LOCUS Exported 7784 bp ds-DNA circular SYN 27-DEC-2017
DEFINITION synthetic circular DNA
FEATURES Location/Qualifiers
source 1..7784
/organism="synthetic DNA construct"
/mol_type="other DNA"
terminator 26..73
/label=T7 terminator
/note="transcription terminator for bacteriophage T7 RNA
polymerase"
CDS complement(140..157)
/codon_start=1
/product="6xHis affinity tag"
/label=6xHis
/translation="HHHHHH"
protein_bind 166..290
/gene="mutant version of attR"
/label=attR2
/bound_moiety="LR Clonase(TM)"
/note="recombination site for the Gateway(R) LR reaction"
CDS complement(331..636)
/codon_start=1
/gene="ccdB"
/product="CcdB, a bacterial toxin that poisons DNA gyrase"
/label=ccdB
/note="Plasmids containing the ccdB gene cannot be
propagated in standard E. coli strains."
/translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
CDS complement(978..1637)
/codon_start=1
/gene="cat"
/product="chloramphenicol acetyltransferase"
/label=Chl
/note="confers resistance to chloramphenicol"
/translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
promoter complement(1691..1721)
/label=lac UV5 promoter
/note="E. coli lac promoter with an ""up"" mutation"
protein_bind complement(1746..1870)
/gene="mutant version of attR"
/label=attR1
/bound_moiety="LR Clonase(TM)"
/note="recombination site for the Gateway(R) LR reaction"
CDS complement(1886..2539)
/codon_start=1
/product="glutathione S-transferase from Schistosoma
japonicum"
/label=GST
/translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
CDS complement(2558..2575)
/codon_start=1
/product="6xHis affinity tag"
/label=6xHis
/translation="HHHHHH"
RBS 2588..2610
/note="efficient ribosome binding site from bacteriophage
T7 gene 10 (Olins and Rangwala, 1989)"
protein_bind 2625..2649
/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)."
promoter complement(2650..2668)
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
promoter 2977..3054
/gene="lacI"
/label=lacI promoter
CDS 3055..4137
/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"
CDS 4946..5137
/codon_start=1
/gene="rop"
/product="Rop protein, which maintains plasmids at low copy
number"
/label=rop
/translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
DELYRSCLARFGDDGENL"
misc_feature 5239..5381
/label=bom
/note="basis of mobility region from pBR322"
rep_origin complement(5567..6155)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(6326..7186)
/codon_start=1
/gene="bla"
/product="beta-lactamase"
/label=AmpR
/note="confers resistance to ampicillin, carbenicillin, and
related antibiotics"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
promoter complement(7187..7291)
/gene="bla"
/label=AmpR promoter
rep_origin complement(7318..7773)
/direction=LEFT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
ORIGIN
1 atccggatat agttcctcct ttcagcaaaa aacccctcaa gacccgttta gaggccccaa
61 ggggttatgc tagttattgc tcagcggtgg cagcagccaa ctcagcttcc tttcgggctt
121 tgttagcagc cggatctcag tggtggtggt ggtggtgctc gagtcaccac tttgtacaag
181 aaagctgaac gagaaacgta aaatgatata aatatcaata tattaaatta gattttgcat
241 aaaaaacaga ctacataata ctgtaaaaca caacatatcc agtcactatg gtcgacctgc
301 agactggctg tgtataaggg agcctgacat ttatattccc cagaacatca ggttaatggc
361 gtttttgatg tcattttcgc ggtggctgag atcagccact tcttccccga taacggagac
421 cggcacactg gccatatcgg tggtcatcat gcgccagctt tcatccccga tatgcaccac
481 cgggtaaagt tcacgggaga ctttatctga cagcagacgt gcactggcca gggggatcac
541 catccgtcgc ccgggcgtgt caataatatc actctgtaca tccacaaaca gacgataacg
601 gctctctctt ttataggtgt aaaccttaaa ctgcatttca ccagtccctg ttctcgtcag
661 caaaagagcc gttcatttca ataaaccggg cgacctcagc catcccttcc tgattttccg
721 ctttccagcg ttcggcacgc agacgacggg cttcattctg catggttgtg cttaccagac
781 cggagatatt gacatcatat atgccttgag caactgatag ctgtcgctgt caactgtcac
841 tgtaatacgc tgcttcatag cacacctctt tttgacatac ttcgggtata catatcagta
901 tatattctta taccgcaaaa atcagcgcgc aaatacgcat actgttatct ggcttttagt
