LOCUS       sequence-355674-        5335 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Trypanosoma brucei PCR only tagging (pPOT) vector for endogenous 
            gene tagging.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5335)
  AUTHORS   Paterou A, Saez Conde J, Tyc J, Sunter JD, Vaughan S, Gull K, Dean S
  TITLE     A comprehensive toolkit for protein localization and functional 
            analysis in trypanosomatids.
  JOURNAL   Open Biol. 2025 Apr;15(4):240361. doi: 10.1098/rsob.240361. Epub 
            2025 Apr 2.
  PUBMED    40169015
REFERENCE   2  (bases 1 to 5335)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Open 
            Biol."; date: "2025-04"; volume: "15(4):240361. doi"; pages: " 
            10.1098/rsob.240361. Epub 2025 Apr 2"
COMMENT     Sequence Label: pPOTv7-blast-nanoluc
FEATURES             Location/Qualifiers
     source          1..5335
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(2460..2478)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2461..2478)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2496..2512)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2496..2512)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2509..2531)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2520..2536
                     /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)."
     promoter        complement(join(2544..2550,2551..2568,2569..2574))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2589..2610
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2727..2744)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2898..3486)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2978..2997)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3657..4448,4449..4517))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVIIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     4280..4299
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4518..4622)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(4700..5155)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4837..4858)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5049..5068
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5281..5303
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     5295..5312
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     5296..5312
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     join(5319..5335,1..3)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        join(5319..5335,1..2)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 gggcgaattg ggcccgtata atgcagacct gctgcaagct aggggcacag caaggtcttc
       61 tgaaattcat gttttttttt tttttactct gcattgcagt ctccgctctt atttagtttt
      121 gctttacgta aggtctcgtt gctgccataa aataagctct agtggaattc atgcctttgt
      181 ctcaagaaga atccaccctc attgaaagag caacggctac aatcaacagc atccccatct
      241 ctgaagacta cagcgtcgcc agcgcagctc tctctagcga cggccgcatc ttcactggtg
      301 tcaatgtata tcattttact gggggacctt gtgcagaact cgtggtgctg ggcactgctg
      361 ctgctgcggc agctggcaac ctgacttgta tcgtcgcgat cggaaatgag aacaggggca
      421 tcttgagccc ctgcggacgg tgccgacagg tgcttctcga cctgcatcct gggatcaaag
      481 ccatagtgaa ggacagtgat ggacagccga cggcagttgg gattcgtgaa ttgctgccct
      541 ctggttatgt gtgggagggc taaccatggt gcccatttag ttggcctttc ccttgtctcg
      601 tgtcttttcc gtggaaaggt tcccggagta atctgatggc acagcaggga ggtgcgcctg
      661 caggttggtt aggaaggggg gatgatgtaa aagaagaaaa tggggggatt cgagccccac
      721 gcgttgctca agctgtgtag cgcaggcgtt tccttacata tttctctaac aggcacggaa
      781 gcctaacaaa tacacttggc ttattttttt gccccctcat gtcttgtaca aatatttgcg
      841 atagcttagc tatcagccac attaatcaaa caagtatacc aacaagcccg aaaacagaaa
      901 ctcaactgca acgggtacct atgggttctg gtagtggttc cggttccggt tctaagctta
      961 tggtatttac tttggaggat tttgtagggg attggcgcca gacggccggc tacaatctgg
     1021 atcaggtact ggagcaaggc ggtgtttcgt cgttgtttca gaacctgggg gtgtcagtaa
     1081 cccctattca gaggatagtc ctttccggtg agaatggcct taaaattgat atccacgtca
     1141 taattcctta tgaagggttg agcggggacc aaatggggca gatagagaag attttcaagg
     1201 tagtttaccc agtcgatgac catcacttca aagttatatt gcattatggg actctggtca
     1261 tcgacggtgt cacacctaat atgattgatt acttcggccg cccatacgaa ggaatagctg
     1321 tgtttgatgg gaagaagata actgtgacgg gaactctgtg gaatggtaac aaaataatcg
     1381 acgagaggct tataaatccc gatggaagcc tcctgttccg tgtaacgatc aacggcgtaa
     1441 ctggttggcg actgtgtgag cggatacttg caggatccgg atcaggatct ggatcaggat
     1501 cgggtagtta agagctccgc tgcgcttaaa tgtctttcat tataatcaat gtataacctt
