LOCUS Exported 6012 bp ds-DNA circular SYN 19-JUL-2021 DEFINITION Encodes human Ran protein. ACCESSION . VERSION . KEYWORDS pET28a-His6-Ran(Q69L) SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6012) TITLE Zeng Lab Plasmids REFERENCE 2 (bases 1 to 6012) AUTHORS . TITLE Direct Submission JOURNAL Exported Jul 19, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6012 /organism="synthetic DNA construct" /mol_type="other DNA" RBS 29..51 /note="efficient ribosome binding site from bacteriophage T7 gene 10 (Olins and Rangwala, 1989)" CDS 70..87 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" CDS 97..114 /codon_start=1 /product="thrombin recognition and cleavage site" /label=thrombin site /translation="LVPRGS" CDS 118..150 /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 847..864 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" primer_bind complement(917..935) /label=T7 Term /note="T7 terminator, reverse primer" terminator 931..978 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" rep_origin 1015..1470 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(1102..1121) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 1312..1333 /label=F1ori-F /note="F1 origin, forward primer" CDS complement(1562..2377) /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 2285..2304 /label=Kan-R /note="Kanamycin resistance gene, reverse primer" primer_bind 2369..2388 /label=pENTR-R /note="pENTR vectors, reverse primer" rep_origin 2499..3087 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 2988..3007 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 3241..3258 /label=L4440 /note="L4440 vector, forward primer" misc_feature 3273..3412 /label=bom /note="basis of mobility region from pBR322" primer_bind complement(3338..3357) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 3498..3520 /label=pGEX 3' /note="pGEX vectors, reverse primer" CDS complement(3514..3705) /codon_start=1 /gene="rop" /product="Rop protein, which maintains plasmids at low copy number" /label=rop /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA DELYRSCLARFGDDGENL" CDS complement(4514..5596) /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 5558..5577 /label=LacI-R /note="LacI, reverse primer" promoter complement(5597..5674) /gene="lacI" /label=lacI promoter primer_bind 5880..5899 /label=pBRrevBam /note="pBR322 vectors, tet region, downstream of BamHI, reverse primer" primer_bind 5983..6002 /label=T7 /note="T7 promoter, forward primer" promoter 5983..6001 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" protein_bind 6002..14 /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)." ORIGIN 1 cggataacaa ttcccctcta gaaataattt tgtttaactt taagaaggag atataccatg 61 ggcagcagcc atcatcatca tcatcacagc agcggcctgg tgccgcgcgg cagccatatg 121 gctagcatga ctggtggaca gcaaatgggt cgcggatccg aattcgagct ccgtcgacgc 181 atggctgcgc agggagagcc ccaggtccag ttcaaacttg tattggttgg tgatggtggt 241 actggaaaaa cgaccttcgt gaaacgtcat ttgactggtg aatttgagaa gaagtatgta 301 gccaccttgg gtgttgaggt tcatccccta gtgttccaca ccaacagagg acctattaag 361 ttcaatgtat gggacacagc cggcctcgag aaattcggtg gactgagaga tggctattat 421 atccaagccc agtgtgccat cataatgttt gatgtaacat cgagagttac ttacaagaat 481 gtgcctaact ggcatagaga tctggtacga gtgtgtgaaa acatccccat tgtgttgtgt 541 ggcaacaaag tggatattaa ggacaggaaa gtgaaggcga aatccattgt cttccaccga 601 aagaagaatc ttcagtacta cgacatttct gccaaaagta actacaactt tgaaaagccc 661 ttcctctggc ttgctaggaa gctcattgga gaccctaact tggaatttgt tgccatgcct 721 gctctcgccc caccagaagt tgtcatggac ccagctttgg cagcacagta tgagcacgac 781 ttagaggttg ctcagacaac tactctcccg gatgaggatg atgacctgtg agcggccgca 841 ctcgagcacc accaccacca ccactgagat ccggctgcta acaaagcccg aaaggaagct 901 gagttggctg ctgccaccgc tgagcaataa ctagcataac cccttggggc ctctaaacgg 961 gtcttgaggg gttttttgct gaaaggagga actatatccg gattggcgaa tgggacgcgc 1021 cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac 1081 ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg 1141 ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt 1201 tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt gggccatcgc 1261 cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct 1321 tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat ttataaggga 1381 ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga 1441 attttaacaa aatattaacg cttacaattt aggtggcact tttcggggaa atgtgcgcgg 1501 aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca tgaattaatt 1561 cttagaaaaa ctcatcgagc atcaaatgaa actgcaattt attcatatca ggattatcaa 1621 taccatattt ttgaaaaagc cgtttctgta atgaaggaga aaactcaccg aggcagttcc 1681 ataggatggc aagatcctgg tatcggtctg cgattccgac tcgtccaaca tcaatacaac 1741 ctattaattt cccctcgtca aaaataaggt tatcaagtga gaaatcacca tgagtgacga 1801 ctgaatccgg tgagaatggc aaaagtttat gcatttcttt ccagacttgt tcaacaggcc 1861 agccattacg