LOCUS       sequence-120526-        6075 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Complementation in Cryptococcus neoformans. Targets DNA constructs 
            to a Safe Haven Site, a small gene-free region. Contains NEO 
            resistance marker..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6075)
  AUTHORS   Arras SD, Chitty JL, Blake KL, Schulz BL, Fraser JA
  TITLE     A genomic safe haven for mutant complementation in Cryptococcus 
            neoformans.
  JOURNAL   PLoS One. 2015 Apr 9;10(4):e0122916. doi: 
            10.1371/journal.pone.0122916. eCollection 2015.
  PUBMED    25856300
REFERENCE   2  (bases 1 to 6075)
  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: "2015-04-9"; volume: "10(4):e0122916. doi"; pages: " 
            10.1371/journal.pone.0122916. eCollection 2015"
COMMENT     Sequence Label: pSDMA57
FEATURES             Location/Qualifiers
     source          1..6075
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(1809..2609)
                     /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="MSMIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQG
                     RPVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDL
                     LSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEE
                     HQGLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQD
                     IALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(1920..1939)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     2530..2549
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     CDS             complement(3565..3930)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSAQLTLTKGNKSWSSTAVAAALELVDPPGCRNSISSLSI
                     PSTSRGGPVPNSPYSESYYARSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEAR
                     TDRPSQQLRSLNGEWKL"
     primer_bind     3699..3721
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3713..3730
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3714..3730
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3740..3759
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3740..3758
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    3767..3874
                     /label=MCS
                     /note="pBluescript multiple cloning site"
     primer_bind     3783..3799
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     3784..3800
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3834..3850)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3834..3850)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(3887..3907)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(3887..3905)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3926..3942)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3926..3942)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3939..3961)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3950..3966
                     /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(3974..3980,3981..3998,3999..4004))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4019..4040
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4157..4174)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4328..4916)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4408..4427)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5087..5878,5879..5947))
                     /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     5710..5729
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5948..6052)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ctcatcagat ctgcccatag acttcaaatc atggaattgg acgtttatct tcttgctttg
       61 gatatccacc ctatcgtaca atcttggcac tttggatcat acgaggcact ttgcggacta
      121 gggaatggtt atcatcaatt tcttggacct ctccatgaca cctggcatcg tcgagctacc
      181 acagccccct gaattcttaa aggcgaaccc cccttcctag gaaccttgag cgctggactg
      241 gatggcactc tgagaccgcc tatgcttaag gacgcgagct tacccttcca ttgatggaag
      301 tcgatcattt atgataatag aacacggcgg aacttgactt tacaggccaa gggaagcaac
      361 aggagaagaa gctaaggtta taacaggctc agcatatttg tatatacggt gccattcaac
      421 gctgaacact ttgtactcac ggtctgcccg ttgaataaca ccatctgctg atgatcatcc
      481 caatttgtga gtcttatttg ccattgatcc tggtgtataa ccgatactga agccctttag
      541 gtctgatcag ttatagacgt ggtatattca ttcttcctag attagctatg ccgaaagaga
      601 tggatacctt gatgtgactg gaatgccatg aagcaggaga ttgtagcagc tcaaattgga
      661 gaaggatgtt gcaaccgtcg gccgtgaagg tcttgtcata tctttgtctg cctgggagaa
      721 tatgtacaag tgccccgatg ttactgatgg aaaacatcgt gtggcgcgcc acgatacttg
      781 tgttaattaa accaaaccag tctgtgccaa gtggattgct tatgcatcga gtggcgagcc
      841 cactgacctc tgggcccctt ttttgaacta taaaaagcgg ccaagtctga tttcggtctc
      901 acaatcatca acatcgactt ctcttttcgc ctcgatcaac ctaaacaacg gtcactcagc
      961 catgaactgg acaatggaat atttcgcctt gccttggatc tttggcctct agactcaatc
     1021 aacgtcagga taaaggtgag cataggttgc tcactcactg tagccttttt ttcccttctt
     1081 cttggtccag atggttccca aggctctata aagtgtatct gtcagttcca cttccttact
     1141 ccgaagcgat tcagagttgt aaagtgcgtc tgaccccctg attatatctc aaagtccctt
     1201 cacaaattca gacatgatgc gataaagtgc cggattgtca ttcgcgtctt gtgactgtat
     1261 agaaagtgcc ggcatcctct tttcttttcc tctttggccg agcgactcag agttgtgaag
     1321 cctgctggta tctgtctgtt cccttgccct gataaagaat atcgaagagc atttctttga
     1381 ttcacaatac gttgacgaat ggtgtgatgg atcctaagca gcgccagccc tttgaacttg
     1441 tgtcgaaaaa aaaaaaaaaa cagcatatga tacgtaaagg gaagaaaagg atggagctga
