LOCUS       sequence-100249-        5843 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Inserts a puromycin resistance gene and a VSG117 gene into the 
            VSG221 expression site of T.brucei downstream of the expression site
            promoter..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5843)
  AUTHORS   Smith TK, Vasileva N, Gluenz E, Terry S, Portman N, Kramer S, 
            Carrington M, Michaeli S, Gull K, Rudenko G
  TITLE     Blocking variant surface glycoprotein synthesis in Trypanosoma 
            brucei triggers a general arrest in translation initiation.
  JOURNAL   PLoS One. 2009 Oct 26;4(10):e7532. doi: 
            10.1371/journal.pone.0007532.
  PUBMED    19855834
REFERENCE   2  (bases 1 to 5843)
  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: "PLoS One.";
            date: "2009-10-26"; volume: "4(10):e7532. doi"; pages: " 
            10.1371/journal.pone.0007532"
COMMENT     Sequence Label: p221- purVSG117UTR
FEATURES             Location/Qualifiers
     source          1..5843
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    30..46
                     /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     35..57
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     54..70
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     54..70
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        91..109
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     CDS             1740..2339
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(1740..1759)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     2236..2256
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     RBS             4084..4092
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     primer_bind     complement(4970..4986)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4970..4986)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(5756..5775)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(5757..5775)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5782..5799)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5782..5798)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5791..5813)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 tatgcttccg gctcgtatgt tgtgtggaat tgtgagcgga taacaatttc acacaggaaa
       61 cagctatgac catgattacg ccaagctcga aattaaccct cactaaaggg aacaaaagct
      121 ggagctccac cgcggtggcg gccgctctag aactagtggg aaatatttat ttgttatttt
      181 ttcttttatt agctggttct tagggtgatt tatgtttccg gattaaacta acaagctgat
      241 agtttggttt acttattgtc tttctgtggg gaggaatgtt ggtgatttgt tcggagaata
      301 tttcggatgc ttttttttta ctagatgagg acatcaactc gccaattgtg gatgggagtt
      361 tgttggctga atagtattat ggatcaggtc agaggcatgt ttttcagtgt tacggaccgc
      421 gtgtggatga aacactagaa tgtgttgcct tttgtcttga gtatgaattt ggccgtaacg
      481 attgttgttg cagtatgttt ttcttcaaca aaaaaaagtt tcaatgaaat ttgtaatcca
      541 ctttaatacg gttacaactt tcgcatttac aactgatgct atatcgatct gacgactaac
      601 cttgatgtta aataggtaag tttttacaga tgctaggctc caatttatag tgaagtaagc
      661 aatcgtaaag ttggtgattc ttcctttatg ggtatgaggt aaggaatatc gacgtgcata
      721 tcactaaagc cttctttagt acttttcaca gggttagaat cactataatg ctgagcgata
