LOCUS       sequence-368167-        7120 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  NMA111-GFP with first nuclear localization signal mutated (amino 
            acids 28 - 30, KRK -> AAA).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7120)
  AUTHORS   Belanger KD, Walter D, Henderson TA, Yelton AL, O'Brien TG, Belanger
            KG, Geier SJ, Fahrenkrog B
  TITLE     Nuclear localisation is crucial for the proapoptotic activity of the
            HtrA-like serine protease Nma111p.
  JOURNAL   J Cell Sci. 2009 Nov 1;122(Pt 21):3931-41. doi: 10.1242/jcs.056887. 
            Epub 2009 Oct 6.
  PUBMED    19808885
REFERENCE   2  (bases 1 to 7120)
  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: "J Cell 
            Sci."; date: "2009-11-1"; volume: "122(Pt 21):3931-41. doi"; pages: 
            " 10.1242/jcs.056887. Epub 2009 Oct 6"
COMMENT     Sequence Label: KBB430
FEATURES             Location/Qualifiers
     source          1..7120
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    1..504
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        541..645
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(646..714,715..1506)
                     /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(864..883)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1677..2265
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2166..2185
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2419..2436
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    2553..2574
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(2589..2594,2595..2612,2613..2619)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2627..2643
                     /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     2632..2654
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2651..2667
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2651..2667
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             3361..3534
                     /codon_start=1
                     /product="IgG-binding unit of Staphylococcus aureus protein
                     A"
                     /label=ProtA
                     /translation="VDNKFNKEQQNAFYEILHLPNLNEEQRNAFIQSLKDDPSQSANLL
                     AEAKKLNDAQAPK"
     CDS             3535..3708
                     /codon_start=1
                     /product="IgG-binding unit of Staphylococcus aureus protein
                     A"
                     /label=ProtA
                     /translation="VDNKFNKEQQNAFYEILHLPNLNEEQRNAVIQSLKDDPSQSANLL
                     AEAKKLNDAQAPK"
     CDS             3754..3774
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             4042..4752
                     /codon_start=1
                     /product="S65T variant of Aequorea victoria green 
                     fluorescent protein (Heim et al., 1995)"
                     /label=GFP (S65T)
                     /note="excitable with blue light"
                     /translation="SKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKF
                     ICTTGKLPVPWPTLVTTFTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTIFFKDDGN
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKAN
                     FKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLEF
                     VTAAGITHGMDELYK"
     primer_bind     complement(4069..4097)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     4692..4712
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     primer_bind     complement(4803..4821)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      4817..4864
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     complement(5071..5090)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(5072..5090)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5097..5114)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5097..5113)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5106..5128)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5254..5709
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5341..5360)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5551..5572
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(5840..6643)
                     /codon_start=1
                     /gene="S. cerevisiae URA3"
                     /product="orotidine-5'-phosphate decarboxylase, required 
                     for uracil biosynthesis"
                     /label=URA3
                     /note="yeast auxotrophic marker, counterselectable with 
