pET101/D-TOPO

LOCUS       pET101-D-TOPO               5753 bp ds-DNA     circular SYN 15-AUG-2016
DEFINITION  synthetic circular DNA
KEYWORDS    pET101-D-TOPO
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5753)
  AUTHORS   Biofeng Lab.
FEATURES             Location/Qualifiers
     source          1..5753
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     promoter        209..227
                     /note="T7 promoter"
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    228..252
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="lac operator"
                     /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)."
     misc_feature    282..296
                     /note="RBS"
     misc_feature    282..288
                     /note="RBS"
     misc_feature    293..296
                     /note="RBS"
     misc_feature    297..310
                     /note="TOPO site"
     CDS             333..374
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /note="V5 tag"
                     /translation="GKPIPNPLLGLDST"
     CDS             384..401
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /note="6xHis"
                     /translation="HHHHHH"
     terminator      416..544
                     /note="T7 terminator"
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(694..722)
                     /note="tet promoter"
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     promoter        845..943
                     /gene="bla"
                     /note="bla promoter"
     CDS             944..1804
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /note="AmpR"
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      1949..2622
                     /direction=RIGHT
                     /note="pBR322"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     misc_feature    2749..2888
                     /note="bom"
                     /note="basis of mobility region from pBR322"
     CDS             complement(2990..3181)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /note="rop"
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4493..5584)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /note="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="MVNVKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAM
                     AELNYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVER
                     SGVEACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSII
                     FSHEDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGD
                     WSAMSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTED
                     SSCYIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTA
                     SPRALADSLMQLARQVSRLESGQ"
     promoter        complement(5576..5653)
                     /gene="lacI"
                     /note="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 agaaggaatt caggagccct
      301 tcaccaaggg cgagctcaat tcgaagcttg aaggtaagcc tatccctaac cctctcctcg
      361 gtctcgattc tacgcgtacc ggtcatcatc accatcacca ttgagtttga tccggctgct
      421 aacaaagccc gaaaggaagc tgagttggct gctgccaccg ctgagcaata actagcataa
      481 ccccttgggg cctctaaacg ggtcttgagg ggttttttgc tgaaaggagg aactatatcc
      541 ggatatcccg caagaggccc ggcagtaccg gcataaccaa gcctatgcct acagcatcca
      601 gggtgacggt gccgaggatg acgatgagcg cattgttaga tttcatacac ggtgcctgac
      661 tgcgttagca atttaactgt gataaactac cgcattaaag cttatcgatg ataagctgtc
      721 aaacatgaga attaattctt gaagacgaaa gggcctcgtg atacgcctat ttttataggt
      781 taatgtcatg ataataatgg tttcttagac gtcaggtggc acttttcggg gaaatgtgcg
      841 cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca
      901 ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt
      961 ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga
     1021 aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga
     1081 actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat
     1141 gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtgttg acgccgggca
     1201 agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt
     1261 cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac
     1321 catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct
     1381 aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga
     1441 gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgcag caatggcaac
     1501 aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat
     1561 agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg
     1621 ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc
     1681 actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc
     1741 aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg
     1801 gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta
