LOCUS Exported 5479 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Tet-inducible RRNA promoter driven plasmid for the generation of stem loop RNAi transcripts from a specific RRNA spacer locus in T. brucei.. ACCESSION . VERSION . KEYWORDS pRPa-iSL(MCS1+2) SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5479) AUTHORS Alsford S, Horn D TITLE Single-locus targeting constructs for reliable regulated RNAi and transgene expression in Trypanosoma brucei. JOURNAL Mol Biochem Parasitol. 2008 Sep;161(1):76-9. doi: 10.1016/j.molbiopara.2008.05.006. Epub 2008 Jun 7. PUBMED 18588918 REFERENCE 2 (bases 1 to 5479) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5479 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind complement(665..682) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(836..1424) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(916..935) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(1595..2455) /codon_start=1 /gene="bla" /product="beta-lactamase" /label=AmpR /note="confers resistance to ampicillin, carbenicillin, and related antibiotics" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" primer_bind 2218..2237 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(2456..2560) /gene="bla" /label=AmpR promoter primer_bind 2628..2646 /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" primer_bind complement(2684..2706) /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind 2806..2825 /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 3035..3054 /label=pBRrevBam /note="pBR322 vectors, tet region, downstream of BamHI, reverse primer" primer_bind complement(5009..5030) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 5221..5240 /label=F1ori-R /note="F1 origin, reverse primer" ORIGIN 1 gaattcgatg cgaggcgaat cgctcagtag tgttagtgca ggcattgttt ctcttttcca 61 tttcggctcg ctctctgtgc cagaaaggct tttggagtct tggttactcg tgggactagg 121 gcaagcgtcg cttccctaat aattgttgct aacgtttcac cgtgcgtgtg tggatatctt 181 ccaagttgtc attttctcta cccctctctt ttccggcgcg gtctaacttc ctccaactag 241 cgcatttcca tgataacggg gtccacatag tccgttgtag tgaaattggc atgccatcag 301 ggccaaggca tacatcgcgg ccgcgataaa aatgatacga tgatctccgt gtggagcaga 361 ccgtggcgac cagtttgtac catggaagtt cttcgataga gttatatcga gtgcggaatt 421 gatgctacgg tcacagtgtc gtatcgtgtg gtctcccatg tgtttttgac ggtgtaccaa 481 ttatctttgc actaatgggc tatggaatct ttccttttaa tcagggggac atgtttatcc 541 cacgccagta gtctggaatt agaatacata ccaactatat ggcgcgccga gctcgaattc 601 tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg 661 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc 721 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt 781 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc 841 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa 901 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc 961 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg 1021 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc 1081 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc 1141 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca 1201 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact 1261 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg 1321 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt 1381 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct 1441 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga 1501 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa 1561 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac 1621 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga 1681 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc 1741 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca 1801 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta 1861 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg 1921 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc 1981 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg 2041 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt 