LOCUS       sequence-242239-        6882 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid for tagging proteins in Cryptococcus neoformans; C-terminal 
            fragment of Venus protein.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6882)
  AUTHORS   So YS, Yang DH, Jung KW, Huh WK, Bahn YS
  TITLE     Molecular Characterization of Adenylyl Cyclase Complex Proteins 
            Using Versatile Protein-Tagging Plasmid Systems in Cryptococcus 
            neoformans.
  JOURNAL   J Microbiol Biotechnol. 2017 Feb 28;27(2):357-364. doi: 
            10.4014/jmb.1609.09036.
  PUBMED    27780958
REFERENCE   2  (bases 1 to 6882)
  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"; doi: "10.4014/jmb.1609"; 
            journalName: "J Microbiol Biotechnol"; date: "2017-02-28- 28"; 
            volume: "27"; issue: "2"; pages: "357-364"
COMMENT     Sequence Label: YSBE418:pNAT_ACA1_Vc_HOG1 terminator
FEATURES             Location/Qualifiers
     source          1..6882
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    107..128
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(143..148,149..166,167..173)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    181..197
                     /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     186..208
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     205..221
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     205..221
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             1192..1443
                     /codon_start=1
                     /product="C-terminal fragment of mVenus for use in 
                     bimolecular fluorescence complementation (BiFC) (Kodama and
                     Hu, 2010)"
                     /label=VC155
                     /translation="DKQKNGIKANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNH
                     YLSYQSALSKDPNEKRDHMVLLEFVTAAGITLGMDELYK"
     primer_bind     1380..1401
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             complement(2015..2578)
                     /codon_start=1
                     /gene="Streptomyces noursei nat1"
                     /product="nourseothricin acetyltransferase"
                     /label=NrsR
                     /note="confers resistance to nourseothricin"
                     /translation="TLDDTAYRYRTSVPGDAEAIEALDGSFTTDTVFRVTATGDGFTLR
                     EVPVDPPLTKVFPDDESDDESDDGEDGDPDSRTFVAYGDDGDLAGFVVVSYSGWNRRLT
                     VEDIEVAPEHRGHGVGRALMGLATEFARERGAGHLWLEVTNVNAPAIHAYRRMGFTLCG
                     LDTALYDGTASDGEQALYMSMPCP"
     primer_bind     complement(3465..3482)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3465..3481)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3474..3496)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             3619..3921
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="QFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDKV
                     SRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     3800..3819
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             4270..5064
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             4270..5064
                     /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="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4324..4343)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4934..4953
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     CDS             join(5082..5150,5151..5942)
                     /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     complement(5300..5319)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6113..6701
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6602..6621
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
       61 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc
      121 tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa
      181 ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac gccaagcttg
      241 gtaccgagct cggatccatc tgcttattcc gagcgtcgct aactctgcta actctcacag
