LOCUS       sequence-150209-        6496 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression for structure determination; may not be full 
            ORF.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6496)
  TITLE     SGC 2016 plasmids
REFERENCE   2  (bases 1 to 6496)
  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: GALTA-c001
FEATURES             Location/Qualifiers
     source          1..6496
                     /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             92..109
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             134..154
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             1357..1374
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1427..1445)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1441..1488
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1525..1980
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1612..1631)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1822..1843
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2073..2888)
                     /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(2073..2888)
                     /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     2796..2815
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2880..2899
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      3010..3598
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3499..3518
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3752..3769
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3784..3926
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3852..3871)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4012..4034
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4028..4219)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5028..6110)
                     /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     6072..6091
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6111..6188)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6394..6413
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
       61 tgtttaactt taagaaggag atatacatat gcaccatcat catcatcatt cttctggtgt
      121 agatctgggt accgagaacc tgtacttcca atccatgtcg cgcagtggaa ccgatcctca
      181 gcaacgccag caggcgtcag aggcggacgc cgcagcagca accttccggg caaacgacca
      241 tcagcatatc cgctacaacc cgctgcagga tgagtgggtg ctggtgtcag ctcaccgcat
      301 gaagcggccc tggcagggtc aagtggagcc ccagcttctg aagacagtgc cccgccatga
      361 ccctctcaac cctctgtgtc ctggggccat ccgagccaac ggagaggtga atccccagta
      421 cgatagcacc ttcctgtttg acaacgactt cccagctctg cagcctgatg cccccagtcc
      481 aggacccagt gatcatcccc ttttccaagc aaagtctgct cgaggagtct gtaaggtcat
      541 gtgcttccac ccctggtcgg atgtaacgct gccactcatg tcggtccctg agatccgggc
      601 tgttgttgat gcatgggcct cagtcacaga ggagctgggt gcccagtacc cttgggtgca
      661 gatctttgaa aacaaaggtg ccatgatggg ctgttctaac ccccaccccc actgccaggt
      721 atgggccagc agtttcctgc cagatattgc ccagcgtgag gagcgatctc agcaggccta
      781 taagagtcag catggagagc ccctgctaat ggagtacagc cgccaggagc tactcaggaa
      841 ggaacgtctg gtcctaacca gtgagcactg gttagtactg gtccccttct gggcaacatg
      901 gccctaccag acactgctgc tgccccgtcg gcatgtgcgg cggctacctg agctgacccc
      961 tgctgagcgt gatgatctag cctccatcat gaagaagctc ttgaccaagt atgacaacct
     1021 ctttgagacg tcctttccct actccatggg ctggcatggg gctcccacag gatcagaggc
     1081 tggggccaac tgggaccatt ggcagctgca cgctcattac taccctccgc tcctgcgctc
     1141 tgccactgtc cggaaattca tggttggcta cgaaatgctt gctcaggctc agagggacct
     1201 cacccctgag caggctgcag agagactaag ggcacttcct gaggttcatt accacctggg
     1261 gcagaaggac agggagacag caaccatcgc ctgacagtaa aggtggatac ggatccgaat
     1321 tcgagctccg tcgacaagct tgcggccgca ctcgagcacc accaccacca ccactgagat
     1381 ccggctgcta acaaagcccg aaaggaagct gagttggctg ctgccaccgc tgagcaataa
     1441 ctagcataac cccttggggc ctctaaacgg gtcttgaggg gttttttgct gaaaggagga
     1501 actatatccg gattggcgaa tgggacgcgc cctgtagcgg cgcattaagc gcggcgggtg
     1561 tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg
     1621 ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg
     1681 ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt
     1741 agggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt
     1801 tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta
     1861 tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa
     1921 atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg tttacaattt
     1981 caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
     2041 attcaaatat gtatccgctc atgaattaat tcttagaaaa actcatcgag catcaaatga
     2101 aactgcaatt tattcatatc aggattatca ataccatatt tttgaaaaag ccgtttctgt
     2161 aatgaaggag aaaactcacc gaggcagttc cataggatgg caagatcctg gtatcggtct
     2221 gcgattccga ctcgtccaac atcaatacaa cctattaatt tcccctcgtc aaaaataagg
     2281 ttatcaagtg agaaatcacc atgagtgacg actgaatccg gtgagaatgg caaaagttta
     2341 tgcatttctt tccagacttg ttcaacaggc cagccattac gctcgtcatc aaaatcactc
     2401 gcatcaacca aaccgttatt cattcgtgat tgcgcctgag cgagacgaaa tacgcgatcg