961 aagccggatc cacgcgttta cgccccgccc tgccactcat cgcagtactg ttgtaattca
1021 ttaagcattc tgccgacatg gaagccatca cagacggcat gatgaacctg aatcgccagc
1081 ggcatcagca ccttgtcgcc ttgcgtataa tatttgccca tggtgaaaac gggggcgaag
1141 aagttgtcca tattggccac gtttaaatca aaactggtga aactcaccca gggattggct
1201 gagacgaaaa acatattctc aataaaccct ttagggaaat aggccaggtt ttcaccgtaa
1261 cacgccacat cttgcgaata tatgtgtaga aactgccgga aatcgtcgtg gtattcactc
1321 cagagcgatg aaaacgtttc agtttgctca tggaaaacgg tgtaacaagg gtgaacacta
1381 tcccatatca ccagctcacc gtctttcatt gccatacgga attccggatg agcattcatc
1441 aggcgggcaa gaatgtgaat aaaggccgga taaaacttgt gcttattttt ctttacggtc
1501 tttaaaaagg ccgtaatatc cagctgaacg gtctggttat aggtacattg agcaactgac
1561 tgaaatgcct caaaatgttc tttacgatgc cattgggata tatcaacggt ggtatatcca
1621 gtgatttttt tctccatttt agcttcctta gctcctgaaa atctcgccgg atcctaactc
1681 aaaatccaca cattatacga gccggaagca taaagtgtaa agcctggggt gcctaatgcg
1741 gccgccatag tgactggata tgttgtgttt tacagtatta tgtagtctgt tttttatgca
1801 aaatctaatt taatatattg atatttatat cattttacgt ttctcgttca gcttttttgt
1861 acaaacttgt catggtacca tccgattttg gaggatggtc gccaccacca aacgtggctt
1921 gccagccctg caaaggccat gctatatact tgctggattt caagtactta tcaatttgtg
1981 ggatagcttc aatacgtttt ttaaaacaaa ctaattttgg gaacgcatcc aggcacattg
2041 ggtccatgta taaaacaaca tcaagagcgt catacaacat gaagtcagga tgggttacat
2101 gatcaccatt taaatatgtt ttatgacata aacgatcttc gaacattttc agcatttcag
2161 gtagcttgct aagaaaatca actttgagag tttcaaagtc tttactatat gcaattctcg
2221 aaacaccgta tctaatatcc aaaaccgctc cttcaagcat tgaaatctct gcacgctctt
2281 ttggacaacc acccaacatg ttgtgcttgt cagctatata acgtatgatg gccatagact
2341 gtgttaattt aacatcacca tcaatataat aaggaagatt gggaaactcc aaacccaatt
2401 caaacttttt gtttcgccat ttatcacctt catcgcgctc atacaaatgc tcttcatatt
2461 tttcttcaag atattccaaa agaagtcgag tgggttgcac aaggccctta attttccaat
2521 aacctagtat aggggacatg gaattgctac tagtgttgtg atggtgatgg tgatgtttca
2581 tggtatatct ccttcttaaa gttaaacaaa attatttcta gaggggaatt gttatccgct
2641 cacaattccc ctatagtgag tcgtattaat ttcgcgggat cgagatctcg atcctctacg
2701 ccggacgcat cgtggccggc atcaccggcg ccacaggtgc ggttgctggc gcctatatcg
2761 ccgacatcac cgatggggaa gatcgggctc gccacttcgg gctcatgagc gcttgtttcg
2821 gcgtgggtat ggtggcaggc cccgtggccg ggggactgtt gggcgccatc tccttgcatg
2881 caccattcct tgcggcggcg gtgctcaacg gcctcaacct actactgggc tgcttcctaa
2941 tgcaggagtc gcataaggga gagcgtcgag atcccggaca ccatcgaatg gcgcaaaacc
3001 tttcgcggta tggcatgata gcgcccggaa gagagtcaat tcagggtggt gaatgtgaaa
3061 ccagtaacgt tatacgatgt cgcagagtat gccggtgtct cttatcagac cgtttcccgc
3121 gtggtgaacc aggccagcca cgtttctgcg aaaacgcggg aaaaagtgga agcggcgatg
3181 gcggagctga attacattcc caaccgcgtg gcacaacaac tggcgggcaa acagtcgttg
3241 ctgattggcg ttgccacctc cagtctggcc ctgcacgcgc cgtcgcaaat tgtcgcggcg
3301 attaaatctc gcgccgatca actgggtgcc agcgtggtgg tgtcgatggt agaacgaagc
3361 ggcgtcgaag cctgtaaagc ggcggtgcac aatcttctcg cgcaacgcgt cagtgggctg
3421 atcattaact atccgctgga tgaccaggat gccattgctg tggaagctgc ctgcactaat
3481 gttccggcgt tatttcttga tgtctctgac cagacaccca tcaacagtat tattttctcc
3541 catgaagacg gtacgcgact gggcgtggag catctggtcg cattgggtca ccagcaaatc
3601 gcgctgttag cgggcccatt aagttctgtc tcggcgcgtc tgcgtctggc tggctggcat
3661 aaatatctca ctcgcaatca aattcagccg atagcggaac gggaaggcga ctggagtgcc
3721 atgtccggtt ttcaacaaac catgcaaatg ctgaatgagg gcatcgttcc cactgcgatg
3781 ctggttgcca acgatcagat ggcgctgggc gcaatgcgcg ccattaccga gtccgggctg
3841 cgcgttggtg cggatatctc ggtagtggga tacgacgata ccgaagacag ctcatgttat
3901 atcccgccgt taaccaccat caaacaggat tttcgcctgc tggggcaaac cagcgtggac
3961 cgcttgctgc aactctctca gggccaggcg gtgaagggca atcagctgtt gcccgtctca
4021 ctggtgaaaa gaaaaaccac cctggcgccc aatacgcaaa ccgcctctcc ccgcgcgttg
4081 gccgattcat taatgcagct ggcacgacag gtttcccgac tggaaagcgg gcagtgagcg
4141 caacgcaatt aatgtaagtt agctcactca ttaggcaccg ggatctcgac cgatgccctt
4201 gagagccttc aacccagtca gctccttccg gtgggcgcgg ggcatgacta tcgtcgccgc
