LOCUS       sequence-205272-        6235 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses AglC from M. voltae with an N-terminal T7 tag and a 
            C-terminal His6 tag. Plasmid has codons optimized for E. coli 
            expression..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6235)
  AUTHORS   Zamora CY, Schocker NS, Chang MM, Imperiali B
  TITLE     Chemoenzymatic Synthesis and Applications of Prokaryote-Specific 
            UDP-Sugars.
  JOURNAL   Methods Enzymol. 2017;597:145-186. doi: 10.1016/bs.mie.2017.06.003. 
            Epub 2017 Jul 5.
  PUBMED    28935101
REFERENCE   2  (bases 1 to 6235)
  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: "Methods 
            Enzymol."; date: "2017"; volume: "597:145-186. doi"; pages: " 
            10.1016/bs.mie.2017.06.003. Epub 2017 Jul 5"
COMMENT     Sequence Label: T7-AglC-His6
FEATURES             Location/Qualifiers
     source          1..6235
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..20
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1..19
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    20..44
                     /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             59..81
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             89..121
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="MASMTGGQQMG"
     CDS             1100..1117
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1170..1188)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1184..1231
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1268..1723
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1355..1374)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1565..1586
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1815..2630)
                     /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(1815..2630)
                     /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     2538..2557
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2622..2641
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2752..3340
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3241..3260
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3494..3511
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3526..3665
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3591..3610)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3751..3773
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3767..3958)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4767..5849)
                     /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     5811..5830
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5850..5927)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6133..6152
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
       61 tgtttaactt taagaaggag atatacatat ggctagcatg actggtggac agcaaatggg
      121 tcgcggatcc atgaatttta tctccattat catccctacg ttcaacgagg aaaaatatat
      181 cactaaatgc ctggagaact ggtttaacca ggattacccg aaagaaaact acgaaatcct
      241 gatctttgac ggtaagtcta ccgacaaaac tctggatgtt atcaaagaac tgcagaagaa
      301 acacaacttc gaaaacatta aaatctatac caacgaaaag cgcaaacagg tatacgcatt
      361 taatgaaggc atcaaaaatg cgaacggcga ttttttcatc attttcggtg ctcatgcgta
      421 tccggaacaa gatttcctga aaaacaacat tgaaacctac cagcgtatca agaaagaaga
      481 accgaaactg gcgggcgtgg gtggtattat caacaaaatc tccgaaaaca tgtctgcgga
      541 gatcgctaaa gtaatctatt ctactccgct gtctggcggc tcttcttttt ggtacgccaa
      601 agagggtttc ttctccaaca ccgtggttta tggtatgtac gacaccaaaa tgattaaaga
      661 gtccgaaatc ctgtttgata ccgacttcat caccggccag gacttcgaat tcaacctgca
      721 cctgatcaag gaaggcttca aactgtacac taacccgaac atcgtttcta gctattacac
      781 ccgttctagc gtcaaaaagt ttatcaaaca gacgatttcc tacggcgccg ctaaaggtct
      841 gatgattcgt aaaggctact ttaacattct gtggctgttc ccattcggtt tcctgttcat
      901 gctgctgagc atcatcatta ccggcctgct gattttcatc tacatcatgg caattctgat
      961 cgacaccatc cgcctgctga tcaaaactcg tgaaccgctg tacatcgcac tgccgattct
     1021 gctgttcctg ttccactgtc tgatcagcta tggtttcttc aaaggtctga ttaagggtaa
     1081 ttctaccttc aaactcgagc accaccacca ccaccactga gatccggctg ctaacaaagc
     1141 ccgaaaggaa gctgagttgg ctgctgccac cgctgagcaa taactagcat aaccccttgg
     1201 ggcctctaaa cgggtcttga ggggtttttt gctgaaagga ggaactatat ccggattggc
     1261 gaatgggacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc
     1321 gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt
     1381 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc
     1441 cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt
     1501 agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt
     1561 aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt
     1621 gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa
     1681 aaatttaacg cgaattttaa caaaatatta acgcttacaa tttaggtggc acttttcggg
     1741 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
     1801 tcatgaatta attcttagaa aaactcatcg agcatcaaat gaaactgcaa tttattcata
     1861 tcaggattat caataccata tttttgaaaa agccgtttct gtaatgaagg agaaaactca
     1921 ccgaggcagt tccataggat ggcaagatcc tggtatcggt ctgcgattcc gactcgtcca
     1981 acatcaatac aacctattaa tttcccctcg tcaaaaataa ggttatcaag tgagaaatca