     1501 ctgttttgac aggcataatt ttgagcgtgc ttcattggcc ggtttatctg tattaacacg
     1561 gaagagatgt agaaactagc ttcctggttt cagagacaag gagccatgaa gatcctgagg
     1621 attcaacggc gttgctgagg gtgggaagga tggtattgtt gtctgtagat agattaatgg
     1681 gcatataatg tcaaaaaaga tgattgacac gatgtctgtc tcgaattcgc agtgtgctaa
     1741 aattgtatca cttctctaat atgtatatat acaccctcta aggaaaatta accccttacc
     1801 gccttcactc agaagaactc gtcaagaagg cgatagaagg cgatgcgctg cgaatcggga
     1861 gcggcgatac cgtaaagcac gaggaagcgg tcagcccatt cgccgccaag ctcttcagca
     1921 atatcacggg tagccaacgc tatgtcctga tagcggtccg ccacacccag ccggccacag
     1981 tcgatgaatc cagaaaagcg gccattttcc accatgatat tcggcaagca ggcatcgcca
     2041 tgggtcacga cgagatcctc gccgtcgggc atgcgcgcct tgagcctggc gaacagttcg
     2101 gctggcgcga gcccctgatg ctcttcgtcc agatcatcct gatcgacaag accggcttcc
     2161 atccgagtac gtgctcgctc gatgcgatgt ttcgcttggt ggtcgaatgg gcaggtagcc
     2221 ggatcaagcg tatgcagccg ccgcattgca tcagccatga tggatacttt ctcggcagga
     2281 gcaaggtggg atgacaggag atcctgcccc ggcacttcgc ccaatagcag ccagtccctt
     2341 cccgcttcag tgacaacgtc gagcacagct gcgcaaggaa cgcccgtcgt ggccagccac
     2401 gatagccgcg ctgcctcgtc ctgcagttca ttcagggcac cggacaggtc ggtcttgaca
     2461 aaaagaaccg ggcgcccctg cgctgacagc cggaacacgg cggcatcaga gcagccgatt
     2521 gtctgttgtg cccagtcata gccgaatagc ctctccaccc aagcggccgg agaacctgcg
     2581 tgcaatccat cttgttcaat catagacatg ttgggcgagt ttactaatgg aaaaagacaa
     2641 ttagcaaaga ggctgcatat taggagacgc ggtgaagatt ggggaggaca atagggtccg
     2701 tgcaccaaaa agtcctgacg gcagctctgt tcaacgcaca tccatctctt ccagctatgc
     2761 gtgtggcacc agcacggatg gattgttgca ccaaaaggac aaggcgagat ggatgcaggt
     2821 cgaggcgatg cggtgctatc ggccctcgtt gtcgaacgcc acgtcgtcct ctcggcgacc
     2881 gtcaatcctg gaaggaaatg accaagcgtg cacccagcat ccagcatcca acatccagca
     2941 cccagcacct agcacccatg atccaatggc cggcatgcag atcagcgtgg ggagagcacc
     3001 atctagcagc ccctggcagg cccagcattg tgctcggatg tgcgataggc ggtgggtgcg
     3061 ataaggtgga ttagtggagg atggaggact gagtggcggg caagggaagg gggcgggaag
     3121 atgactcacc ttgggcagtg ggtgggggat gcggtatagg ggatgtatgt ggatgctggc
     3181 ggaggataga agctgtaagc tgagctggtg ccaacggtga cgctgtgaga gtggtctcgg
     3241 cgaccctgac ggcgactcgg cgttaccccg tgtggcacac cggacgcgcg tcgagtgacg
     3301 ttgctgctat tgttgttacc atcatcctct cctccgacaa ccatacactc agtgcacacc
     3361 atgccactcg aatcctgcat gcttatgtga gtcctcccct tcccgtgcgc cgctctccag
     3421 ctcacatcct cgcagccatg gactcaaccc tatctcggtc tattcttttg atttataagg
     3481 gattttgccg atttcggcct attggttaaa aaaatgagct gatttaacaa aaatttaacg
     3541 cgaattttaa caaaatatta acgcttacaa tttccattcg ccattcaggc tgcgcaactg
     3601 ttgggaaggg cgatcggtgc gggcctcttc gctattacgc cagctggcga aagggggatg
     3661 tgctgcaagg cgattaagtt gggtaacgcc agggttttcc cagtcacgac gttgtaaaac
     3721 gacggccagt gagcgcgcgt aatacgactc actatagggc gaattgggta ccgggccccc
     3781 cctcgaggtc gacggtatcg ataagcttga tatcgaattc ctgcagcccg ggggatccac
     3841 tagttctaga gcggccgcca ccgcggtgga gctccagctt ttgttccctt tagtgagggt
     3901 taattgcgcg cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc
     3961 tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat
     4021 gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc
     4081 tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg
     4141 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag
     4201 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag
     4261 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
     4321 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
     4381 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
     4441 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
     4501 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg
     4561 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
     4621 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
     4681 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
     4741 ggtggcctaa ctacggctac actagaagga cagtatttgg tatctgcgct ctgctgaagc
     4801 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
     4861 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
     4921 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
     4981 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa
     5041 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa
     5101 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc
     5161 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga
     5221 taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa
     5281 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt
     5341 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg
     5401 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc
     5461 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg
     5521 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag
     5581 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt
     5641 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt
     5701 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac
     5761 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac
     5821 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag
     5881 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa
     5941 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga
     6001 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc
     6061 cccgaaaagt gccac
//