4261 acttatgact gtcttcttta tcatgcaact cgtaggacag gtgccggcag cgctctgggt
4321 cattttcggc gaggaccgct ttcgctggag cgcgacgatg atcggcctgt cgcttgcggt
4381 attcggaatc ttgcacgccc tcgctcaagc cttcgtcact ggtcccgcca ccaaacgttt
4441 cggcgagaag caggccatta tcgccggcat ggcggcccca cgggtgcgca tgatcgtgct
4501 cctgtcgttg aggacccggc taggctggcg gggttgcctt actggttagc agaatgaatc
4561 accgatacgc gagcgaacgt gaagcgactg ctgctgcaaa acgtctgcga cctgagcaac
4621 aacatgaatg gtcttcggtt tccgtgtttc gtaaagtctg gaaacgcgga agtcagcgcc
4681 ctgcaccatt atgttccgga tctgcatcgc aggatgctgc tggctaccct gtggaacacc
4741 tacatctgta ttaacgaagc gctggcattg accctgagtg atttttctct ggtcccgccg
4801 catccatacc gccagttgtt taccctcaca acgttccagt aaccgggcat gttcatcatc
4861 agtaacccgt atcgtgagca tcctctctcg tttcatcggt atcattaccc ccatgaacag
4921 aaatccccct tacacggagg catcagtgac caaacaggaa aaaaccgccc ttaacatggc
4981 ccgctttatc agaagccaga cattaacgct tctggagaaa ctcaacgagc tggacgcgga
5041 tgaacaggca gacatctgtg aatcgcttca cgaccacgct gatgagcttt accgcagctg
5101 cctcgcgcgt ttcggtgatg acggtgaaaa cctctgacac atgcagctcc cggagacggt
5161 cacagcttgt ctgtaagcgg atgccgggag cagacaagcc cgtcagggcg cgtcagcggg
5221 tgttggcggg tgtcggggcg cagccatgac ccagtcacgt agcgatagcg gagtgtatac
5281 tggcttaact atgcggcatc agagcagatt gtactgagag tgcaccatat atgcggtgtg
5341 aaataccgca cagatgcgta aggagaaaat accgcatcag gcgctcttcc gcttcctcgc
5401 tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg
5461 cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag
5521 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
5581 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
5641 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
5701 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
5761 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
5821 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
5881 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
5941 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
6001 ctagaaggac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
6061 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
6121 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
6181 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
6241 aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta
6301 tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag
6361 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga
6421 tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac
6481 cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc
6541 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta
6601 gttcgccagt taatagtttg cgcaacgttg ttgccattgc tgcaggcatc gtggtgtcac
6661 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat
6721 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa
6781 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg
6841 tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag
6901 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc
6961 cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct
7021 caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat
7081 cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg
7141 ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc
7201 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
7261 tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgaaa
7321 ttgtaaacgt taatattttg ttaaaattcg cgttaaattt ttgttaaatc agctcatttt
7381 ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag accgagatag
7441 ggttgagtgt tgttccagtt tggaacaaga gtccactatt aaagaacgtg gactccaacg
7501 tcaaagggcg aaaaaccgtc tatcagggcg atggcccact acgtgaacca tcaccctaat
7561 caagtttttt ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa gggagccccc
7621 gatttagagc ttgacgggga aagccggcga acgtggcgag aaaggaaggg aagaaagcga
7681 aaggagcggg cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta accaccacac
7741 ccgccgcgct taatgcgccg ctacagggcg cgtcccattc gcca
//