      781 tttcggtttt ttactaagaa aagacactgg ataatacatt tgtcgcagcn tttgatatga
      841 taccttattt cccnatttgt ttatttttaa gagttatgag gcgttctttt cacacgagaa
      901 acacgtgata aatgaaggtc ttgcgctgtc ttaattgtta actgtgctct cggagagaga
      961 gagggtggga tttttttctt taaataaagg gagatgtatt cggaggagtt gcattaacgt
     1021 tgtcccatgt tttacaaaaa acgagtctga aagaacgccg actcaatggc catccaagcg
     1081 gaataacctt ctaaaagaat catatcccta ttaccacacc agtttatatt acaggggagg
     1141 ttattacaga aatctaggat atcagactca aggtgtggcg gacgtctcga accgatcgtg
     1201 agttggcctg agcgtgccgt cgttaccgga gtgcccgtat tcgatttatg tcgactgtgt
     1261 ggctgcgaaa acccttagcg ttaccactgt gacgattgcg tgcgatgcaa agagaccgtt
     1321 gtcggttcat gcatgacttt aatctcatct acgctttatc ccgtggttcc ttcgtccatt
     1381 gaggttgggc ccccctcgac tagatgggtc ccattgtttg cctcttccgc tgtgtggagt
     1441 gcgcctacac gcacttctca cttcgtaagt ggtggtggcg taagtattgc ctaatgttga
     1501 ctctatgttc tcctctcctc accccctcgc ggtgctgatt tctgacagat cttcaaacac
     1561 tagtttaagc aaaggactat tcatccgttt atattagcaa cagtaggtac tagcaccact
     1621 aacaacaaca acaaagcact tctgcaggaa ttaattcgat agcttgatgg ccgccacgac
     1681 cggtgccgcc accatcccct gacccacgcc cctgacccct cacaaggaga cgaccttcca
     1741 tgaccgagta caagcccacg gtgcgcctcg ccacccgcga cgacgtcccc cgggccgtac
     1801 gcaccctcgc cgccgcgttc gccgactacc ccgccacgcg ccacaccgtc gacccggacc
     1861 gccacatcga gcgggtcacc gagctgcaag aactcttcct cacgcgcgtc gggctcgaca
     1921 tcggcaaggt gtgggtcgcg gacgacggcg ccgcggtggc ggtctggacc acgccggaga
     1981 gcgtcgaagc gggggcggtg ttcgccgaga tcggcccgcg catggccgag ttgagcggtt
     2041 cccggctggc cgcgcagcaa cagatggaag gcctcctggc gccgcaccgg cccaaggagc
     2101 ccgcgtggtt cctggccacc gtcggcgtct cgcccgacca ccagggcaag ggtctgggca
     2161 gcgccgtcgt gctccccgga gtggaggcgg ccgagcgcgc cggggtgccc gccttcctgg
     2221 agacctccgc gccccgcaac ctccccttct acgagcggct cggcttcacc gtcaccgccg
     2281 acgtcgaggt gcccgaagga ccgcgcacct ggtgcatgac ccgcaagccc ggtgcctgac
     2341 gcccgcccca cgacccgcag cgcccgaccg aaaggagcgc acgaccccat ggggggatca
     2401 tcatcgatga attaattcga gctcggtacc cctagaaagt gtgacaacgt cgcaccatgt
     2461 gtaggttttc atttatgttc tttctttctt tctttnntgt gaatttgttt tctgtctcaa
     2521 atgtttttaa ttcgcttggg acctatgttt ttcttgtttt tttgctcacc ctttgtgtag
     2581 gaggcaccct gtcacgtctg tggttgcgtg tatgccttcc ttccccttat tcgcttcttc
     2641 ctgtcgtgtc acacctcttt ctccctctcc ctttccgcct tttctttcaa tcttgttttc
     2701 tcgaccagcc ctactagagg agaaagaata gtaacccttt catcaaagaa aatagtcgaa
     2761 gcttatggac tgccatacaa aggagacact aggggtcaca caatggaggc gatcaacgat
     2821 gctaacacta tcactgcttt acgccatcac tccagcggac ggcgccaaag aagcccttga
     2881 atacaaaact tggacaaacc actgcggact ggcggccaca ctgagaaagg ttgccggtgg
     2941 agtattaacg aaactgaaaa gccacattag ctaccggaaa aaactggaag aaatggaaac
     3001 gaagctacga atctacgcac taaaaggaga cggagtggga gagcaaaaat cagcggagat
     3061 actagcaaca acggcagccc taatgcgaca aaaagcactc acaccagaag aagcaaattt
     3121 gaaaacagcg ctgaaggcgg caggattcgc aggcgaagga gcggcagccg tcagcagcta
     3181 cctgatgaca ctcgggacac tgacaacaag cggatctgcg cactgcctaa gcaacgaagg
     3241 cggcgacggt gacggaaaag accaacttgc gccgaaaggc tgccggcacg gcacagaagc
