LOCUS Exported 5628 bp DNA circular SYN 02-JUL-2021 DEFINITION BBF10K_000540 in pOpen_v4. ACCESSION . VERSION . KEYWORDS . SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5628) AUTHORS . TITLE Direct Submission JOURNAL Exported Jul 2, 2021 from SnapGene 5.3.1 https://www.snapgene.com COMMENT This file was created using tools provided by Twist Bioscience FEATURES Location/Qualifiers source 1..5628 /organism="BBF10K_000540 in pOpen_v4" primer_bind 21..51 /label=Gibson-for regulatory 46..55 /regulatory_class="other" /label=Kozak sequence /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 52..1134 /codon_start=1 /product="G-protein coupled receptor 183" /label=GPR183 /translation="MDIQMANNFTPPSATPQGNDCDLYAHHSTARIVMPLHYSLVFIIG LVGNLLALVVIVQNRKKINSTTLYSTNLVISDILFTTALPTRIAYYAMGFDWRIGDALC RITALVFYINTYAGVNFMTCLSIDRFIAVVHPLRYNKIKRIEHAKGVCIFVWILVFAQT LPLLINPMSKQEAERITCMEYPNFEETKSLPWILLGACFIGYVLPLIIILICYSQICCK LFRTAKQNPLTEKSGVNKKALNTIILIIVVFVLCFTPYHVAIIQHMIKKLRFSNFLECS QRHSFQISLHFTVCLMNFNCCMDPFIYFFACKGYKRKVMRMLKRQVSVSISSAVKSAPE ENSREMTETQMMIHSKSSNGK" primer_bind complement(1135..1159) /label=Gibson-rev terminator 1278..1325 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" primer_bind complement(1330..1346) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" misc_feature 1505..1755 /gene="incC" /label=incC /note="incompatibility region of the bacterial F plasmid" CDS complement(1758..2513) /codon_start=1 /gene="repE" /product="replication initiation protein for the bacterial F plasmid" /label=repE /translation="MAETAVINHKKRKNSPRIVQSNDLTEAAYSLSRDQKRMLYLFVDQ IRKSDGTLQEHDGICEIHVAKYAEIFGLTSAEASKDIRQALKSFAGKEVVFYRPEEDAG DEKGYESFPWFIKRAHSPSRGLYSVHINPYLIPFFIGLQNRFTQFRLSETKEITNPYAM RLYESLCQYRKPDGSGIVSLKIDWIIERYQLPQSYQRMPDFRRRFLQVCVNEINSRTPM RLSYIEKKKGRQTTHIVFSFRDITSMTTG" rep_origin 2604..2823 /label=ori2 /note="secondary origin of replication for the bacterial F plasmid; also known as oriS" protein_bind 3243..3264 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." promoter 3279..3309 /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 3317..3333 /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)." primer_bind 3341..3357 /label=M13 rev /note="common sequencing primer, one of multiple similar variants" CDS 3430..4275 /codon_start=1 /label=repL /translation="MLAKVTFLSCITMSDFTFSGYELACFVTHSGLSRSAGHILSQCAN LAATTSEYFIHKPHRLIAAETGYSQSTVVRAFREAVNKGILSVEIVIGDHRERRANLYR FTPSFLAFAQQAKNALIESKLKISSAATKVKAVLAKTLALFNFLSTPPCQNDTPSPCQD DVAIKNKKSQVKKTKRSVSGGAGTTSLKKLTSWIAKAKAKADNLRLSKKRTQKHEFKQK VEAAARKYAYLKNKRSPDIGGISNFDNLPHCMTVNEALNAVLAKNKDNEQWGIPAGFRG " misc_feature 4383..4534 /label=nnpp promoters CDS 4574..5389 /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" terminator complement(5499..5542) /label=bacterial terminator /note="putative bacterial transcription terminator" primer_bind 5560..5576 /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" terminator 5577..5608 /label=tonB terminator /note="bidirectional E. coli tonB-P14 transcription terminator" ORIGIN 1 aagaattact gacccctcgg cagactgagt cgcccggggg gatccgccac catggacatc 61 cagatggcca acaacttcac cccccccagc gccacccccc agggcaacga ctgcgacctg 121 tacgcccacc acagcaccgc cagaatcgtg atgcccctgc actacagcct ggtgttcatc 