LOCUS       sequence-354447-        5658 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5658)
  TITLE     LIC N-terminal fusion vectors for E. coli expression
REFERENCE   2  (bases 1 to 5658)
  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"
COMMENT     Sequence Label: pET His6 Sumo TEV LIC cloning vector (1S)
FEATURES             Location/Qualifiers
     source          1..5658
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1310..1501
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(1495..1517)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    1603..1742
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     1658..1677
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(1757..1774)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1928..2516)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2008..2027)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(2627..2646)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             2638..3453
                     /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             2638..3453
                     /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     complement(2711..2730)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      complement(3546..4001)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3683..3704)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3895..3914
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     terminator      complement(4038..4085)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     4081..4099
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(4152..4169)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(4235..4255)
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             complement(4262..4555)
                     /codon_start=1
                     /gene="S. cerevisiae SMT3 (truncated)"
                     /product="cleavable ubiquitin-like protein tag"
                     /label=SUMO
                     /translation="MSDSEVNQEAKPEVKPEVKPETHINLKVSDGSSEIFFKIKKTTPL
                     RRLMEAFAKRQGKEMDSLRFLYDGIRIQADQTPEDLDMEDNDIIEAHREQIGG"
     CDS             complement(4574..4591)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             4610..4632
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    4647..4671
                     /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     complement(4671..4690)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4672..4690)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4774..4793)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        4999..5076
                     /gene="lacI"
                     /label=lacI promoter
     CDS             join(5077..5658,1..501)
                     /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(5096..5115)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 cgtctgcgtc tggctggctg gcataaatat ctcactcgca atcaaattca gccgatagcg
       61 gaacgggaag gcgactggag tgccatgtcc ggttttcaac aaaccatgca aatgctgaat
      121 gagggtatcg ttcccactgc gatgctggtt gccaacgatc agatggcgct gggcgcaatg
      181 cgcgccatta ccgagtccgg gctgcgcgtt ggtgcggata tctcggtagt gggatacgac
      241 gataccgaag acagctcatg ttatatcccg ccgttaacca ccatcaaaca ggattttcgc
      301 ctgctggggc aaaccagcgt ggaccgcttg ctgcaactct ctcagggcca ggcggtgaag
      361 ggcaatcagc tgttgcccgt ctcactggtg aaaagaaaaa ccaccctggc gcccaatacg
      421 caaaccgcct ctccccgcgc gttggccgat tcattaatgc agctggcacg acaggtttcc
      481 cgactggaaa gcgggcagtg agcgcaacgc aattaatgta agttagctca ctcattaggc
      541 accgggatct cgaccgatgc ccttgagagc cttcaaccca gtcagctcct tccggtgggc
      601 gcggggcatg actatcgtcg ccgcacttat gactgtcttc tttatcatgc aactcgtagg
      661 acaggtgccg gcagcgctct gggtcatttt cggcgaggac cgctttcgct ggagcgcgac
      721 gatgatcggc ctgtcgcttg cggtattcgg aatcttgcac gccctcgctc aagccttcgt
      781 cactggtccc gccaccaaac gtttcggcga gaagcaggcc attatcgccg gcatggcggc
      841 cccacgggtg cgcatgatcg tgctcctgtc gttgaggacc cggctaggct ggcggggttg
      901 ccttactggt tagcagaatg aatcaccgat acgcgagcga acgtgaagcg actgctgctg
      961 caaaacgtct gcgacctgag caacaacatg aatggtcttc ggtttccgtg tttcgtaaag
     1021 tctggaaacg cggaagtcag cgccctgcac cattatgttc cggatctgca tcgcaggatg
     1081 ctgctggcta ccctgtggaa cacctacatc tgtattaacg aagcgctggc attgaccctg
     1141 agtgattttt ctctggtccc gccgcatcca taccgccagt tgtttaccct cacaacgttc
     1201 cagtaaccgg gcatgttcat catcagtaac ccgtatcgtg agcatcctct ctcgtttcat
     1261 cggtatcatt acccccatga acagaaatcc cccttacacg gaggcatcag tgaccaaaca
     1321 ggaaaaaacc gcccttaaca tggcccgctt tatcagaagc cagacattaa cgcttctgga
     1381 gaaactcaac gagctggacg cggatgaaca ggcagacatc tgtgaatcgc ttcacgacca
     1441 cgctgatgag ctttaccgca gctgcctcgc gcgtttcggt gatgacggtg aaaacctctg
     1501 acacatgcag ctcccggaga cggtcacagc ttgtctgtaa gcggatgccg ggagcagaca
     1561 agcccgtcag ggcgcgtcag cgggtgttgg cgggtgtcgg ggcgcagcca tgacccagtc