2101 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt 2161 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata 2221 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc 2281 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac 2341 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa 2401 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct 2461 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat 2521 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc 2581 cacctgacgt ctaagaaacc attattatca tgacattaac ctataaaaat aggcgtatca 2641 cgaggccctt tcgtctcgcg cgtttcggtg atgacggtga aaacctctga cacatgcagc 2701 tcccggagac ggtcacagct tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg 2761 gcgcgtcagc gggtgttggc gggtgtcggg gctggcttaa ctatgcggca tcagagcaga 2821 ttgtactgag agtgcaccat tcgacgctct cccttatgcg actcctgcat taggaagcag 2881 cccagtagta ggttgaggcc gttgagcacc gccgccgcaa ggaatggtgc aaggagatgg 2941 cgcccaacag tcccccggcc acggggcctg ccaccatacc cacgccgaaa caagcgctca 3001 tgagcccgaa gtggcgagcc cgatcttccc catcggtgat gtcggcgata taggcgccag 3061 caaccgcacc tgtggcgccg gtgatgccgg ccacgatgcg tccggcgtag aggatctggc 3121 taggaatact gcatagataa caaacgcatc aaacacaact ggtggcacta tttcaatcat 3181 gtcgacggcg cgccctcgga cgagtgctgg ggcgtcggtt tccactatcg gcgagtactt 3241 ctacacagcc atcggtccag acggccgcgc ttctgcgggc gatttgtgta cgcccgacag 3301 tcccggctcc ggatcggacg attgcgtcgc atcgaccctg cgcccaagct gcatcatcga 3361 aattgccgtc aaccaagctc tgatagagtt ggtcaagacc aatgcggagc atatacgccc 3421 ggagccgcgg cgatcctgca agctccggat gcctccgctc gaagtagcgc gtctgctgct 3481 ccatacaagc caaccacggc ctccagaaga agatgttggc gacctcgtat tgggaatccc 3541 cgaacatcgc ctcgctccag tcaatgaccg ctgttatgcg gccattgtcc gtcaggacat 3601 tgttggagcc gaaatccgcg tgcacgaggt gccggacttc ggggcagtcc tcggcccaaa 3661 gcatcagctc atcgagagcc tgcgcgacgg acgcactgac ggtgtcgtcc atcacagttt 3721 gccagtgata cacatgggga tcagcaatcg cgcatatgaa atcacgccat gtagtgtatt 3781 gaccgattcc ttgcggtccg aatgggccga acccgctcgt ctggctaaga tcggccgcag 3841 cgatcgcatc catggcctcc gcgaccggct gcagaacagc gggcagttcg gtttcaggca 3901 ggtcttgcgt gacaccctgt gcacggcggg agatgcaata ggtcaggctc tcgctgaatt 3961 ccccaatgtc aagcacttcc ggaatcggga gcgcggccga tgcaaagtgc cgataaacat 4021 aacgatcttt gtagaaacca tcggcgcagc tatttacccg caggacatat ccacgccctc 4081 ctacatcgaa gctgaaagca cgagattctt cgccctccga gagctgcatc aggtcggaga 4141 cgctgtcgaa cttttcgatc agaaacttct cgacagacgt cgcggtgagt tcaggctttt 4201 tcatcactag acttcgttgc agttgagttt ctgttttcgg gcttgttggt atacttgttt 4261 gattaatgtg gctgatagct aagctatcgc aaatatttgt acaagacatg agggggcaaa 4321 aaaataagcc aagtgtattt gttaggcttc cgtgcctgtt agagaaatat gtaaggaaac 4381 gcgtgcgcta cacagcttga gcacccatcg aattgggacg agaacatgtt aaactcagta 4441 ctcacccgat aacccctatt ttcaatgcac gcccacccag agacacggaa tagcataaaa 4501 caaccccatc actgccacaa aatggacatt ttgcacaaaa cgcactattg actagtattc 4561 gactatacta ataagagata cctatcagac acgatatact gtgaaaagga cattaagctc 4621 actatgcatc tgaaaacgct ttcacctcac aacctactta cacacacctt tccgcttaac 4681 ccgctagctt tccacccagc gcgggtgcat tctggctctt atatatactt attgtcatga 4741 cagagtatat tgtactgtgt tgataaggga cgggtaactg tattgaagag ccgatgcttt 4801 tgacatgtta gatataatat gttttattgt aaagtcaata caacacacaa taggataata 4861 atgataaagt taaaaaagta tatatagtaa tagaaatata tcttatatag gaaagattaa 4921 gcagtaaaag tagcgcttac ggcgtacacc ctatcaatga tagagtggta cccgggctta 4981 aggatccgag atagggttga gtgttgttcc agtttggaac aagagtccac tattaaagaa 5041 cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga 5101 accatcaccc taatcaagtt ttttggggtc gaggtgccgt aaagcactaa atcggaaccc 5161 taaagggagc ccccgattta gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga 5221 agggaagaaa gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg 5281 cgtaaccacc acacccgccg cgcttaatgc gccgctacag ggcgcgtcca ttcgccattc 5341 aggctgcgca actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccagctg 5401 gcgaaagggg gatgtgctgc aaggcgatta agttgggtaa cgccagggtt ttctagaagc 5461 ttctcgaggg cccctgcag //