      301 ggttccccgg cccccccttc tatgcccaag gcatcagcat ctgctccttc tccacctcct
      361 ccccctcctt ctggctcagc accggctgct cctacatctg gttctggagc tgctgccctt
      421 ttggctgatt tgaatcgtgg tggcgctgtc acttctggat tgcgaaaggt cgatagttcc
      481 cagatgaccc acaagaaccc ctcacttcgt tctgccggca ctgtctctga caatgcaaag
      541 aagggcccgc ctctcaaacc caagcctggg gcaaagcctg caaagaagcc tgcgaagatc
      601 gagcttgaag atgggaacaa atggatcatt gttcgtctct tttctttgtc ctttcatggt
      661 ctcgttttgt gactgactta taacaggaaa accaagagga caacaaatct attaaaatcg
      721 acaacactga acttcaccac actgttcaca tctttggttg cgtcaactct gtggttcaga
      781 tctccggtaa aatcaacgct gtaaccatgg gtaagtatag tcaatgctct tagcacatat
      841 gcatatagct gataactcgc agctggatgt aagaagacgt ctgtcgtcct cgacaccgcc
      901 gtatcctctt tctctatcac ttcgtccccc tcctttgaag tccagatcat cggctccatt
      961 cccaccatcc aaatcgacac cactgactct ggccaggtat acctctcaaa ggattgtatg
     1021 gaagtggttg aaatcgttac cagcaagtcc agtagcatca acatttctgt gcccaccggc
     1081 gaagatggtg atttcgtcga aaggcctgtg cctgagcaga tgaagtcaag gataatcgac
     1141 ggaaaactgg ttaccgagat tgtagagcac tcaggaggtt ctggtggtac tgacaagcag
     1201 aagaacggca tcaaggccaa cttcaagatc cgccacaaca tcgaggacgg cggcgtgcag
     1261 ctcgccgacc actaccagca gaacaccccc atcggcgacg gccccgtgct gctgcccgac
     1321 aaccactacc tgagctacca gtccgccctg agcaaagacc ccaacgagaa gcgcgatcac
     1381 atggtcctgc tggagttcgt gaccgccgcc gggatcactc tcggcatgga cgagctgtac
     1441 aagtagggcc gcgtaaaaga atcgttcttc tttctttctt tctttttatt tataacgaaa
     1501 gaagagatgt tggcgtatac ttgaacttat gaatgatgtg ttgtattcag gagcctgaaa
     1561 gtaatgtggt gtgtggtggg cggtgggacg aggaagagtc gggataagtt gggatgatag
     1621 atggtggagc aagattgcat aagcaaaagt tgagaagcgc aaagttgaga agaagtgtgg
     1681 acggctcgtg ttgcgggttc agggagcaga agggagggaa tgtggagatt gttcggagag
     1741 agggatagat tggtacggat ccactagtaa cggccgccag tgtgctggaa ttcgcccttg
     1801 gagccatgaa gatcctgagg attcaacggc gttgctgagg gtgggaagga tggtattgtt
     1861 gtctgtagat agattaatgg gcatataatg tcaaaaaaga tgattgacac gatgtctgtc
     1921 tcggattcgc agtgtgctaa aattgtatca cttctctaat atgtatatat acaccctcta
     1981 aggaaaatta accccttacc gccttcacga attcttaggg gcagggcatg ctcatgtaga
     2041 gcgcctgctc gccgtccgag gcggtgccgt cgtacagggc ggtgtccagg ccgcagaggg
     2101 tgaaccccat ccgccggtac gcgtggatcg ccggtgcgtt gacgttggtg acctccagcc
     2161 agaggtgccc ggcgccccgc tcgcgggcga actccgtcgc gagccccatc aacgcgcgcc
     2221 cgaccccgtg cccccggtgc tccggggcga cctcgatgtc ctcgacggtc agccggcggt
     2281 tccagccgga gtacgagacg accacgaagc ccgccaggtc gccgtcgtcc ccgtacgcga
     2341 cgaacgtccg ggagtccggg tcgccgtcct ccccgtcgtc cgattcgtcg tccgattcgt
     2401 cgtcggggaa caccttggtc aggggcgggt ccaccggcac ctcccgcagg gtgaagccgt
     2461 ccccggtggc ggtgacgcgg aagacggtgt cggtggtgaa ggacccatcc agtgcctcga
     2521 tggcctcggc gtcccccggg acactggtgc ggtaccggta agccgtgtcg tcaagagtgg
     2581 cggccgccat agacatgttg ggcgagttta ctaatggaaa aagacaatta gcaaagaggc
     2641 tgcatattag gagacgcggt gaagattggg gaggacaata gggtccgtgc accaaaaagt
     2701 cctgacggca gctctgttca acgcacatcc atctcttcca gctatgcgtg tggcaccagc
     2761 acggatggat tgttgcacca aaaggacaag gcgagatgga tgcaggtcga ggcgatgcgg
     2821 tgctatcggc cctcgttgtc gaacgccacg tcgtcctctc ggcgaccgtc aatcctggaa
     2881 ggaaatgacc aagcgtgcac ccagcatcca gcatccaaca tccagcaccc agcacctagc
     2941 acccatgatc caatggccgg catgcagatc agcgtgggga gagcaccatc tagcagcccc
     3001 tggcaggccc agcattgtgc tcggatgtgc gataggcggt gggtgcgata aggtggatta
     3061 gtggaggatg gaggactgag tggcgggcaa gggaaggggg cgggaagatg actcaccttg
     3121 ggcagtgggt gggggatgcg gtatagggga tgtatgtgga tgctggcgga ggatagaagc
     3181 tgtaagctga gctggtgcca acggtgacgc tgtgagagtg gtctcggcga ccctgacggc
     3241 gactcggcgt taccccgtgt ggcacaccgg acgcgcgtcg agtgacgttg ctgctattgt
     3301 tgttaccatc atcctctcct ccgacaacca tacactcagc gcacaccatg ccactcgaat
     3361 cctgcatgca agggcgaatt ctgcagatat ccatcacact ggcggccgct cgagcatgca
     3421 tctagagggc ccaattcgcc ctatagtgaa tcgtattaca attcactggc cgtcgtttta
     3481 caacgtcgtg actgggaaaa ccctggcgtt acccaactta atcgccttgc agcacatccc
     3541 cctttcgcca gctggcgtaa tagcgaagag gcccgcaccg atcgcccttc ccaacagttg