     2041 ccatgagtga cgactgaatc cggtgagaat ggcaaaagtt tatgcatttc tttccagact
     2101 tgttcaacag gccagccatt acgctcgtca tcaaaatcac tcgcatcaac caaaccgtta
     2161 ttcattcgtg attgcgcctg agcgagacga aatacgcgat cgctgttaaa aggacaatta
     2221 caaacaggaa tcgaatgcaa ccggcgcagg aacactgcca gcgcatcaac aatattttca
     2281 cctgaatcag gatattcttc taatacctgg aatgctgttt tcccggggat cgcagtggtg
     2341 agtaaccatg catcatcagg agtacggata aaatgcttga tggtcggaag aggcataaat
     2401 tccgtcagcc agtttagtct gaccatctca tctgtaacat cattggcaac gctacctttg
     2461 ccatgtttca gaaacaactc tggcgcatcg ggcttcccat acaatcgata gattgtcgca
     2521 cctgattgcc cgacattatc gcgagcccat ttatacccat ataaatcagc atccatgttg
     2581 gaatttaatc gcggcctaga gcaagacgtt tcccgttgaa tatggctcat aacacccctt
     2641 gtattactgt ttatgtaagc agacagtttt attgttcatg accaaaatcc cttaacgtga
     2701 gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc
     2761 tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt
     2821 ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc
     2881 gcagatacca aatactgtcc ttctagtgta gccgtagtta ggccaccact tcaagaactc
     2941 tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg
     3001 cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg
     3061 gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga
     3121 actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc
     3181 ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg
     3241 gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg
     3301 atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt
     3361 tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc
     3421 tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg
     3481 aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa gagcgcctga tgcggtattt
     3541 tctccttacg catctgtgcg gtatttcaca ccgcaatggt gcactctcag tacaatctgc
     3601 tctgatgccg catagttaag ccagtataca ctccgctatc gctacgtgac tgggtcatgg
     3661 ctgcgccccg acacccgcca acacccgctg acgcgccctg acgggcttgt ctgctcccgg
     3721 catccgctta cagacaagct gtgaccgtct ccgggagctg catgtgtcag aggttttcac
     3781 cgtcatcacc gaaacgcgcg aggcagctgc ggtaaagctc atcagcgtgg tcgtgaagcg
     3841 attcacagat gtctgcctgt tcatccgcgt ccagctcgtt gagtttctcc agaagcgtta
     3901 atgtctggct tctgataaag cgggccatgt taagggcggt tttttcctgt ttggtcactg
     3961 atgcctccgt gtaaggggga tttctgttca tgggggtaat gataccgatg aaacgagaga
     4021 ggatgctcac gatacgggtt actgatgatg aacatgcccg gttactggaa cgttgtgagg
     4081 gtaaacaact ggcggtatgg atgcggcggg accagagaaa aatcactcag ggtcaatgcc
     4141 agcgcttcgt taatacagat gtaggtgttc cacagggtag ccagcagcat cctgcgatgc
     4201 agatccggaa cataatggtg cagggcgctg acttccgcgt ttccagactt tacgaaacac
     4261 ggaaaccgaa gaccattcat gttgttgctc aggtcgcaga cgttttgcag cagcagtcgc
     4321 ttcacgttcg ctcgcgtatc ggtgattcat tctgctaacc agtaaggcaa ccccgccagc
     4381 ctagccgggt cctcaacgac aggagcacga tcatgcgcac ccgtggggcc gccatgccgg
     4441 cgataatggc ctgcttctcg ccgaaacgtt tggtggcggg accagtgacg aaggcttgag
     4501 cgagggcgtg caagattccg aataccgcaa gcgacaggcc gatcatcgtc gcgctccagc
     4561 gaaagcggtc ctcgccgaaa atgacccaga gcgctgccgg cacctgtcct acgagttgca
     4621 tgataaagaa gacagtcata agtgcggcga cgatagtcat gccccgcgcc caccggaagg
     4681 agctgactgg gttgaaggct ctcaagggca tcggtcgaga tcccggtgcc taatgagtga
     4741 gctaacttac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt
     4801 gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgcc
     4861 agggtggttt ttcttttcac cagtgagacg ggcaacagct gattgccctt caccgcctgg
     4921 ccctgagaga gttgcagcaa gcggtccacg ctggtttgcc ccagcaggcg aaaatcctgt
     4981 ttgatggtgg ttaacggcgg gatataacat gagctgtctt cggtatcgtc gtatcccact
     5041 accgagatat ccgcaccaac gcgcagcccg gactcggtaa tggcgcgcat tgcgcccagc
     5101 gccatctgat cgttggcaac cagcatcgca gtgggaacga tgccctcatt cagcatttgc
     5161 atggtttgtt gaaaaccgga catggcactc cagtcgcctt cccgttccgc tatcggctga
     5221 atttgattgc gagtgagata tttatgccag ccagccagac gcagacgcgc cgagacagaa
     5281 cttaatgggc ccgctaacag cgcgatttgc tggtgaccca atgcgaccag atgctccacg
     5341 cccagtcgcg taccgtcttc atgggagaaa ataatactgt tgatgggtgt ctggtcagag
     5401 acatcaagaa ataacgccgg aacattagtg caggcagctt ccacagcaat ggcatcctgg
     5461 tcatccagcg gatagttaat gatcagccca ctgacgcgtt gcgcgagaag attgtgcacc
     5521 gccgctttac aggcttcgac gccgcttcgt tctaccatcg acaccaccac gctggcaccc
     5581 agttgatcgg cgcgagattt aatcgccgcg acaatttgcg acggcgcgtg cagggccaga
     5641 ctggaggtgg caacgccaat cagcaacgac tgtttgcccg ccagttgttg tgccacgcgg
     5701 ttgggaatgt aattcagctc cgccatcgcc gcttccactt tttcccgcgt tttcgcagaa
     5761 acgtggctgg cctggttcac cacgcgggaa acggtctgat aagagacacc ggcatactct
     5821 gcgacatcgt ataacgttac tggtttcaca ttcaccaccc tgaattgact ctcttccggg
     5881 cgctatcatg ccataccgcg aaaggttttg cgccattcga tggtgtccgg gatctcgacg
     5941 ctctccctta tgcgactcct gcattaggaa gcagcccagt agtaggttga ggccgttgag
     6001 caccgccgcc gcaaggaatg gtgcatgcaa ggagatggcg cccaacagtc ccccggccac
     6061 ggggcctgcc accataccca cgccgaaaca agcgctcatg agcccgaagt ggcgagcccg
     6121 atcttcccca tcggtgatgt cggcgatata ggcgccagca accgcacctg tggcgccggt
     6181 gatgccggcc acgatgcgtc cggcgtagag gatcgagatc tcgatcccgc gaaat
//
