LOCUS Exported 6428 bp ds-DNA circular SYN 12-SEP-2022 DEFINITION Inducible expression vector for Trypanosoma brucei. ACCESSION . VERSION . KEYWORDS pBA310 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6428) AUTHORS Ishii M, Akiyoshi B TITLE Targeted protein degradation using deGradFP in Trypanosoma brucei. JOURNAL Wellcome Open Res. 2022 Jun 23;7:175. doi: 10.12688/wellcomeopenres.17964.1. eCollection 2022. PUBMED 35865221 REFERENCE 2 (bases 1 to 6428) AUTHORS . TITLE Direct Submission JOURNAL Exported Sep 12, 2022 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6428 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind complement(66..83) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(237..825) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(317..336) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(996..1856) /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 1619..1638 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(1857..1961) /gene="bla" /label=AmpR promoter primer_bind 2029..2047 /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" primer_bind complement(2085..2107) /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind 2207..2226 /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 2436..2455 /label=pBRrevBam /note="pBR322 vectors, tet region, downstream of BamHI, reverse primer" CDS complement(2865..3239) /codon_start=1 /gene="Sh ble from Streptoalloteichus hindustanus" /product="antibiotic-binding protein" /label=BleoR /note="confers resistance to bleomycin, phleomycin, and Zeocin(TM)" /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR EFALRDPAGNCVHFVAEEQD" terminator 3705..3752 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" terminator 3837..3884 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" primer_bind 4136..4155 /label=T7 /note="T7 promoter, forward primer" promoter 4136..4154 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" protein_bind 4161..4179 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" protein_bind 4292..4310 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" protein_bind 4313..4331 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" CDS 4467..5180 /codon_start=1 /product="enhanced GFP" /label=EGFP /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE FVTAAGITLGMDELYK" CDS 4467..5180 /codon_start=1 /product="the original enhanced GFP (Yang et al., 1996)" /label=EGFP /note="mammalian codon-optimized" /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE FVTAAGITLGMDELYK" primer_bind complement(4509..4530) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(4770..4789) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 5117..5138 /label=EGFP-C /note="EGFP, forward primer" promoter complement(5655..5673) /label=SP6 promoter /note="promoter for bacteriophage SP6 RNA polymerase" primer_bind complement(5656..5673) /label=SP6 /note="SP6 promoter, forward primer" ORIGIN 1 ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc 61 gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct 121 cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg 181 tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc 241 cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga 301 aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct 361 cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg 421 gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag 481 ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat 541 cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac 601 aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac 661 tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc 721 ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt 781 tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc 841 ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg 901 agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca 961 atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca 1021 cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag 1081 ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac 1141 ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc 1201 agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct 1261 agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc 1321 gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg 1381 cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc 1441 gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat 1501 tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag 1561 tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat 1621 aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg 1681 cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca 1741 cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga 1801 aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc 1861 ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag cggatacata 1921 tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg 1981 ccacctgacg tctaagaaac cattattatc atgacattaa cctataaaaa taggcgtatc 2041 acgaggccct ttcgtctcgc gcgtttcggt gatgacggtg aaaacctctg acacatgcag 2101 ctcccggaga