     3301 agacttcgac gcaggagccg gcccggcaga atctgaagta gccgacagcg gcttcgcgca
     3361 agtaccaggc aaacaggacg gagcaaacgc aggccaagca aacatgtgcg cattgttcac
     3421 acaccaagca acgccgcaca gctcacaggg catatacata accggggcac agacaaaacc
     3481 ttcattcggg tacggcatgc tgacaatcgg cacgacggac cagaccatcg gcttgaaact
     3541 ttcggacata aagggcaaac aagcagacag cgcgcagaaa ttctggagca gctgccacgc
     3601 agcagtcaaa gccgcccagg atatgaaggc agacccagcc ctaaaggtcg accagacgct
     3661 cctagctgtt cttgtggctt ctccggagat ggcggaaata ctgaaactag aagcggcagc
     3721 atcacagcaa aaaggaccag aggaagtgac gatcgacctg gccaccgaga aaaacaatta
     3781 tttcggaacc aacaacaaca aactagagcc gctctggact aaaatcaaag gacagaatat
     3841 agttgacctg gcggcgacca aaggcagcac gaaagagtta ggaacagtca cagacacggc
     3901 cgagctacaa aaacttttaa gttattatta cacggtcaac aaagaagaac agaaaaaaac
     3961 agcggagaaa ataactaaac tcgaaaccga actagcagat caaaaaggca aatcccctga
     4021 aagcgagtgc aataaaatat ctgaggaacc caaatgcaac gaggacaaga tatgcagttg
     4081 gcataaggag gttaaagcgg gagaaaagca ctgcaaattt aactcaacaa aagcaaaaga
     4141 aaagggggtc tctgtaacac aaactcaaac tgcaggagga accgaagcga caacagataa
     4201 atgcaaaggg aaattgggag atacctgcaa gaaggagagc aactgcaaat gggaaaataa
     4261 tgcttgcaaa gattcctcta ttctagtaac caaaaaattc gccctcagtg ctgctgcgtt
     4321 tgcggccttg cttttttagg aattcctgca gccctttccc ccctcaaatt tcccccctcc
     4381 ttttaaaatt ttccttgcta cttgaaaact ttttgatata ttttaacacc aaaaccagcc
     4441 gagattttgt gttctgtgtt ttgtaagttg actgtctgat tgtctagaaa tattttctgg
     4501 caactaaaat ttttttcttt tttcctgttt tttttgtagg taggtaggaa tggggggggg
     4561 ggtagttagg taggttagtt aggttagtta gggggttagt taggggggtt aggcttagga
     4621 ttaggattag acttaggctt aggattagga ttaggattag gattagggtt aattttttcc
     4681 tctttttttt taactcacac ctctatcctg gatttttaat tttttttttt agccattcgc
     4741 ggctcctttt tttttttttg cgccaatgtt taatttttta ttgtgttttc aatttttttg
     4801 tcaaccatgc agcggctgtt ttgttatgcg gaccctaacc ctcctccccc ccccccgccc
     4861 gcgcacctcc atttttaaaa atttttttac cgcgtccttc aaccagaatt tttttaaatt
     4921 ttttaatttt ttttattttc cgtggttttg aatcttaatt tttcgggggg atccactagt
     4981 tctagagcgg ccagcttatc gataccgtca cgaagagcag ggggtgcaac ggaagagtga
     5041 aaggattaga ggaaatggtg gaattagcat gatcggagat tgtgagggtt aggaaaaacg
     5101 tgaagaggga aatggtaact ggacatcgtg cgcgaggatg gcgaaggctt gaagcgtgta
     5161 gagagcgtgt gtaaatggcc caaacggtgt gggtggggta ataacgcaga gtgctgaggt
     5221 gtttgacaag tgcccgcgag gggtgctggg gaggaggaag caagacagta aaatcgtcgt
     5281 gagacggtgg ctgtgctcgg gaatccgtac aaatatactg aaaattattg atggttgata
     5341 tgcgtgggaa cgacgcagga tgatgctgca tcgccaagat gcttcgggag aacaggaaaa
     5401 taacgacgaa tcaacactgg tgatttgata ctataaaaat cgcgtgggct ttgctgcttc
     5461 gctttcataa atgccacctg caatgaattt ggagggtgga ggtgagctgc cctgtgtatg
     5521 ggtgggtatt gcagatgcag cggcgctttt gggtgatttt tggtttacag tttgctgttg
     5581 ttgggaaggt tggtggtggc aacaaaattg gagaaaatga tttcatctgg aaacctactt
     5641 caaaacccaa taaacaatcg gacctcgaag tgcaagtgtc tcccagaacg taaacaaggc
     5701 cttgaagacg accagcagat gaagcacctc gagggggggc ccggtaccca attcgcccta
     5761 tagtgagtcg tattacaatt cactggccgt cgttttacaa cgtcgtgact gggaaaaccc
     5821 tggcgttacc caacttaatc gcc
//