181 atcggcctgg tgggcaacct gctggccctg gtggtgatcg tgcagaacag aaagaagatc 241 aacagcacca ccctgtacag caccaacctg gtgatcagcg acatcctgtt caccaccgcc 301 ctgcccacca gaatcgccta ctacgccatg ggcttcgact ggagaatcgg cgacgccctg 361 tgcagaatca ccgccctggt gttctacatc aacacctacg ccggcgtgaa cttcatgacc 421 tgcctgagca tcgacagatt catcgccgtg gtgcaccccc tgagatacaa caagatcaag 481 agaatcgagc acgccaaggg cgtgtgcatc ttcgtgtgga tactggtgtt cgcccagacc 541 ctgcccctgc tgatcaaccc catgagcaag caggaggccg agagaatcac ctgcatggag 601 taccccaact tcgaggagac caagagcctg ccctggatac tgctgggcgc ctgcttcatc 661 ggctacgtgc tgcccctgat catcatcctg atctgctaca gccagatctg ctgcaagctg 721 ttcagaaccg ccaagcagaa ccccctgacc gagaagagcg gcgtgaacaa gaaggccctg 781 aacaccatca tcctgatcat cgtggtgttc gtgctgtgct tcacccccta ccacgtggcc 841 atcatccagc acatgatcaa gaagctgaga ttcagcaact tcctggagtg cagccagaga 901 cacagcttcc agatcagcct gcacttcacc gtgtgcctga tgaacttcaa ctgctgcatg 961 gaccccttca tctacttctt cgcctgcaag ggctacaaga gaaaggtgat gagaatgctg 1021 aagagacagg tgagcgtgag catcagcagc gccgtgaaga gcgcccccga ggagaacagc 1081 agagagatga ccgagaccca gatgatgatc cacagcaaga gcagcaacgg caagggatcc 1141 gccaccaact tcagcctgct agttgcagta tctaacccgc tgaggtcagt gacaaggtca 1201 ttagggataa cagggtaatc ctaacggagt attagccgaa cgctgaggtg tcaatcgtcg 1261 gagccgctga gcaataacta gcataacccc ttggggcctc taaacgggtc ttgaggggtt 1321 ttttgcatgg tcatagctgt ttcctgagag cttggcaggt gtcgcgtatg catcgatcac 1381 gtgtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact cggatcgatc 1441 cggcgcgcac cgtgggaaaa actccaggta gaggtacaca cgcggatagc caattcagag 1501 taataaactg tgataatcaa ccctcatcaa tgatgacgaa ctaacccccg atatcaggtc 1561 acatgacgaa gggaaagaga aggaaatcaa ctgtgacaaa ctgccctcaa atttggcttc 1621 cttaaaaatt acagttcaaa aagtatgaga aaatccatgc aggctgaagg aaacagcaaa 1681 actgtgacaa attaccctca gtaggtcaga acaaatgtga cgaaccaccc tcaaatctgt 1741 gacagataac cctcagacta tcctgtcgtc atggaagtga tatcgcggaa ggaaaatacg 1801 atatgagtcg tctggcggcc tttctttttc tcaatgtatg agaggcgcat tggagttctg 1861 ctgttgatct cattaacaca gacttgcagg aagcggcggc ggaagtcagg catacgctgg 1921 taactttgag gcagctggta acgctctatg atccagtcga ttttcagaga cacgatgcct 1981 gagccatccg gcttacgata ctgacacagg gattcgtata aacgcatggc atacggattg 2041 gtgatttctt ttgtttcact aagccgaaac tgcgtaaacc ggttctgtaa cccgataaag 2101 aagggaatga gatatgggtt gatatgtaca ctgtaaagcc ctctggatgg actgtgcgca 2161 cgtttgataa accaaggaaa agattcatag cctttttcat caccggcatc ctcttcaggg 2221 cgataaaaaa ccacttcctt ccccgcgaaa ctcttcaatg cctgccgtat atccttactg 2281 gcttccgcag aggtcaatcc gaatatttca gcatatttag caacatggat ctcgcagata 2341 ccgtcatgtt cctgtagggt gccatcagat tttctgatct ggtcaacgaa cagatacagc 2401 atacgttttt gatcccggga gagactatat gccgcctcag tgaggtcgtt tgactggacg 2461 attcgcgggc tatttttacg tttcttgtga ttgataaccg ctgtttccgc catgacagat 2521 ccatgtgaag tgtgacaagt ttttagattg tcacactaaa taaaaaagag tcaataagca 2581 gggataactt tgtgaaaaaa cagcttcttc tgagggcaat ttgtcacagg gttaagggca 2641 atttgtcaca gacaggactg tcatttgagg gtgatttgtc acactgaaag ggcaatttgt 2701 cacaacacct tctgtagaac cagcatggat aaaggcctac aaggcgctct aaaaaagaag 2761 atctaaaaac tataaaaaaa ataattataa aaatatcccc gtggataagt ggataacccc 2821 aagggaagtt ttttcaggca tcgtgtgtaa gcagaatata taagtgctgt tccctggtgc 2881 ttcctcgctc actcgaaatt