cggtcacagc ttgtctgtaa gcggatgccg ggagcagaca agcccgtcag 2161 ggcgcgtcag cgggtgttgg cgggtgtcgg ggctggctta actatgcggc atcagagcag 2221 attgtactga gagtgcacca ttcgacgctc tcccttatgc gactcctgca ttaggaagca 2281 gcccagtagt aggttgaggc cgttgagcac cgccgccgca aggaatggtg caaggagatg 2341 gcgcccaaca gtcccccggc cacggggcct gccaccatac ccacgccgaa acaagcgctc 2401 atgagcccga agtggcgagc ccgatcttcc ccatcggtga tgtcggcgat ataggcgcca 2461 gcaaccgcac ctgtggcgcc ggtgatgccg gccacgatgc gtccggcgta gaggatctgg 2521 ctaggaatac tgcatagata acaaacgcat caaacacaac tggtggcact atttcaatca 2581 tgtcgacaca ccaagacgag aaaatttgag ttaaacaaga tgcctacaac cctatattta 2641 ttatcttcaa gtgatcttct cttctcttgt gcgctgtacg taaatgtgtt gcaaataagg 2701 catagggctg agtacaggca ccacacgcga ggaaaatcat aagcactagc cgtgtcaact 2761 cacagcgact acagaacaat tttggccaca caacccggtg ttaggatctc cgaggcctcg 2821 gacccgtcgg gccgcgtcgg accggcggtg ttggtcggcg tcggtcagtc ctgctcctcg 2881 gccacgaagt gcacgcagtt gccggccggg tcgcgcaggg cgaactcccg cccccacggc 2941 tgctcgccga tctcggtcat ggccggcccg gaggcgtccc ggaagttcgt ggacacgacc 3001 tccgaccact cggcgtacag ctcgtccagg ccgcgcaccc acacccaggc cagggtgttg 3061 tccggcacca cctggtcctg gaccgcgctg atgaacaggg tcacgtcgtc ccggaccaca 3121 ccggcgaagt cgtcctccac gaagtcccgg gagaacccga gccggtcggt ccagaactcg 3181 accgctccgg cgacgtcgcg cgcggtgagc accggaacgg cactggtcaa cttggccatg 3241 gtgatagctt attttatggc agcaacgaga ccttacgtaa agcaaaacta aataagagcg 3301 gagactgcaa tgcagagtaa aaaaaaaaaa aacatgaatt tcagaagacc ttgctgtgcc 3361 cgggtctgag agcggtcagt tgctctcctg ctccgtacgc cgtaagcgct acttttactg 3421 cttaatcttt cctatataag atatatttct attactatat atactttttt aactttatca 3481 ttattatcct attgtgtgtt gtattgactt tacaataaaa catattatat ctaacatgtc 3541 aaaagcatcg gctcttcaat acagttaccc gtcccttatc aacacagtac aatatactct 3601 gtcatgacaa taagtatata taagagccag aatgcacccg cgctgggtgg aaagctagct 3661 tgcatgcctg caggggccca atccggatat agttctcctt tcagcaaaaa acccctcaag 3721 acccgtttag aggccccaag gggttatgct agttattgct cagcggtggc agcagccaac 3781 tcagcttcct ttcgggcttt gttagcagcc ctatccggat atagttctcc tttcagcaaa 3841 aaacccctca agacccgttt agaggcccca aggggttatg ctagttattg ctcagcggtg 3901 gcagcagcca actcagcttc ctttcgggct ttgttagcag cctgcagaat actgcataga 3961 taacaaacgc atcaaacaca actggtggca ctatttcaat catgtcgagg aggaccgaat 4021 actaataacg acacttgcgg tcaaaaagta gaagaacaaa tgctcaacga tgagtgaatc 4081 aggttagggt agttggaaaa ttatacagaa tgtctttggc aacacaccgg ctgattaata 4141 cgactcacta tagggagatc tccctatcag tgatagagat ctccccgggc cccccctcga 4201 gtccattttg tggcagtgat ggggttgttt tatgctattc cgtgtctctg ggtgggcgtg 4261 cattgaaaat aggggttatc gggtagggat ctccctatca gtgatagaga tctccctatc 4321 agtgatagag atccctgagt actgagttta acatgttctc gtcccgggct gcacgcgcct 4381 tcgagttttt tttccttttc cccatttttt tcaacttgaa gacttcaatt acaccaaaaa 4441 gtaaaattca caagcttatg actagtgtga gcaagggcga ggagctgttc accggggtgg 4501 tgcccatcct ggtcgagctg gacggcgacg taaacggcca caagttcagc gtgtccggcg 4561 agggcgaggg cgatgccacc tacggcaagc tgaccctgaa gttcatctgc accaccggca 4621 agctgcccgt gccctggccc accctcgtga ccaccctgac ctacggcgtg cagtgcttca 4681 gccgctaccc cgaccacatg aagcagcacg acttcttcaa gtccgccatg cccgaaggct 4741 acgtccagga gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg 4801 tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg 4861 aggacggcaa catcctgggg cacaagctgg agtacaacta caacagccac aacgtctata 4921 tcatggccga caagcagaag aacggcatca aggtgaactt caagatccgc cacaacatcg 4981 aggacggcag cgtgcagctc gccgaccact accagcagaa cacccccatc ggcgacggcc 5041 ccgtgctgct gcccgacaac cactacctga gcacccagtc cgccctgagc aaagacccca 5101 acgagaagcg cgatcacatg gtcctgctgg agttcgtgac cgccgccggg atcactctcg 5161 gcatggacga gctgtacaag cggtaccgtt ctagaatcga taccggtcga ggacacaagc 5221 ggtcacgtga acaattgccg cgggatatcc atatgttaat taaggcgcgc ctgatcagtt 5281 taaacggatc ccgctgcgct taagtgtctt tcattataat caatgtataa cctttatgta 5341 gtatttcaat ctatgccgct gtgtacgtgc actgcggtgc ctatccttcg gcattagaga 5401 gtcactgttt gtgtagatcg tagctgcatg tctgacatct ggttcattag tgccgttttg 5461 agtctgcact gttgtgcaac ttgatatgca tcactgcaca ccaaccgtgt gtttcctcct 5521 ttgggtgcac cctttaatcc ttgtcttctc cttttttgtc tctttccccc tcgaaaaggt 5581 gtcaaactac tgccgcataa actacggtta tcccaaattt aagagaaagc aataaagcat 5641 cacctagctt gtattctata gtgtcaccta aatcgtatgt gtatgataca taaggttatg 5701 tattaattgt agccgcgttc taacgacaat atgtacaagc ctaattgtgt agcatctggc 5761 ttactgaagc agaccctatc atctctctcg taaactgccg tcagagtcgg tttggttgga 5821 cgaaccttct gagtttctgg taacgccgtc ccgcacccgg aaatggtcag cgaaccaatg 5881 gtacccatta aacactaaag aacattgttg caaacttgca aattaatgtt acacacttgt 5941 atttaatgtt gcacacttgt aattaatatg ggacacatta aaagttattg tgtataatag 6001 cgttaactgc ggattgttta atattcgttc gtataaggac cattaaatag ctttgttgca 6061 cccattaaac actaaagaac attgttgcaa acttgcaaat taatgttaca cacttgtatt 6121 taatgttgca cacggatcgc ggccgcgatc cgtaattaat atggcacaca ttaaaagtta 6181 ttgtgtataa tagcgttaac tgcgcattgt ttaatattcg ttcgtataag gaccattaaa 6241 tagctttgtt gcacacatta aacactaaag aacattgttt caaacttgca aattaacgtt 6301 acacacttgt atttaatgtt gcacacttgt aattaatatg gcagacatta aaagttattg 6361 tgtataatag cgttaactgc gcattgttta atattcgttc gtataaggac cattaaagag 6421 ctcgaatt //