                     5-fluoroorotic acid (5-FOA)"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
                     ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     promoter        complement(6644..6859)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(6883..6902)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7002..7024
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(7062..7080)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 gatcgcttgc ctgtaactta cacgcgcctc gtatctttta atgatggaat aatttgggaa
       61 tttactctgt gtttatttat ttttatgttt tgtatttgga ttttagaaag taaataaaga
      121 aggtagaaga gttacggaat gaagaaaaaa aaataaacaa aggtttaaaa aatttcaaca
      181 aaaagcgtac tttacatata tatttattag acaagaaaag cagattaaat agatatacat
      241 tcgattaacg ataagtaaaa tgtaaaatca caggattttc gtgtgtggtc ttctacacag
      301 acaagatgaa acaattcggc attaatacct gagagcagga agagcaagat aaaaggtagt
      361 atttgttggc gatcccccta gagtctttta catcttcgga aaacaaaaac tattttttct
      421 ttaatttctt tttttacttt ctatttttaa tttatatatt tatattaaaa aatttaaatt
      481 ataattattt ttatagcacg tgatgaaaag gacccaggtg gcacttttcg gggaaatgtg
      541 cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga
      601 caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat
      661 ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca
      721 gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc
      781 gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca
      841 atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg
      901 caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca
      961 gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata
     1021 accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag
     1081 ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg
     1141 gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca
     1201 acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta
     1261 atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct
     1321 ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca
     1381 gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag
     1441 gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat
     1501 tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt
     1561 taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa
     1621 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
     1681 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
     1741 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
     1801 agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag
     1861 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
     1921 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
     1981 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
     2041 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
     2101 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
     2161 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     2221 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
     2281 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta
     2341 tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc
     2401 agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cccaatacgc
     2461 aaaccgcctc tccccgcgcg ttggccgatt cattaatgca gctggcacga caggtttccc
     2521 gactggaaag cgggcagtga gcgcaacgca attaatgtga gttagctcac tcattaggca
     2581 ccccaggctt tacactttat gcttccggct cgtatgttgt gtggaattgt gagcggataa
     2641 caatttcaca caggaaacag ctatgacatg attacgaatt cgatccattg agtcatcagc
     2701 ctcttcttga aaaaacgatt ctgaaataac cggaacttgt ttagattcct ttttgtaggt
     2761 taatcttatc ctgtacttgc tgatattgtg gttattagca gagatttttt tcaaaacatc
     2821 atctaaagta ggctttttgg ataagtcaat ttcagtgtcc cttaaccctt tagagcggct
     2881 ttttatggtg ataggcattt tcaaattaaa ttcaaaatat gtatctctct caaatagctc
     2941 aattggtttc tagatagcta aatagaatat aatcttactg tcctccgttc tgtaaaattc
     3001 acgctcttag tcccttttca taattcctta actttttgcg tacaaaatga tatgtttatt
     3061 atatttttct tttttttttt tcaaattttt tctttttctt gaaaaatttt tcaaattgga
     3121 aagctcatct ctcttgaatg tataatactt tcttcctcta actttcaaaa agttttacat
     3181 agccaagaag ttttccttac atcggtatac tactgttata taagttattc ttcgagaaac
     3241 aattagatat cattcatcgg ataaatctaa gttgcccatt gctttcaata actccgatca
     3301 aattaactca aatcaactaa aacagtaatg tcagcatgcc ttgcgcaaca cgatgaagcc
     3361 gtggacaaca aattcaacaa agaacaacaa aacgcgttct atgagatctt acatttacct
     3421 aacttaaacg aagaacaacg aaacgccttc atccaaagtt taaaagatga cccaagccaa
