LOCUS       sequence-349860-        5340 bp    DNA     circular SYN 14-APR-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5340)
  TITLE     LIC N-terminal fusion vectors for E. coli expression
REFERENCE   2  (bases 1 to 5340)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: pET His6 TEV LIC cloning vector (1B)
FEATURES             Location/Qualifiers
     source          1..5340
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     959..978
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(998..1075)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1281..1300
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     1384..1403
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1384..1402
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1403..1427
                     /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             1442..1464
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1483..1500
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1501..1521
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             1587..1604
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1657..1675)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1671..1718
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1755..2210
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1842..1861)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2052..2073
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2303..3118)
                     /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(2303..3118)
                     /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     3026..3045
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3110..3129
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      3240..3828
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3729..3748
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3982..3999
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4014..4153
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4079..4098)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4239..4261
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4255..4446)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(5255..5340,1..997))
                     /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"
ORIGIN
        1 tgggcgccag ggtggttttt cttttcacca gtgagacggg caacagctga ttgcccttca
       61 ccgcctggcc ctgagagagt tgcagcaagc ggtccacgct ggtttgcccc agcaggcgaa
      121 aatcctgttt gatggtggtt aacggcggga tataacatga gctgtcttcg gtatcgtcgt
      181 atcccactac cgagatatcc gcaccaacgc gcagcccgga ctcggtaatg gcgcgcattg
      241 cgcccagcgc catctgatcg ttggcaacca gcatcgcagt gggaacgata ccctcattca
      301 gcatttgcat ggtttgttga aaaccggaca tggcactcca gtcgccttcc cgttccgcta
      361 tcggctgaat ttgattgcga gtgagatatt tatgccagcc agccagacgc agacgcgccg
      421 agacagaact taatgggccc gctaacagcg cgatttgctg gtgacccaat gcgaccagat
      481 gctccacgcc cagtcgcgta ccgtcttcat gggagaaaat aatactgttg atgggtgtct
      541 ggtcagagac atcaagaaat aacgccggaa cattagtgca ggcagcttcc acagcaatgg
      601 catcctggtc atccagcgga tagttaatga tcagcccact gacgcgttgc gcgagaagat
      661 tgtgcaccgc cgctttacag gcttcgacgc cgcttcgttc taccatcgac accaccacgc
      721 tggcacccag ttgatcggcg cgagatttaa tcgccgcgac aatttgcgac ggcgcgtgca
      781 gggccagact ggaggtggca acgccaatca gcaacgactg tttgcccgcc agttgttgtg
      841 ccacgcggtt gggaatgtaa ttcagctccg ccatcgccgc ttccactttt tcccgcgttt
      901 tcgcagaaac gtggctggcc tggttcacca cgcgggaaac ggtctgataa gagacaccgg
      961 catactctgc gacatcgtat aacgttactg gtttcacatt caccaccctg aattgactct
     1021 cttccgggcg ctatcatgcc ataccgcgaa aggttttgcg ccattcgatg gtgtccggga
     1081 tctcgacgct ctcccttatg cgactcctgc attaggaagc agcccagtag taggttgagg
     1141 ccgttgagca ccgccgccgc aaggaatggt gcatgcaagg agatggcgcc caacagtccc
     1201 ccggccacgg ggcctgccac catacccacg ccgaaacaag cgctcatgag cccgaagtgg
     1261 cgagcccgat cttccccatc ggtgatgtcg gcgatatagg cgccagcaac cgcacctgtg
     1321 gcgccggtga tgccggccac gatgcgtccg gcgtagagga tcgagatctc gatcccgcga
     1381 aattaatacg actcactata ggggaattgt gagcggataa caattcccct ctagaaataa
     1441 ttttgtttaa ctttaagaag gagatatacc atgggttctt ctcaccatca ccatcaccat
     1501 gaaaacctgt acttccaatc caatattgga agtggataac ggatccgaat tcgagcgccg
     1561 tcgacaagct tgcggccgca ctcgagcacc accaccacca ccactgagat ccggctgcta
     1621 acaaagcccg aaaggaagct gagttggctg ctgccaccgc tgagcaataa ctagcataac