     1561 tatgtagtat ttcaatctat gccgctgtgt acgtgcactg cggtgcctat ccttcggcat
     1621 tagagagtca ctgtttgtgt agatcgtagc tgcatgtctg acatctggtt cattagtgcc
     1681 gttttgagtc tgcactgttg tgcaacttga tattcatcac tgcacaccaa ccgtgtgttt
     1741 cctcctttgg gtgcaccctt taatccttgt cttctccttt tttgtctctt tccccctcga
     1801 aaaggtgtca aactactgcc gcataaacta cggttatccc aaatttaaga gaaagcaata
     1861 aagcatcagc tagcatgcct ttgtctcaag aagaatccac cctcattgaa agagcaacgg
     1921 ctacaatcaa cagcatcccc atctctgaag actacagcgt cgccagcgca gctctctcta
     1981 gcgacggccg catcttcact ggtgtcaatg tatatcattt tactggggga ccttgtgcag
     2041 aactcgtggt gctgggcact gctgctgctg cggcagctgg caacctgact tgtatcgtcg
     2101 cgatcggaaa tgagaacagg ggcatcttga gcccctgcgg acggtgccga caggtgcttc
     2161 tcgatctgca tcctgggatc aaagccatag tgaaggacag tgatggacag ccgacggcag
     2221 ttgggattcg tgaattgctg ccctctggtt atgtgtggga gggctaactc gagtgcccat
     2281 ttagttggct tttcccttgt ctcgtgtctt ttccgtggaa aggttcccgg agtaatctga
     2341 tggcacagca gggaggtgcg cctgcaggtt ggttaggaag gggggatgat gtaaaagaag
     2401 aaaatggggg gattcgagcc ccgcacaggt ctctcaaatt ggatgcatag cttgagtatt
     2461 ctatagtgtc acctaaatag cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat
     2521 tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg
     2581 ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag
     2641 tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt
     2701 ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg
     2761 ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg
     2821 gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag
     2881 gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga
     2941 cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct
     3001 ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc
     3061 tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg
     3121 gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc
     3181 tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca
     3241 ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag
     3301 ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct
     3361 ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc
     3421 accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga
     3481 tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca
     3541 cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat
     3601 taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac
     3661 caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt
     3721 gcctgactcc ccgtcgtgta gataattacg atacgggagg gcttaccatc tggccccagt
     3781 gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag
     3841 ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct
     3901 attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt
     3961 gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc
     4021 tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt
     4081 agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg
     4141 gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg
     4201 actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct
     4261 tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc
     4321 attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt
     4381 tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt
     4441 tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg
     4501 aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta tcagggttat
     4561 tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg
     4621 cgcacatttc cccgaaaagt gccacctgat gcggtgtgaa ataccgcaca gatgcgtaag
     4681 gagaaaatac cgcatcagga aattgtaagc gttaatattt tgttaaaatt cgcgttaaat
     4741 ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat cccttataaa
     4801 tcaaaagaat agaccgagat agggttgagt gttgttccag tttggaacaa gagtccacta
     4861 ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca
     4921 ctacgtgaac catcacccta atcaagtttt ttggggtcga ggtgccgtaa agcactaaat
     4981 cggaacccta aagggagccc ccgatttaga gcttgacggg gaaagccggc gaacgtggcg
     5041 agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag tgtagcggtc
     5101 acgctgcgcg taaccaccac acccgccgca cttaatgcgc cgctacaggg cgcgtccatt
     5161 cgccattcag gctgcgcaac tgttgggaag ggcgatcggt gcgggcctct tcgctattac
     5221 gccagctggc gaaaggggga tgtgctgcaa ggcgattaag ttgggtaacg ccagggtttt
     5281 cccagtcacg acgttgtaaa acgacggcca gtgaattgta atacgactca ctata
//