     2461 ctgttaaaag gacaattaca aacaggaatc gaatgcaacc ggcgcaggaa cactgccagc
     2521 gcatcaacaa tattttcacc tgaatcagga tattcttcta atacctggaa tgctgttttc
     2581 ccggggatcg cagtggtgag taaccatgca tcatcaggag tacggataaa atgcttgatg
     2641 gtcggaagag gcataaattc cgtcagccag tttagtctga ccatctcatc tgtaacatca
     2701 ttggcaacgc tacctttgcc atgtttcaga aacaactctg gcgcatcggg cttcccatac
     2761 aatcgataga ttgtcgcacc tgattgcccg acattatcgc gagcccattt atacccatat
     2821 aaatcagcat ccatgttgga atttaatcgc ggcctagagc aagacgtttc ccgttgaata
     2881 tggctcataa caccccttgt attactgttt atgtaagcag acagttttat tgttcatgac
     2941 caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa
     3001 aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc
     3061 accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt
     3121 aactggcttc agcagagcgc agataccaaa tactgtcctt ctagtgtagc cgtagttagg
     3181 ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc
     3241 agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt
     3301 accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga
     3361 gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct
     3421 tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg
     3481 cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca
     3541 cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa
     3601 cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt
     3661 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga
     3721 taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga
     3781 gcgcctgatg cggtattttc tccttacgca tctgtgcggt atttcacacc gcatatatgg
     3841 tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagtatac actccgctat
     3901 cgctacgtga ctgggtcatg gctgcgcccc gacacccgcc aacacccgct gacgcgccct
     3961 gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct
     4021 gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gaggcagctg cggtaaagct
     4081 catcagcgtg gtcgtgaagc gattcacaga tgtctgcctg ttcatccgcg tccagctcgt
     4141 tgagtttctc cagaagcgtt aatgtctggc ttctgataaa gcgggccatg ttaagggcgg
     4201 ttttttcctg tttggtcact gatgcctccg tgtaaggggg atttctgttc atgggggtaa
     4261 tgataccgat gaaacgagag aggatgctca cgatacgggt tactgatgat gaacatgccc
     4321 ggttactgga acgttgtgag ggtaaacaac tggcggtatg gatgcggcgg gaccagagaa
     4381 aaatcactca gggtcaatgc cagcgcttcg ttaatacaga tgtaggtgtt ccacagggta
     4441 gccagcagca tcctgcgatg cagatccgga acataatggt gcagggcgct gacttccgcg
     4501 tttccagact ttacgaaaca cggaaaccga agaccattca tgttgttgct caggtcgcag
     4561 acgttttgca gcagcagtcg cttcacgttc gctcgcgtat cggtgattca ttctgctaac
     4621 cagtaaggca accccgccag cctagccggg tcctcaacga caggagcacg atcatgcgca
     4681 cccgtggggc cgccatgccg gcgataatgg cctgcttctc gccgaaacgt ttggtggcgg
     4741 gaccagtgac gaaggcttga gcgagggcgt gcaagattcc gaataccgca agcgacaggc
     4801 cgatcatcgt cgcgctccag cgaaagcggt cctcgccgaa aatgacccag agcgctgccg
     4861 gcacctgtcc tacgagttgc atgataaaga agacagtcat aagtgcggcg acgatagtca
     4921 tgccccgcgc ccaccggaag gagctgactg ggttgaaggc tctcaagggc atcggtcgag
     4981 atcccggtgc ctaatgagtg agctaactta cattaattgc gttgcgctca ctgcccgctt
     5041 tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag
     5101 gcggtttgcg tattgggcgc cagggtggtt tttcttttca ccagtgagac gggcaacagc
     5161 tgattgccct tcaccgcctg gccctgagag agttgcagca agcggtccac gctggtttgc
     5221 cccagcaggc gaaaatcctg tttgatggtg gttaacggcg ggatataaca tgagctgtct
     5281 tcggtatcgt cgtatcccac taccgagata tccgcaccaa cgcgcagccc ggactcggta
     5341 atggcgcgca ttgcgcccag cgccatctga tcgttggcaa ccagcatcgc agtgggaacg
     5401 atgccctcat tcagcatttg catggtttgt tgaaaaccgg acatggcact ccagtcgcct
     5461 tcccgttccg ctatcggctg aatttgattg cgagtgagat atttatgcca gccagccaga
     5521 cgcagacgcg ccgagacaga acttaatggg cccgctaaca gcgcgatttg ctggtgaccc
     5581 aatgcgacca gatgctccac gcccagtcgc gtaccgtctt catgggagaa aataatactg
     5641 ttgatgggtg tctggtcaga gacatcaaga aataacgccg gaacattagt gcaggcagct
     5701 tccacagcaa tggcatcctg gtcatccagc ggatagttaa tgatcagccc actgacgcgt
     5761 tgcgcgagaa gattgtgcac cgccgcttta caggcttcga cgccgcttcg ttctaccatc
     5821 gacaccacca cgctggcacc cagttgatcg gcgcgagatt taatcgccgc gacaatttgc
     5881 gacggcgcgt gcagggccag actggaggtg gcaacgccaa tcagcaacga ctgtttgccc
     5941 gccagttgtt gtgccacgcg gttgggaatg taattcagct ccgccatcgc cgcttccact
     6001 ttttcccgcg ttttcgcaga aacgtggctg gcctggttca ccacgcggga aacggtctga
     6061 taagagacac cggcatactc tgcgacatcg tataacgtta ctggtttcac attcaccacc
     6121 ctgaattgac tctcttccgg gcgctatcat gccataccgc gaaaggtttt gcgccattcg
     6181 atggtgtccg ggatctcgac gctctccctt atgcgactcc tgcattagga agcagcccag
     6241 tagtaggttg aggccgttga gcaccgccgc cgcaaggaat ggtgcatgca aggagatggc
     6301 gcccaacagt cccccggcca cggggcctgc caccataccc acgccgaaac aagcgctcat
     6361 gagcccgaag tggcgagccc gatcttcccc atcggtgatg tcggcgatat aggcgccagc
     6421 aaccgcacct gtggcgccgg tgatgccggc cacgatgcgt ccggcgtaga ggatcgagat
     6481 ctcgatcccg cgaaat
//