ctcgtcatca aaatcactcg catcaaccaa accgttattc attcgtgatt 1921 gcgcctgagc gagacgaaat acgcgatcgc tgttaaaagg acaattacaa acaggaatcg 1981 aatgcaaccg gcgcaggaac actgccagcg catcaacaat attttcacct gaatcaggat 2041 attcttctaa tacctggaat gctgttttcc cggggatcgc agtggtgagt aaccatgcat 2101 catcaggagt acggataaaa tgcttgatgg tcggaagagg cataaattcc gtcagccagt 2161 ttagtctgac catctcatct gtaacatcat tggcaacgct acctttgcca tgtttcagaa 2221 acaactctgg cgcatcgggc ttcccataca atcgatagat tgtcgcacct gattgcccga 2281 cattatcgcg agcccattta tacccatata aatcagcatc catgttggaa tttaatcgcg 2341 gcctagagca agacgtttcc cgttgaatat ggctcataac accccttgta ttactgttta 2401 tgtaagcaga cagttttatt gttcatgacc aaaatccctt aacgtgagtt ttcgttccac 2461 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc 2521 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat 2581 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat 2641 actgtccttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct 2701 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt 2761 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg 2821 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta 2881 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg 2941 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg 3001 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc 3061 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg 3121 gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat 3181 aaccgtatta ccgcctttga gtgagctgat accgctcgcc gcagccgaac gaccgagcgc 3241 agcgagtcag tgagcgagga agcggaagag cgcctgatgc ggtattttct ccttacgcat 3301 ctgtgcggta tttcacaccg caatggtgca ctctcagtac aatctgctct gatgccgcat 3361 agttaagcca gtatacactc cgctatcgct acgtgactgg gtcatggctg cgccccgaca 3421 cccgccaaca cccgctgacg cgccctgacg ggcttgtctg ctcccggcat ccgcttacag 3481 acaagctgtg accgtctccg ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa 3541 acgcgcgagg cagctgcggt aaagctcatc agcgtggtcg tgaagcgatt cacagatgtc 3601 tgcctgttca tccgcgtcca gctcgttgag tttctccaga agcgttaatg tctggcttct 3661 gataaagcgg gccatgttaa gggcggtttt ttcctgtttg gtcactgatg cctccgtgta 3721 agggggattt ctgttcatgg gggtaatgat accgatgaaa cgagagagga tgctcacgat 3781 acgggttact gatgatgaac atgcccggtt actggaacgt tgtgagggta aacaactggc 3841 ggtatggatg cggcgggacc agagaaaaat cactcagggt caatgccagc gcttcgttaa 3901 tacagatgta ggtgttccac agggtagcca gcagcatcct gcgatgcaga tccggaacat 3961 aatggtgcag ggcgctgact tccgcgtttc cagactttac gaaacacgga aaccgaagac 4021 cattcatgtt gttgctcagg tcgcagacgt tttgcagcag cagtcgcttc acgttcgctc 4081 gcgtatcggt gattcattct gctaaccagt aaggcaaccc cgccagccta gccgggtcct 4141 caacgacagg agcacgatca tgcgcacccg tggggccgcc atgccggcga taatggcctg 4201 cttctcgccg aaacgtttgg tggcgggacc agtgacgaag gcttgagcga gggcgtgcaa 4261 gattccgaat accgcaagcg acaggccgat catcgtcgcg ctccagcgaa agcggtcctc 4321 gccgaaaatg acccagagcg ctgccggcac ctgtcctacg agttgcatga taaagaagac 4381 agtcataagt gcggcgacga tagtcatgcc ccgcgcccac cggaaggagc tgactgggtt 4441 gaaggctctc aagggcatcg gtcgagatcc cggtgcctaa tgagtgagct aacttacatt 4501 aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta 4561 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgccagg gtggtttttc 4621 ttttcaccag tgagacgggc aacagctgat tgcccttcac cgcctggccc tgagagagtt 4681 gcagcaagcg gtccacgctg gtttgcccca gcaggcgaaa atcctgtttg atggtggtta 4741 acggcgggat ataacatgag ctgtcttcgg tatcgtcgta tcccactacc gagatatccg 4801 caccaacgcg cagcccggac tcggtaatgg cgcgcattgc gcccagcgcc atctgatcgt 4861 tggcaaccag catcgcagtg ggaacgatgc cctcattcag catttgcatg gtttgttgaa 4921 aaccggacat ggcactccag tcgccttccc gttccgctat cggctgaatt tgattgcgag 4981 tgagatattt atgccagcca gccagacgca gacgcgccga gacagaactt aatgggcccg 5041 ctaacagcgc gatttgctgg tgacccaatg cgaccagatg ctccacgccc agtcgcgtac 5101 cgtcttcatg ggagaaaata atactgttga tgggtgtctg gtcagagaca tcaagaaata 5161 acgccggaac attagtgcag gcagcttcca cagcaatggc atcctggtca tccagcggat 5221 agttaatgat cagcccactg acgcgttgcg cgagaagatt gtgcaccgcc gctttacagg 5281 cttcgacgcc gcttcgttct accatcgaca ccaccacgct ggcacccagt tgatcggcgc 5341 gagatttaat cgccgcgaca atttgcgacg gcgcgtgcag ggccagactg gaggtggcaa 5401 cgccaatcag caacgactgt ttgcccgcca gttgttgtgc cacgcggttg ggaatgtaat 5461 tcagctccgc catcgccgct tccacttttt cccgcgtttt cgcagaaacg tggctggcct 5521 ggttcaccac gcgggaaacg gtctgataag agacaccggc atactctgcg acatcgtata 5581 acgttactgg tttcacattc accaccctga attgactctc ttccgggcgc tatcatgcca 5641 taccgcgaaa ggttttgcgc cattcgatgg tgtccgggat ctcgacgctc tcccttatgc 5701 gactcctgca ttaggaagca gcccagtagt aggttgaggc cgttgagcac cgccgccgca 5761 aggaatggtg catgcaagga gatggcgccc aacagtcccc cggccacggg gcctgccacc 5821 atacccacgc cgaaacaagc gctcatgagc ccgaagtggc gagcccgatc ttccccatcg 5881 gtgatgtcgg cgatataggc gccagcaacc gcacctgtgg cgccggtgat gccggccacg 5941 atgcgtccgg cgtagaggat cgagatctcg atcccgcgaa attaatacga ctcactatag 6001 gggaattgtg ag //