     3601 cgcagcctat acgtacggca gtttaaggtt tacacctata aaagagagag ccgttatcgt
     3661 ctgtttgtgg atgtacagag tgatattatt gacacgccgg ggcgacggat ggtgatcccc
     3721 ctggccagtg cacgtctgct gtcagataaa gtctcccgtg aactttaccc ggtggtgcat
     3781 atcggggatg aaagctggcg catgatgacc accgatatgg ccagtgtgcc ggtctccgtt
     3841 atcggggaag aagtggctga tctcagccac cgcgaaaatg acatcaaaaa cgccattaac
     3901 ctgatgttct ggggaatata aatgtcaggc atgagattat caaaaaggat cttcacctag
     3961 atccttttca cgtagaaagc cagtccgcag aaacggtgct gaccccggat gaatgtcagc
     4021 tactgggcta tctggacaag ggaaaacgca agcgcaaaga gaaagcaggt agcttgcagt
     4081 gggcttacat ggcgatagct agactgggcg gttttatgga cagcaagcga accggaattg
     4141 ccagctgggg cgccctctgg taaggttggg aagccctgca aagtaaactg gatggctttc
     4201 ttgccgccaa ggatctgatg gcgcagggga tcaagctctg atcaagagac aggatgagga
     4261 tcgtttcgca tgattgaaca agatggattg cacgcaggtt ctccggccgc ttgggtggag
     4321 aggctattcg gctatgactg ggcacaacag acaatcggct gctctgatgc cgccgtgttc
     4381 cggctgtcag cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc cggtgccctg
     4441 aatgaactgc aagacgaggc agcgcggcta tcgtggctgg ccacgacggg cgttccttgc
     4501 gcagctgtgc tcgacgttgt cactgaagcg ggaagggact ggctgctatt gggcgaagtg
     4561 ccggggcagg atctcctgtc atctcacctt gctcctgccg agaaagtatc catcatggct
     4621 gatgcaatgc ggcggctgca tacgcttgat ccggctacct gcccattcga ccaccaagcg
     4681 aaacatcgca tcgagcgagc acgtactcgg atggaagccg gtcttgtcga tcaggatgat
     4741 ctggacgaag agcatcaggg gctcgcgcca gccgaactgt tcgccaggct caaggcgagc
     4801 atgcccgacg gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg
     4861 gtggaaaatg gccgcttttc tggattcatc gactgtggcc ggctgggtgt ggcggaccgc
     4921 tatcaggaca tagcgttggc tacccgtgat attgctgaag agcttggcgg cgaatgggct
     4981 gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat cgccttctat
     5041 cgccttcttg acgagttctt ctgaattgaa aaaggaagag tatgagtatt caacatttcc
     5101 gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct cacccagaaa
     5161 cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt tacatcgaac
     5221 tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt tttccaatga
     5281 tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtattgac gccgggcaag
     5341 agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac tcaccagtca
     5401 cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct gccataacca
     5461 tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg aaggagctaa
     5521 ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg gaaccggagc
     5581 tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgtagca atggcaacaa
     5641 cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa caattaatag
     5701 actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt ccggctggct
     5761 ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc attgcagcac
     5821 tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg agccaggcaa
     5881 ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt
     5941 aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt catttttaat
     6001 ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg
     6061 agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc
     6121 ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg
     6181 tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag
     6241 cgcagatacc aaatactgtt cttctagtgt agccgtagtt aggccaccac ttcaagaact
     6301 ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg
     6361 gcgataagtc gtgtcttacc gggttggatt caagacgata gttaccggat aaggcgcagc
     6421 ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg
     6481 aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg
     6541 cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag
     6601 ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc
     6661 gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct
     6721 ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc
     6781 ctgattctgt ggataaccgt attaccgcct ttgagtgagc tgataccgct cgccgcagcc
     6841 gaacgaccga gcgcagcgaa tcagtgagcg aggaagcgga ag
//
