LOCUS Exported 5537 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Bacterial expression for structure determination; may not be full ORF. ACCESSION . VERSION . KEYWORDS 5JJZ (CDYL2) SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5537) TITLE SGC Toronto Q2 Q3 2016 REFERENCE 2 (bases 1 to 5537) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5537 /organism="synthetic DNA construct" /mol_type="other DNA" 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 94..138 /codon_start=1 /product="peptide tag that allows for enzymatic biotinylation" /label=AviTag(TM) /translation="GLNDIFEAQKIEWHE" CDS 355..372 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" CDS 402..419 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" primer_bind complement(472..490) /label=T7 Term /note="T7 terminator, reverse primer" terminator 486..533 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" rep_origin 570..1025 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(657..676) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 867..888 /label=F1ori-F /note="F1 origin, forward primer" CDS complement(1117..1932) /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 1840..1859 /label=Kan-R /note="Kanamycin resistance gene, reverse primer" primer_bind 1924..1943 /label=pENTR-R /note="pENTR vectors, reverse primer" rep_origin 2054..2642 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 2543..2562 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 2796..2813 /label=L4440 /note="L4440 vector, forward primer" misc_feature 2828..2967 /label=bom /note="basis of mobility region from pBR322" primer_bind complement(2893..2912) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 3053..3075 /label=pGEX 3' /note="pGEX vectors, reverse primer" CDS complement(3069..3260) /codon_start=1 /gene="rop" /product="Rop protein, which maintains plasmids at low copy number" /label=rop /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA DELYRSCLARFGDDGENL" CDS complement(4069..5151) /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 5113..5132 /label=LacI-R /note="LacI, reverse primer" promoter complement(5152..5229) /gene="lacI" /label=lacI promoter primer_bind 5435..5454 /label=pBRrevBam /note="pBR322 vectors, tet region, downstream of BamHI, reverse primer" ORIGIN 1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt 61 tgtttaactt taagaaggag atataccatg agcggcctga acgatatttt tgaagcgcag 121 aaaattgaat ggcatgaagg cagcgctgga ggttcaggtg gggaccttta cgaggttgaa 181 aggattgtag acaagaggaa gaacaagaaa ggaaaatggg agtatcttat ccgatggaaa 241 ggctacggga gcaccgagga cacgtgggag ccggagcacc acctcttgca ctgtgaggag 301 tttattgatg aattcaatgg gttgcacatg tccaaggaca agggtggaag tggtcatcac 361 caccatcatc actgatgacg aagcttgcgg ccgcactcga gcaccaccac caccaccact 421 gagatccggc tgctaacaaa gcccgaaagg aagctgagtt ggctgctgcc accgctgagc 481 aataactagc ataacccctt ggggcctcta aacgggtctt gaggggtttt ttgctgaaag 541 gaggaactat atccggattg gcgaatggga cgcgccctgt agcggcgcat taagcgcggc 601 gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc 661 tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa 721 tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact 781 tgattagggt gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt 841 gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa 901 ccctatctcg gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt 961 aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat taacgcttac 1021 aatttaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa 1081 atacattcaa atatgtatcc gctcatgaat taattcttag aaaaactcat cgagcatcaa 1141 atgaaactgc aatttattca tatcaggatt atcaatacca tatttttgaa aaagccgttt 1201 ctgtaatgaa ggagaaaact caccgaggca gttccatagg atggcaagat cctggtatcg 1261 gtctgcgatt ccgactcgtc caacatcaat acaacctatt aatttcccct cgtcaaaaat 1321 aaggttatca agtgagaaat caccatgagt gacgactgaa tccggtgaga atggcaaaag 1381 tttatgcatt tctttccaga cttgttcaac aggccagcca ttacgctcgt catcaaaatc 1441 actcgcatca accaaaccgt tattcattcg tgattgcgcc tgagcgagac gaaatacgcg 1501 atcgctgtta aaaggacaat tacaaacagg aatcgaatgc aaccggcgca ggaacactgc 1561 cagcgcatca acaatatttt cacctgaatc aggatattct tctaatacct ggaatgctgt 1621 tttcccgggg atcgcagtgg tgagtaacca tgcatcatca ggagtacgga taaaatgctt 1681 gatggtcgga agaggcataa attccgtcag ccagtttagt ctgaccatct catctgtaac 1741 atcattggca acgctacctt tgccatgttt cagaaacaac tctggcgcat