     1861 atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg
     1921 tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     1981 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     2041 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     2101 agcgcagata ccaaatactg tccttctagt gtagccgtag ttaggccacc acttcaagaa
     2161 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     2221 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     2281 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     2341 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
     2401 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     2461 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     2521 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     2581 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
     2641 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
     2701 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc tgatgcggta
     2761 ttttctcctt acgcatctgt gcggtatttc acaccgcaat ggtgcactct cagtacaatc
     2821 tgctctgatg ccgcatagtt aagccagtat acactccgct atcgctacgt gactgggtca
     2881 tggctgcgcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc
     2941 cggcatccgc ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt
     3001 caccgtcatc accgaaacgc gcgaggcagc tgcggtaaag ctcatcagcg tggtcgtgaa
     3061 gcgattcaca gatgtctgcc tgttcatccg cgtccagctc gttgagtttc tccagaagcg
     3121 ttaatgtctg gcttctgata aagcgggcca tgttaagggc ggttttttcc tgtttggtca
     3181 ctgatgcctc cgtgtaaggg ggatttctgt tcatgggggt aatgataccg atgaaacgag
     3241 agaggatgct cacgatacgg gttactgatg atgaacatgc ccggttactg gaacgttgtg
     3301 agggtaaaca actggcggta tggatgcggc gggaccagag aaaaatcact cagggtcaat
     3361 gccagcgctt cgttaataca gatgtaggtg ttccacaggg tagccagcag catcctgcga
     3421 tgcagatccg gaacataatg gtgcagggcg ctgacttccg cgtttccaga ctttacgaaa
     3481 cacggaaacc gaagaccatt catgttgttg ctcaggtcgc agacgttttg cagcagcagt
     3541 cgcttcacgt tcgctcgcgt atcggtgatt cattctgcta accagtaagg caaccccgcc
     3601 agcctagccg ggtcctcaac gacaggagca cgatcatgcg cacccgtggc caggacccaa
     3661 cgctgcccga gatgcgccgc gtgcggctgc tggagatggc ggacgcgatg gatatgttct
     3721 gccaagggtt ggtttgcgca ttcacagttc tccgcaagaa ttgattggct ccaattcttg
     3781 gagtggtgaa tccgttagcg aggtgccgcc ggcttccatt caggtcgagg tggcccggct
     3841 ccatgcaccg cgacgcaacg cggggaggca gacaaggtat agggcggcgc ctacaatcca
     3901 tgccaacccg ttccatgtgc tcgccgaggc ggcataaatc gccgtgacga tcagcggtcc
     3961 aatgatcgaa gttaggctgg taagagccgc gagcgatcct tgaagctgtc cctgatggtc
     4021 gtcatctacc tgcctggaca gcatggcctg caacgcgggc atcccgatgc cgccggaagc
     4081 gagaagaatc ataatgggga aggccatcca gcctcgcgtc gcgaacgcca gcaagacgta
     4141 gcccagcgcg tcggccgcca tgccggcgat aatggcctgc ttctcgccga aacgtttggt
     4201 ggcgggacca gtgacgaagg cttgagcgag ggcgtgcaag attccgaata ccgcaagcga
     4261 caggccgatc atcgtcgcgc tccagcgaaa gcggtcctcg ccgaaaatga cccagagcgc
     4321 tgccggcacc tgtcctacga gttgcatgat aaagaagaca gtcataagtg cggcgacgat
     4381 agtcatgccc cgcgcccacc ggaaggagct gactgggttg aaggctctca agggcatcgg
     4441 tcgagatccc ggtgcctaat gagtgagcta acttacatta attgcgttgc gctcactgcc
     4501 cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg
     4561 gagaggcggt ttgcgtattg ggcgccaggg tggtttttct tttcaccagt gagacgggca
     4621 acagctgatt gcccttcacc gcctggccct gagagagttg cagcaagcgg tccacgctgg
     4681 tttgccccag caggcgaaaa tcctgtttga tggtggttaa cggcgggata taacatgagc
     4741 tgtcttcggt atcgtcgtat cccactaccg agatatccgc accaacgcgc agcccggact
     4801 cggtaatggc gcgcattgcg cccagcgcca tctgatcgtt ggcaaccagc atcgcagtgg
     4861 gaacgatgcc ctcattcagc atttgcatgg tttgttgaaa accggacatg gcactccagt
     4921 cgccttcccg ttccgctatc ggctgaattt gattgcgagt gagatattta tgccagccag
     4981 ccagacgcag acgcgccgag acagaactta atgggcccgc taacagcgcg atttgctggt
     5041 gacccaatgc gaccagatgc tccacgccca gtcgcgtacc gtcttcatgg gagaaaataa
     5101 tactgttgat gggtgtctgg tcagagacat caagaaataa cgccggaaca ttagtgcagg
     5161 cagcttccac agcaatggca tcctggtcat ccagcggata gttaatgatc agcccactga
     5221 cgcgttgcgc gagaagattg tgcaccgccg ctttacaggc ttcgacgccg cttcgttcta
     5281 ccatcgacac caccacgctg gcacccagtt gatcggcgcg agatttaatc gccgcgacaa
     5341 tttgcgacgg cgcgtgcagg gccagactgg aggtggcaac gccaatcagc aacgactgtt
     5401 tgcccgccag ttgttgtgcc acgcggttgg gaatgtaatt cagctccgcc atcgccgctt
     5461 ccactttttc ccgcgttttc gcagaaacgt ggctggcctg gttcaccacg cgggaaacgg
     5521 tctgataaga gacaccggca tactctgcga catcgtataa cgttactggt ttcacattca
     5581 ccaccctgaa ttgactctct tccgggcgct atcatgccat accgcgaaag gttttgcgcc
     5641 attcgatggt gtccgggatc tcgacgctct cccttatgcg actcctgcat taggaagcag
     5701 cccagtagta ggttgaggcc gttgagcacc gccgccgcaa ggaatggtgc atg
//