LOCUS       sequence-71840-e        3351 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Gateway adapted transfer of Neomycin resistance gene.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3351)
  AUTHORS   Xu D, Brandan CP, Basombrio MA, Tarleton RL
  TITLE     Evaluation of high efficiency gene knockout strategies for 
            Trypanosoma cruzi.
  JOURNAL   BMC Microbiol. 2009 May 11;9:90. doi: 10.1186/1471-2180-9-90.
  PUBMED    19432966
REFERENCE   2  (bases 1 to 3351)
  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: "BMC 
            Microbiol."; date: "2009-05-11"; volume: "9:90. doi"; pages: " 
            10.1186/1471-2180-9-90"
COMMENT     Sequence Label: pDONR221-Neo
FEATURES             Location/Qualifiers
     source          1..3351
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(106..125)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(107..125)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(130..146)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(130..146)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(248..267)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             259..1068
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(326..345)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      1215..1803
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1704..1723
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1957..1974
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(2133..2160)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(2252..2338)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     2387..2409
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2401..2418
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2402..2418
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    2434..2533
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             2537..3340
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MGSAIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQ
                     GRPVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQD
                     LLSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDE
                     EHQGLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQ
                     DIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2600..2619)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3210..3229
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     protein_bind    complement(join(3341..3351,1..89))
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
ORIGIN
        1 ttgtacaaag ttggcattat aagaaagcat tgcttatcaa tttgttgcaa cgaacaggtc
       61 actatcagtc aaaataaaat cattatttgc catccagctg atatccccta tagtgagtcg
      121 tattacatgg tcatagctgt ttcctggcag ctctggcccg tgtctcaaaa tctctgatgt
      181 tacattgcac aagataaaat aatatcatca tgaacaataa aactgtctgc ttacataaac
      241 agtaatacaa ggggtgttat gagccatatt caacgggaaa cgtcgaggcc gcgattaaat
      301 tccaacatgg atgctgattt atatgggtat aaatgggctc gcgataatgt cgggcaatca
      361 ggtgcgacaa tctatcgctt gtatgggaag cccgatgcgc cagagttgtt tctgaaacat
      421 ggcaaaggta gcgttgccaa tgatgttaca gatgagatgg tcagactaaa ctggctgacg
      481 gaatttatgc ctcttccgac catcaagcat tttatccgta ctcctgatga tgcatggtta
      541 ctcaccactg cgatccccgg aaaaacagca ttccaggtat tagaagaata tcctgattca
      601 ggtgaaaata ttgttgatgc gctggcagtg ttcctgcgcc ggttgcattc gattcctgtt
      661 tgtaattgtc cttttaacag cgatcgcgta tttcgtctcg ctcaggcgca atcacgaatg
      721 aataacggtt tggttgatgc gagtgatttt gatgacgagc gtaatggctg gcctgttgaa
      781 caagtctgga aagaaatgca taaacttttg ccattctcac cggattcagt cgtcactcat
      841 ggtgatttct cacttgataa ccttattttt gacgagggga aattaatagg ttgtattgat
      901 gttggacgag tcggaatcgc agaccgatac caggatcttg ccatcctatg gaactgcctc
      961 ggtgagtttt ctccttcatt acagaaacgg ctttttcaaa aatatggtat tgataatcct
     1021 gatatgaata aattgcagtt tcatttgatg ctcgatgagt ttttctaatc agaattggtt
     1081 aattggttgt aacactggca gagcattacg ctgacttgac gggacggcgc aagctcatga
     1141 ccaaaatccc ttaacgtgag ttacgcgtcg ttccactgag cgtcagaccc cgtagaaaag
     1201 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
     1261 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
     1321 aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag
     1381 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
     1441 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
     1501 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
     1561 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
     1621 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
     1681 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
     1741 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
     1801 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac
     1861 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga
     1921 gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg
     1981 gaagagcgcc caatacgcaa accgcctctc cccgcgcgtt ggccgattca ttaatgcagc
     2041 tggcacgaca ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat taatacgcgt
     2101 accgctagcc aggaagagtt tgtagaaacg caaaaaggcc atccgtcagg atggccttct
     2161 gcttagtttg atgcctggca gtttatggcg ggcgtcctgc ccgccaccct ccgggccgtt
     2221 gcttcacaac gttcaaatcc gctcccggcg gatttgtcct actcaggaga gcgttcaccg
     2281 acaaacaaca gataaaacga aaggcccagt cttccgactg agcctttcgt tttatttgat
     2341 gcctggcagt tccctactct cgcgttaacg ctagcatgga tgttttccca gtcacgacgt
     2401 tgtaaaacga cggccagtct taagctcggg ccccaaataa tgattttatt ttgactgata
     2461 gtgacctgtt cgttgcaaca cattgatgag caatgctttt ttataatgcc aactttgtac
     2521 aaaaaagcag gctaccatgg gatcggccat tgaacaagat ggattgcacg caggttctcc
     2581 ggccgcttgg gtggagaggc tattcggcta tgactgggca caacagacaa tcggctgctc
     2641 tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg gttctttttg tcaagaccga
     2701 cctgtccggt gccctgaatg aactgcagga cgaggcagcg cggctatcgt ggctggccac
     2761 gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa gggactggct
     2821 gctattgggc gaagtgccgg ggcaggatct cctgtcatct caccttgctc ctgccgagaa
     2881 agtatccatc atggctgatg caatgcggcg gctgcatacg cttgatccgg ctacctgccc
     2941 attcgaccac caagcgaaac atcgcatcga gcgagcacgt actcggatgg aagccggtct
     3001 tgtcgatcag gatgatctgg acgaagagca tcaggggctc gcgccagccg aactgttcgc
     3061 caggctcaag gcgcgcatgc ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg
     3121 cttgccgaat atcatggtgg aaaatggccg cttttctgga ttcatcgact gtggccggct
     3181 gggtgtggcg gaccgctatc aggacatagc gttggctacc cgtgatattg ctgaagagct
     3241 tggcggcgaa tgggctgacc gcttcctcgt gctttacggt atcgccgctc ccgattcgca
     3301 gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga acccagcttt c
//