cccggggata gctttatgct tgtaaaccgt tttgtgaaaa 2941 aatttttaaa ataaaaaagg ggacctctag ggtccccaat taattagtaa tataatctat 3001 taaaggtcat tcaaaaggtc atccaccgga tcaattcccc tgctcgcgca ggctgggtgc 3061 caagctctcg ggtaacatca aggcccgatc cttggagccc ttgccctccc gcacgatgat 3121 cgtgccgtga tcgaaatcca gatccttgac ccgcagttgc aaaccctcac tgatccgatt 3181 cattaatgca gctggcacga caggtttccc gactggaaag cgggcagtga gcgcaacgca 3241 attaatgtga gttagctcac tcattaggca ccccaggctt tacactttat gcttccggct 3301 cgtatgttgt gtggaattgt gagcggataa caatttcaca caggaaacag ctatgaccat 3361 gattacgcca agcgcggtac ctgtcgcggc aacgcgctaa cagacgtagt aagaaccacc 3421 agcattgtaa tgctggctaa agtcactttc ctgagctgta taacgatgag cgattttact 3481 ttttctggct atgaattggc ctgctttgta acacactccg gtctatcccg tagcgccggg 3541 catatcctgt cgcaatgtgc aaatctcgcg gcaacaacca gtgaatactt cattcacaag 3601 cctcaccgcc tgatcgcggc agaaactggt tatagccaat caaccgtcgt tcgtgcattc 3661 cgtgaagctg taaacaaagg tatcctgtct gtagagattg ttatcggcga tcaccgtgaa 3721 cgtcgcgcta acctgtaccg gtttacacca tcctttttgg ccttcgcaca acaagccaaa 3781 aatgcgctga tagaaagcaa attaaagatc tcttcagcgg caaccaaggt taaagctgtt 3841 ctcgctaaga cattggcttt atttaatttt ttatccacac ccccatgtca aaatgatacc 3901 ccctccccct gtcaggatga cgtggcaata aagaataaga agtcacaagt taaaaaaaca 3961 aaaagatcag tttccggcgg tgccggaaca accagcctca aaaaattgac ttcatggatc 4021 gctaaggcaa aagcaaaggc tgacaatctg cggttatcca aaaaacgcac tcaaaaacat 4081 gagttcaagc agaaagtaga ggcggctgcg cggaaatatg cttacctgaa gaacaagcgt 4141 tcgcctgata ttggcgggat atcaaacttc gataacctac cgcattgcat gacggtaaac 4201 gaagctctta atgcggtttt agccaaaaat aaagataacg aacaatgggg tataccggca 4261 ggattcagag ggtaatgaat tgctctaatt ataaccatgc atactttcaa cacctctagt 4321 ttgccatgag gcaaactcat aggtgtcctg gtaagaggac actgttgcca aaactggacg 4381 ccccattatt gcaattaata aacaactaac ggacaattct acctaacaat aagtggagtt 4441 gcggccggcc gcgatccgct agcaaagcca cgttgtgtct caaaatctct gatgttacat 4501 tgcacaagat aaaaatatat catcatgaac aataaaactg tctgcttaca taaacagtaa 4561 tacaaggggt gttatgagcc atattcaacg ggaaacgtct tgctcgcgcc cgcgattaaa 4621 ttccaacatg gatgctgatt tatatgggta taaatgggct cgcgataatg tcgggcaatc 4681 aggtgcgaca atctatcgat tgtatgggaa gcccgatgcg ccagagttgt ttctgaaaca 4741 tggcaaaggt agcgttgcca atgatgttac agatgagatg gtcagactaa actggctgac 4801 ggaatttatg cctcttccga ccatcaagca ttttatccgt actcctgatg atgcatggtt 4861 actcaccact gcgatccccg ggaaaacagc attccaggta ttagaagaat atcctgattc 4921 aggtgaaaat attgttgatg cgctggcagt gttcctgcgc cggttgcatt cgattcctgt 4981 ttgtaattgt ccttttaaca gcgatcgcgt atttcgcctc gctcaggcgc aatcacgaat 5041 gaataacggt ttggttgatg cgagtgattt tgatgacgag cgtaatggct ggcctgttga 5101 acaagtctgg aaagaaatgc ataaactgtt gccattctca ccggattcag tcgtcactca 5161 tggtgatttc tcacttgata accttatttt tgacgagggg aaattaatag gttgtattga 5221 tgttggacga gtcggaatcg cagaccgata ccaggatctt gccatcctat ggaactgcct 5281 cggtgagttt tctccttcat tacagaaacg gctttttcaa aaatatggta ttgataatcc 5341 tgatatgaat aaattgcagt ttcatttgat gctcgatgag tttttctaat cagaattggt 5401 taattggttg taacactggc gagctctgcc acctgacgtc taagaaacca ttattatcat 5461 gacattaacc tataaaaata ggcgtatcac gaggcagaat ttcagataaa aaaaatcctt 5521 agctttcgct aaggatgatt tctgcacctg caccagtcag taaaacgacg gccagtagtc 5581 aaaagcctcc gaccggaggc ttttgacttg gttcaggtgg agtgggag //