     1681 cccttggggc ctctaaacgg gtcttgaggg gttttttgct gaaaggagga actatatccg
     1741 gattggcgaa tgggacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac
     1801 gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc
     1861 ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt
     1921 agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg
     1981 ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac
     2041 gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta
     2101 ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat
     2161 ttaacaaaaa tttaacgcga attttaacaa actagtaacg tttacaattt caggtggcac
     2221 ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat
     2281 gtatccgctc atgaattaat tcttagaaaa actcatcgag catcaaatga aactgcaatt
     2341 tattcatatc aggattatca ataccatatt tttgaaaaag ccgtttctgt aatgaaggag
     2401 aaaactcacc gaggcagttc cataggatgg caagatcctg gtatcggtct gcgattccga
     2461 ctcgtccaac atcaatacaa cctattaatt tcccctcgtc aaaaataagg ttatcaagtg
     2521 agaaatcacc atgagtgacg actgaatccg gtgagaatgg caaaagttta tgcatttctt
     2581 tccagacttg ttcaacaggc cagccattac gctcgtcatc aaaatcactc gcatcaacca
     2641 aaccgttatt cattcgtgat tgcgcctgag cgagacgaaa tacgcgatcg ctgttaaaag
     2701 gacaattaca aacaggaatc gaatgcaacc ggcgcaggaa cactgccagc gcatcaacaa
     2761 tgttttcacc tgaatcagga tattcttcta atacctggaa tgctgttttc ccggggatcg
     2821 cagtggtgag taaccatgca tcatcaggag tacggataaa atgcttgatg gtcggaagag
     2881 gcataaattc cgtcagccag tttagtctga ccatctcatc tgtaacatca ttggcaacgc
     2941 tacctttgcc atgtttcaga aacaactctg gcgcatcggg cttcccatac aatcgataga
     3001 ttgtcgcacc tgattgcccg acattatcgc gagcccattt atacccatat aaatcagcat
     3061 ccatgttgga atttaatcgc ggcctagagc aagacgtttc ccgttgaata tggctcataa
     3121 caccccttgt attactgttt atgtaagcag acagttttat tgttcatgac caaaatccct
     3181 taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct
     3241 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca
     3301 gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc
     3361 agcagagcgc agataccaaa tactgtcctt ctagtgtagc cgtagttagg ccaccacttc
     3421 aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct
     3481 gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag
     3541 gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc
     3601 tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg
     3661 agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag
     3721 cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt
     3781 gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac
     3841 gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg
     3901 ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc
     3961 cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga gcgcctgatg
     4021 cggtattttc tccttacgca tctgtgcggt atttcacacc gcaatggtgc actctcagta
     4081 caatctgctc tgatgccgca tagttaagcc agtatacact ccgctatcgc tacgtgactg
     4141 ggtcatggct gcgccccgac acccgccaac acccgctgac gcgccctgac gggcttgtct
     4201 gctcccggca tccgcttaca gacaagctgt gaccgtctcc gggagctgca tgtgtcagag
     4261 gttttcaccg tcatcaccga aacgcgcgag gcagctgcgg taaagctcat cagcgtggtc
     4321 gtgaagcgat tcacagatgt ctgcctgttc atccgcgtcc agctcgttga gtttctccag
     4381 aagcgttaat gtctggcttc tgataaagcg ggccatgtta agggcggttt tttcctgttt
     4441 ggtcactgat gcctccgtgt aagggggatt tctgttcatg ggggtaatga taccgatgaa
     4501 acgagagagg atgctcacga tacgggttac tgatgatgaa catgcccggt tactggaacg
     4561 ttgtgagggt aaacaactgg cggtatggat gcggcgggac cagagaaaaa tcactcaggg
     4621 tcaatgccag cgcttcgtta atacagatgt aggtgttcca cagggtagcc agcagcatcc
     4681 tgcgatgcag atccggaaca taatggtgca gggcgctgac ttccgcgttt ccagacttta
     4741 cgaaacacgg aaaccgaaga ccattcatgt tgttgctcag gtcgcagacg ttttgcagca
     4801 gcagtcgctt cacgttcgct cgcgtatcgg tgattcattc tgctaaccag taaggcaacc
     4861 ccgccagcct agccgggtcc tcaacgacag gagcacgatc atgcgcaccc gtggggccgc
     4921 catgccggcg ataatggcct gcttctcgcc gaaacgtttg gtggcgggac cagtgacgaa
     4981 ggcttgagcg agggcgtgca agattccgaa taccgcaagc gacaggccga tcatcgtcgc
     5041 gctccagcga aagcggtcct cgccgaaaat gacccagagc gctgccggca cctgtcctac
     5101 gagttgcatg ataaagaaga cagtcataag tgcggcgacg atagtcatgc cccgcgccca
     5161 ccggaaggag ctgactgggt tgaaggctct caagggcatc ggtcgagatc ccggtgccta
     5221 atgagtgagc taacttacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa
     5281 cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat
//