     3481 agcgctaacc ttttagcaga agctaaaaag ctaaatgatg ctcaggcgcc gaaagtagac
     3541 aacaaattca acaaagaaca acaaaacgcg ttctatgaga tcttacattt acctaactta
     3601 aacgaagaac aacgaaacgc cgtcatccaa agtttaaaag atgacccaag ccaaagcgct
     3661 aaccttttag cagaagctaa aaagctaaat gatgctcagg cgccgaaagt agacgcgaat
     3721 tcggggctgc aggaattcga tatcccaacg accgaaaacc tgtattttca gggcgcccat
     3781 atggccatgg caatgaccat atcgttgagc aatataaaga aaagagacca ttctaaaatt
     3841 tccgatggaa cttctggtga atcatcgctt gtagctgctg ctcagttgga aagcgccacc
     3901 ggagatcaag aggaagaata tacagaccat gaaatcatta tagagccgtt gcactttgct
     3961 aataataata acactgtact cacagattca gaaaattatc taagatggca aaataccatc
     4021 tccaacgttg ttaaatcagt gagcaaagga gaagaactct tcactggagt tgtcccaatt
     4081 cttgttgaat tagatggtga tgttaatggg cacaaatttt ctgtcagtgg agagggtgaa
     4141 ggtgatgcaa catacggaaa acttaccctt aaatttattt gcactactgg aaaactacct
     4201 gttccatggc caacacttgt cactactttc acttatggtg ttcagtgctt ttcaagatac
     4261 ccggatcata tgaaacggca tgactttttc aagagtgcca tgcccgaagg ttatgtacag
     4321 gaaagaacta tatttttcaa agatgacggg aactacaaga cacgtgctga agtcaagttt
     4381 gaaggtgata cccttgttaa tagaatcgag ttaaaaggta ttgattttaa agaagatgga
     4441 aacattcttg ggcacaaatt ggaatacaac tataactcac acaatgtata catcatggca
     4501 gacaaacaaa agaatggaat caaagctaac ttcaaaatta gacacaacat tgaagatgga
     4561 agcgttcaac tagcagacca ttatcaacaa aatactccaa ttggcgatgg ccctgtcctt
     4621 ttaccagaca accattacct gtccacacaa tctgcccttt cgaaagatcc caacgaaaag
     4681 agagaccaca tggtccttct tgagtttgta acagctgctg ggattacaca tggcatggat
     4741 gaactataca aaggatccgg ctgctaacaa agcccgaaag gaagctgagt tggctgctgc
     4801 caccgctgag caataactag cataacccct tggggcctct aaacgggtct tgaggggttt
     4861 tttgctgaaa ggaggaacta tatccggata tccacaggac gggtgtggtc gccatgatcg
     4921 cgtagtcgat agtggctcca agtagcgaag cgagcaggac tgggcggcgg ccaaagcggt
     4981 cggacagtgc tccgagaacg ggtgcgcata gaaattgcat caacgcatat agcgctagag
     5041 cggccgccac cgcggtggag ctccaattcg ccctatagtg agtcgtatta caattcactg
     5101 gccgtcgttt tacaacgtcg tgactgggaa aaccctggcg ttacccaact taatcgcctt
     5161 gcagcacatc cccctttcgc cagctggcgt aatagcgaag aggcccgcac cgatcgccct
     5221 tcccaacagt tgcgcagcct gaatggcgaa tggacgcgcc ctgtagcggc gcattaagcg
     5281 cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg
     5341 ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc
     5401 taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa
     5461 aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc
     5521 ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac
     5581 tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt
     5641 ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa atattaacgc
     5701 ttacaatttc ctgatgcggt attttctcct tacgcatctg tgcggtattt cacaccgcat
     5761 agggtaataa ctgatataat taaattgaag ctctaatttg tgagtttagt atacatgcat
     5821 ttacttataa tacagttttt tagttttgct ggccgcatct tctcaaatat gcttcccagc
     5881 ctgcttttct gtaacgttca ccctctacct tagcatccct tccctttgca aatagtcctc
     5941 ttccaacaat aataatgtca gatcctgtag agaccacatc atccacggtt ctatactgtt
     6001 gacccaatgc gtctcccttg tcatctaaac ccacaccggg tgtcataatc aaccaatcgt
     6061 aaccttcatc tcttccaccc atgtctcttt gagcaataaa gccgataaca aaatctttgt
     6121 cgctcttcgc aatgtcaaca gtacccttag tatattctcc agtagatagg gagcccttgc
     6181 atgacaattc tgctaacatc aaaaggcctc taggttcctt tgttacttct tctgccgcct
     6241 gcttcaaacc gctaacaata cctgggccca ccacaccgtg tgcattcgta atgtctgccc
     6301 attctgctat tctgtataca cccgcagagt actgcaattt gactgtatta ccaatgtcag
     6361 caaattttct gtcttcgaag agtaaaaaat tgtacttggc ggataatgcc tttagcggct
     6421 taactgtgcc ctccatggaa aaatcagtca agatatccac atgtgttttt agtaaacaaa
     6481 ttttgggacc taatgcttca actaactcca gtaattcctt ggtggtacga acatccaatg
     6541 aagcacacaa gtttgtttgc ttttcgtgca tgatattaaa tagcttggca gcaacaggac
     6601 taggatgagt agcagcacgt tccttatatg tagctttcga catgatttat cttcgtttcc
     6661 tgcaggtttt tgttctgtgc agttgggtta agaatactgg gcaatttcat gtttcttcaa
     6721 cactacatat gcgtatatat accaatctaa gtctgtgctc cttccttcgt tcttccttct
     6781 gttcggagat taccgaatca aaaaaatttc aaggaaaccg aaatcaaaaa aaagaataaa
     6841 aaaaaaatga tgaattgaaa aggtggtatg gtgcactctc agtacaatct gctctgatgc
     6901 cgcatagtta agccagcccc gacacccgcc aacacccgct gacgcgccct gacgggcttg
     6961 tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct gcatgtgtca
     7021 gaggttttca ccgtcatcac cgaaacgcgc gagacgaaag ggcctcgtga tacgcctatt
     7081 tttataggtt aatgtcatga taataatggt ttcttagacg
//