     1621 acgtagcgat agcggagtgt atactggctt aactatgcgg catcagagca gattgtactg
     1681 agagtgcacc attgcggtgt gaaataccgc acagatgcgt aaggagaaaa taccgcatca
     1741 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag
     1801 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag
     1861 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
     1921 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
     1981 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
     2041 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
     2101 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg
     2161 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
     2221 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
     2281 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
     2341 ggtggcctaa ctacggctac actagaagga cagtatttgg tatctgcgct ctgctgaagc
     2401 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
     2461 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
     2521 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
     2581 ttttggtcat gaacaataaa actgtctgct tacataaaca gtaatacaag gggtgttatg
     2641 agccatattc aacgggaaac gtcttgctct aggccgcgat taaattccaa catggatgct
     2701 gatttatatg ggtataaatg ggctcgcgat aatgtcgggc aatcaggtgc gacaatctat
     2761 cgattgtatg ggaagcccga tgcgccagag ttgtttctga aacatggcaa aggtagcgtt
     2821 gccaatgatg ttacagatga gatggtcaga ctaaactggc tgacggaatt tatgcctctt
     2881 ccgaccatca agcattttat ccgtactcct gatgatgcat ggttactcac cactgcgatc
     2941 cccgggaaaa cagcattcca ggtattagaa gaatatcctg attcaggtga aaacattgtt
     3001 gatgcgctgg cagtgttcct gcgccggttg cattcgattc ctgtttgtaa ttgtcctttt
     3061 aacagcgatc gcgtatttcg tctcgctcag gcgcaatcac gaatgaataa cggtttggtt
     3121 gatgcgagtg attttgatga cgagcgtaat ggctggcctg ttgaacaagt ctggaaagaa
     3181 atgcataaac ttttgccatt ctcaccggat tcagtcgtca ctcatggtga tttctcactt
     3241 gataacctta tttttgacga ggggaaatta ataggttgta ttgatgttgg acgagtcgga
     3301 atcgcagacc gataccagga tcttgccatc ctatggaact gcctcggtga gttttctcct
     3361 tcattacaga aacggctttt tcaaaaatat ggtattgata atcctgatat gaataaattg
     3421 cagtttcatt tgatgctcga tgagtttttc taagaattaa ttcatgagcg gatacatatt
     3481 tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc
     3541 acctgaaatt gtaaacgtta ctagtttgtt aaaattcgcg ttaaattttt gttaaatcag
     3601 ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa aagaatagac
     3661 cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa agaacgtgga
     3721 ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac gtgaaccatc
     3781 accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg
     3841 gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa aggaagggaa
     3901 gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac
     3961 caccacaccc gccgcgctta atgcgccgct acagggcgcg tcccattcgc caatccggat
     4021 atagttcctc ctttcagcaa aaaacccctc aagacccgtt tagaggcccc aaggggttat
     4081 gctagttatt gctcagcggt ggcagcagcc aactcagctt cctttcgggc tttgttagca
     4141 gccggatctc agtggtggtg gtggtggtgc tcgagtgcgg ccgcaagctt gtcgacggcg
     4201 ctcgaattcg gatccgttat ccacttccaa tattggattg gaagtacagg ttttcctcga
     4261 tcccaccaat ctgttctctg tgagcctcaa taatatcgtt atcctccatg tccaaatctt
     4321 caggggtctg atcagcttga attctaatac cgtcgtacaa gaatcttaag gagtccattt
     4381 ccttaccctg tcttttagcg aacgcttcca tcagccttct taaaggagtg gtctttttga
     4441 tcttgaagaa gatctctgaa gatccatcgg acacctttaa attgatgtga gtctcaggct
     4501 tgacttctgg cttgacctct ggcttagctt cttgattgac ttctgagtcc gacatgctag
     4561 ccatagaaga accatggtga tggtgatggt gagaagaacc catggtatat ctccttctta
     4621 aagttaaaca aaattatttc tagaggggaa ttgttatccg ctcacaattc ccctatagtg
     4681 agtcgtatta atttcgcggg atcgagatct cgatcctcta cgccggacgc atcgtggccg
     4741 gcatcaccgg cgccacaggt gcggttgctg gcgcctatat cgccgacatc accgatgggg
     4801 aagatcgggc tcgccacttc gggctcatga gcgcttgttt cggcgtgggt atggtggcag
     4861 gccccgtggc cgggggactg ttgggcgcca tctccttgca tgcaccattc cttgcggcgg
     4921 cggtgctcaa cggcctcaac ctactactgg gctgcttcct aatgcaggag tcgcataagg
     4981 gagagcgtcg agatcccgga caccatcgaa tggcgcaaaa cctttcgcgg tatggcatga
     5041 tagcgcccgg aagagagtca attcagggtg gtgaatgtga aaccagtaac gttatacgat
     5101 gtcgcagagt atgccggtgt ctcttatcag accgtttccc gcgtggtgaa ccaggccagc
     5161 cacgtttctg cgaaaacgcg ggaaaaagtg gaagcggcga tggcggagct gaattacatt
     5221 cccaaccgcg tggcacaaca actggcgggc aaacagtcgt tgctgattgg cgttgccacc
     5281 tccagtctgg ccctgcacgc gccgtcgcaa attgtcgcgg cgattaaatc tcgcgccgat
     5341 caactgggtg ccagcgtggt ggtgtcgatg gtagaacgaa gcggcgtcga agcctgtaaa
     5401 gcggcggtgc acaatcttct cgcgcaacgc gtcagtgggc tgatcattaa ctatccgctg
     5461 gatgaccagg atgccattgc tgtggaagct gcctgcacta atgttccggc gttatttctt
     5521 gatgtctctg accagacacc catcaacagt attattttct cccatgaaga cggtacgcga
     5581 ctgggcgtgg agcatctggt cgcattgggt caccagcaaa tcgcgctgtt agcgggccca
     5641 ttaagttctg tctcggcg
//