cgggcttccc 1801 atacaatcga tagattgtcg cacctgattg cccgacatta tcgcgagccc atttataccc 1861 atataaatca gcatccatgt tggaatttaa tcgcggccta gagcaagacg tttcccgttg 1921 aatatggctc ataacacccc ttgtattact gtttatgtaa gcagacagtt ttattgttca 1981 tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga 2041 tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa 2101 aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga 2161 aggtaactgg cttcagcaga gcgcagatac caaatactgt ccttctagtg tagccgtagt 2221 taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt 2281 taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat 2341 agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct 2401 tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca 2461 cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag 2521 agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc 2581 gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga 2641 aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca 2701 tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc tttgagtgag 2761 ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg 2821 aagagcgcct gatgcggtat tttctcctta cgcatctgtg cggtatttca caccgcaatg 2881 gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagtata cactccgcta 2941 tcgctacgtg actgggtcat ggctgcgccc cgacacccgc caacacccgc tgacgcgccc 3001 tgacgggctt gtctgctccc ggcatccgct tacagacaag ctgtgaccgt ctccgggagc 3061 tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg cgaggcagct gcggtaaagc 3121 tcatcagcgt ggtcgtgaag cgattcacag atgtctgcct gttcatccgc gtccagctcg 3181 ttgagtttct ccagaagcgt taatgtctgg cttctgataa agcgggccat gttaagggcg 3241 gttttttcct gtttggtcac tgatgcctcc gtgtaagggg gatttctgtt catgggggta 3301 atgataccga tgaaacgaga gaggatgctc acgatacggg ttactgatga tgaacatgcc 3361 cggttactgg aacgttgtga gggtaaacaa ctggcggtat ggatgcggcg ggaccagaga 3421 aaaatcactc agggtcaatg ccagcgcttc gttaatacag atgtaggtgt tccacagggt 3481 agccagcagc atcctgcgat gcagatccgg aacataatgg tgcagggcgc tgacttccgc 3541 gtttccagac tttacgaaac acggaaaccg aagaccattc atgttgttgc tcaggtcgca 3601 gacgttttgc agcagcagtc gcttcacgtt cgctcgcgta tcggtgattc attctgctaa 3661 ccagtaaggc aaccccgcca gcctagccgg gtcctcaacg acaggagcac gatcatgcgc 3721 acccgtgggg ccgccatgcc ggcgataatg gcctgcttct cgccgaaacg tttggtggcg 3781 ggaccagtga cgaaggcttg agcgagggcg tgcaagattc cgaataccgc aagcgacagg 3841 ccgatcatcg tcgcgctcca gcgaaagcgg tcctcgccga aaatgaccca gagcgctgcc 3901 ggcacctgtc ctacgagttg catgataaag aagacagtca taagtgcggc gacgatagtc 3961 atgccccgcg cccaccggaa ggagctgact gggttgaagg ctctcaaggg catcggtcga 4021 gatcccggtg cctaatgagt gagctaactt acattaattg cgttgcgctc actgcccgct 4081 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga 4141 ggcggtttgc gtattgggcg ccagggtggt ttttcttttc accagtgaga cgggcaacag 4201 ctgattgccc ttcaccgcct ggccctgaga gagttgcagc aagcggtcca cgctggtttg 4261 ccccagcagg cgaaaatcct gtttgatggt ggttaacggc gggatataac atgagctgtc 4321 ttcggtatcg tcgtatccca ctaccgagat atccgcacca acgcgcagcc cggactcggt 4381 aatggcgcgc attgcgccca gcgccatctg atcgttggca accagcatcg cagtgggaac 4441 gatgccctca ttcagcattt gcatggtttg ttgaaaaccg gacatggcac tccagtcgcc 4501 ttcccgttcc gctatcggct gaatttgatt gcgagtgaga tatttatgcc agccagccag 4561 acgcagacgc gccgagacag aacttaatgg gcccgctaac agcgcgattt gctggtgacc 4621 caatgcgacc agatgctcca cgcccagtcg cgtaccgtct tcatgggaga aaataatact 4681 gttgatgggt gtctggtcag agacatcaag aaataacgcc ggaacattag tgcaggcagc 4741 ttccacagca atggcatcct ggtcatccag cggatagtta atgatcagcc cactgacgcg 4801 ttgcgcgaga agattgtgca ccgccgcttt acaggcttcg acgccgcttc gttctaccat 4861 cgacaccacc acgctggcac ccagttgatc ggcgcgagat ttaatcgccg cgacaatttg 4921 cgacggcgcg tgcagggcca gactggaggt ggcaacgcca atcagcaacg actgtttgcc 4981 cgccagttgt tgtgccacgc ggttgggaat gtaattcagc tccgccatcg ccgcttccac 5041 tttttcccgc gttttcgcag aaacgtggct ggcctggttc accacgcggg aaacggtctg 5101 ataagagaca ccggcatact ctgcgacatc gtataacgtt actggtttca cattcaccac 5161 cctgaattga ctctcttccg ggcgctatca tgccataccg cgaaaggttt tgcgccattc 5221 gatggtgtcc gggatctcga cgctctccct tatgcgactc ctgcattagg aagcagccca 5281 gtagtaggtt gaggccgttg agcaccgccg ccgcaaggaa tggtgcatgc aaggagatgg 5341 cgcccaacag tcccccggcc acggggcctg ccaccatacc cacgccgaaa caagcgctca 5401 tgagcccgaa gtggcgagcc cgatcttccc catcggtgat gtcggcgata taggcgccag 5461 caaccgcacc tgtggcgccg gtgatgccgg ccacgatgcg tccggcgtag aggatcgaga 5521 tctcgatccc gcgaaat //