LOCUS       sequence-355613-        5342 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 5342)
  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 5342)
  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: pPOTv7.1-phleo-mNG
FEATURES             Location/Qualifiers
     source          1..5342
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(2467..2485)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2468..2485)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2503..2519)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2503..2519)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2516..2538)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2527..2543
                     /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(2551..2557,2558..2575,2576..2581))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2596..2617
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2734..2751)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2905..3493)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2985..3004)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3664..4455,4456..4524))
                     /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     4287..4306
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4525..4629)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(4707..5162)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4844..4865)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5056..5075
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5288..5310
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     5302..5319
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     5303..5319
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     join(5326..5342,1..3)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        join(5326..5342,1..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 atggctaaac
      181 tcacttctgc tgtaccagta ctcactgcta gagatgtagc tggagctgta gagttctgga
      241 ctgatagact cggattctct agagatttcg tagaggatga tttcgctgga gtagtaagag
      301 atgatgtaac tctcttcata tctgctgtac aagatcaagt agtaccagat aacactctcg
      361 cttgggtatg ggtaagagga ctcgatgagc tctacgctga gtggtctgag gtagtatcta
      421 ctaacttcag agatgcttct ggaccagcta tgactgagat aggagagcaa ccgtggggaa
      481 gagagttcgc tctcagagat ccagctggaa actgcgtaca cttcgtagct gaggagcaag
      541 attaaccatg gtgcccattt agttggcctt tcccttgtct cgtgtctttt ccgtggaaag
      601 gttcccggag taatctgatg gcacagcagg gaggtgcgcc tgcaggttgg ttaggaaggg
      661 gggatgatgt aaaagaagaa aatgggggga ttcgagcccc acgcgttgct caagctgtgt
      721 agcgcaggcg tttccttaca tatttctcta acaggcacgg aagcctaaca aatacacttg
      781 gcttattttt ttgccccctc atgtcttgta caaatatttg cgatagctta gctatcagcc
      841 acattaatca aacaagtata ccaacaagcc cgaaaacaga aactcaactg caacgggtac
      901 ctatgggttc tggtagtggt tccggttccg gttctaagct tgtctcgaaa ggtgaggaag
      961 acaatatggc cagcttgcca gcaacgcacg agcttcatat atttgggtcg atcaacggtg
     1021 tggatttcga catggtgggt caaggcacag gtaatcctaa tgacgggtat gaagaactca
     1081 atttgaagag cactaaggga gatctccagt tttccccgtg gattcttgtg ccgcatattg
     1141 gttatggttt tcatcagtac ttaccatacc ccgatggaat gtcgcccttc caagcagcta
     1201 tggtggacgg ctcggggtat caagtacatc gcaccatgca atttgaggac ggagcatccc
     1261 tcaccgtaaa ctaccgctat acgtatgagg ggagccacat aaaaggtgag gctcaagtta
     1321 aaggcacagg cttcccagct gacggcccgg tcatgaccaa ttcattaacc gctgctgatt
     1381 ggtgccgttc taagaagacc taccctaacg acaagacgat cattagtact tttaagtgga
     1441 gttataccac aggaaacgga aagcggtatc gaagcactgc tcgaacaacc tacacatttg
     1501 cgaagccaat ggccgcgaac tacttgaaga accagccaat gtatgtgttt cgaaaaacgg
     1561 aactgaagca ttcgaaaacc gagcttaatt ttaaagagtg gcagaaggcg tttacagacg
     1621 tgatggggat ggacgaattg tataagggat ccggatcagg atctggatca ggatcgggta
     1681 gttaagagct ccgctgcgct taaatgtctt tcattataat caatgtataa cctttatgta
     1741 gtatttcaat ctatgccgct gtgtacgtgc actgcggtgc ctatccttcg gcattagaga
     1801 gtcactgttt gtgtagatcg tagctgcatg tctgacatct ggttcattag tgccgttttg
     1861 agtctgcact gttgtgcaac ttgatattca tcactgcaca ccaaccgtgt gtttcctcct
     1921 ttgggtgcac cctttaatcc ttgtcttctc cttttttgtc tctttccccc tcgaaaaggt
     1981 gtcaaactac tgccgcataa actacggtta tcccaaattt aagagaaagc aataaagcat
     2041 cagctagcat ggctaaactc acttctgctg taccagtact cactgctaga gatgtagctg
     2101 gagctgtaga gttctggact gatagactcg gattctctag agatttcgta gaggatgatt
     2161 tcgctggagt agtaagagat gatgtaactc tcttcatatc tgctgtacaa gatcaagtag
     2221 taccagataa cactctcgct tgggtatggg taagaggact cgatgagctc tacgctgagt
     2281 ggtctgaggt agtatctact aacttcagag atgcttctgg accagctatg actgagatag
     2341 gagagcaacc gtggggaaga gagttcgctc tcagagatcc agctggaaac tgcgtacact
     2401 tcgtagctga ggagcaagat taactcgagg cacaggtctc tcaaattgga tgcatagctt
     2461 gagtattcta tagtgtcacc taaatagctt ggcgtaatca tggtcatagc tgtttcctgt
     2521 gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa
     2581 agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc
     2641 tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag
     2701 aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt
     2761 cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga
     2821 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg
     2881 taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa
     2941 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt
     3001 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct
     3061 gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct
     3121 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc
     3181 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt
     3241 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc
     3301 tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat
     3361 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa
     3421 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa
     3481 aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga
     3541 aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct
     3601 tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga
     3661 cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc
     3721 catagttgcc tgactccccg tcgtgtagat aattacgata cgggagggct taccatctgg
     3781 ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat
     3841 aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat
     3901 ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg
     3961 caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc
     4021 attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa
     4081 agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc
     4141 actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt
     4201 ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag
     4261 ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt
     4321 gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag
     4381 atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac
     4441 cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc
     4501 gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca
     4561 gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg
     4621 ggttccgcgc acatttcccc gaaaagtgcc acctgatgcg gtgtgaaata ccgcacagat
     4681 gcgtaaggag aaaataccgc atcaggaaat tgtaagcgtt aatattttgt taaaattcgc
     4741 gttaaatttt tgttaaatca gctcattttt taaccaatag gccgaaatcg gcaaaatccc
     4801 ttataaatca aaagaataga ccgagatagg gttgagtgtt gttccagttt ggaacaagag
     4861 tccactatta aagaacgtgg actccaacgt caaagggcga aaaaccgtct atcagggcga
     4921 tggcccacta cgtgaaccat caccctaatc aagttttttg gggtcgaggt gccgtaaagc
     4981 actaaatcgg aaccctaaag ggagcccccg atttagagct tgacggggaa agccggcgaa
     5041 cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt
     5101 agcggtcacg ctgcgcgtaa ccaccacacc cgccgcactt aatgcgccgc tacagggcgc
     5161 gtccattcgc cattcaggct gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg
     5221 ctattacgcc agctggcgaa agggggatgt gctgcaaggc gattaagttg ggtaacgcca
     5281 gggttttccc agtcacgacg ttgtaaaacg acggccagtg aattgtaata cgactcacta
     5341 ta
//
