagcattt gcatggtttg
     4801 ttgaaaaccg gacatggcac tccagtcgcc ttcccgttcc gctatcggct gaatttgatt
     4861 gcgagtgaga tatttatgcc agccagccag acgcagacgc gccgagacag aacttaatgg
     4921 gcccgctaac agcgcgattt gctggtgacc caatgcgacc agatgctcca cgcccagtcg
     4981 cgtaccgtct tcatgggaga aaataatact gttgatgggt gtctggtcag agacatcaag
     5041 aaataacgcc ggaacattag tgcaggcagc ttccacagca atggcatcct ggtcatccag
     5101 cggatagtta atgatcagcc cactgacgcg ttgcgcgaga agattgtgca ccgccgcttt
     5161 acaggcttcg acgccgcttc gttctaccat cgacaccacc acgctggcac ccagttgatc
     5221 ggcgcgagat ttaatcgccg cgacaatttg cgacggcgcg tgcagggcca gactggaggt
     5281 ggcaacgcca atcagcaacg actgtttgcc cgccagttgt tgtgccacgc ggttgggaat
     5341 gtaattcagc tccgccatcg ccgcttccac tttttcccgc gttttcgcag aaacgtggct
     5401 ggcctggttc accacgcggg aaacggtctg ataagagaca ccggcatact ctgcgacatc
     5461 gtataacgtt actggtttca cattcaccac cctgaattga ctctcttccg ggcgctatca
     5521 tgccataccg cgaaaggttt tgcgccattc gatggtgtcc gggatctcga cgctctccct
     5581 tatgcgactc ctgcattagg aagcagccca gtagtaggtt gaggccgttg agcaccgccg
     5641 ccgcaaggaa tggtgcatg
//
