LOCUS       sequence-367998-        6450 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  NMA111-GFP with first nuclear localization signal mutated (amino 
            acids 9 - 11, KKR -> AAA).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6450)
  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 6450)
  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: KBB428
FEATURES             Location/Qualifiers
     source          1..6450
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    89..110
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(125..130,131..148,149..155)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    163..179
                     /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     168..190
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     187..203
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     187..203
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        224..242
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     270..286
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     271..287
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             908..1618
                     /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(935..963)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     1558..1578
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     primer_bind     complement(1669..1687)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1683..1730
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     complement(1937..1956)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1938..1956)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1963..1980)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1963..1979)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1972..1994)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2120..2575
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2207..2226)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2417..2438
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2706..3509)
                     /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(3510..3725)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(3749..3768)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3868..3890
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3928..3946)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_feature    3987..4490
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        4527..4631
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4632..4700,4701..5492)
                     /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(4850..4869)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5663..6251
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6152..6171
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6405..6422
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg gcacgacagg tttcccgact
       61 ggaaagcggg cagtgagcgc aacgcaatta atgtgagtta gctcactcat taggcacccc
      121 aggctttaca ctttatgctt ccggctcgta tgttgtgtgg aattgtgagc ggataacaat
      181 ttcacacagg aaacagctat gaccatgatt acgccaagct cgaaattaac cctcactaaa
      241 gggaacaaaa gctggtaccg ggccccccct cgaggtcgac ggtatcgata agcttgatat
      301 cgaattcctg caggcatcag catcagcaag atccaagaac ctagagtcac gattgaaata
      361 tttaataata aatgctatgt atcatagata aactagaaag aggcaagaaa aatttaacaa
      421 gtttcttaat tgaatattat tttactgtcg ctaccattat tagacgccat ctcagcgtgt
      481 gacatgataa tttttctttt tttctttttt tttcctcaag gcgatgatat gaaactttca
      541 aaaaatttat atgtaaataa aaaagttcta cgagggttta aatacggaag tttatttaga
      601 agatatttat actctgaata cacacgtaga gtacagtaaa ggttttttag atctactaat
      661 gaccatatcg ttgagcaata tagctgctgc tgaccattct aaaatttccg atggaacttc
      721 tggtgaatca tcgcttgtaa aaaggaaaca gttggaaagc gccaccggag atcaagagga
      781 agaatataca gaccatgaaa tcattataga gccgttgcac tttgctaata ataataacac
      841 tgtactcaca gattcagaaa attatctaag atggcaaaat accatctcca acgttgttaa
      901 atcagtgagc aaaggagaag aactcttcac tggagttgtc ccaattcttg ttgaattaga
      961 tggtgatgtt aatgggcaca aattttctgt cagtggagag ggtgaaggtg atgcaacata
     1021 cggaaaactt acccttaaat ttatttgcac tactggaaaa ctacctgttc catggccaac
     1081 acttgtcact actttcactt atggtgttca gtgcttttca agatacccgg atcatatgaa
     1141 acggcatgac tttttcaaga gtgccatgcc cgaaggttat gtacaggaaa gaactatatt
     1201 tttcaaagat gacgggaact acaagacacg tgctgaagtc aagtttgaag gtgataccct
     1261 tgttaataga atcgagttaa aaggtattga ttttaaagaa gatggaaaca ttcttgggca
     1321 caaattggaa tacaactata actcacacaa tgtatacatc atggcagaca aacaaaagaa
     1381 tggaatcaaa gctaacttca aaattagaca caacattgaa gatggaagcg ttcaactagc
     1441 agaccattat caacaaaata ctccaattgg cgatggccct gtccttttac cagacaacca
     1501 ttacctgtcc acacaatctg ccctttcgaa agatcccaac gaaaagagag accacatggt
     1561 ccttcttgag tttgtaacag ctgctgggat tacacatggc atggatgaac tatacaaagg
     1621 atccggctgc taacaaagcc cgaaaggaag ctgagttggc tgctgccacc gctgagcaat
     1681 aactagcata accccttggg gcctctaaac gggtcttgag gggttttttg ctgaaaggag
     1741 gaactatatc cggatatcca caggacgggt gtggtcgcca tgatcgcgta gtcgatagtg
     1801 gctccaagta gcgaagcgag caggactggg cggcggccaa agcggtcgga cagtgctccg
     1861 agaacgggtg cgcatagaaa ttgcatcaac gcatatagcg ctagagcggc cgccaccgcg
     1921 gtggagctcc aattcgccct atagtgagtc gtattacaat tcactggccg tcgttttaca
     1981 acgtcgtgac tgggaaaacc ctggcgttac ccaacttaat cgccttgcag cacatccccc
     2041 tttcgccagc tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg
     2101 cagcctgaat ggcgaatgga cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg
     2161 gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc
     2221 ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc
     2281 cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt
     2341 gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag
     2401 tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg
     2461 gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag
     2521 ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat taacgcttac aatttcctga
     2581 tgcggtattt tctccttacg catctgtgcg gtatttcaca ccgcataggg taataactga
     2641 tataattaaa ttgaagctct aatttgtgag tttagtatac atgcatttac ttataataca
     2701 gttttttagt tttgctggcc gcatcttctc aaatatgctt cccagcctgc ttttctgtaa
     2761 cgttcaccct ctaccttagc atcccttccc tttgcaaata gtcctcttcc aacaataata
     2821 atgtcagatc ctgtagagac cacatcatcc acggttctat actgttgacc caatgcgtct
     2881 cccttgtcat ctaaacccac accgggtgtc ataatcaacc aatcgtaacc ttcatctctt
     2941 ccacccatgt ctctttgagc aataaagccg ataacaaaat ctttgtcgct cttcgcaatg
     3001 tcaacagtac ccttagtata ttctccagta gatagggagc ccttgcatga caattctgct
     3061 aacatcaaaa ggcctctagg ttcctttgtt acttcttctg ccgcctgctt caaaccgcta
     3121 acaatacctg ggcccaccac accgtgtgca ttcgtaatgt ctgcccattc tgctattctg
     3181 tatacacccg cagagtactg caatttgact gtattaccaa tgtcagcaaa ttttctgtct
     3241 tcgaagagta aaaaattgta cttggcggat aatgccttta gcggcttaac tgtgccctcc
     3301 atggaaaaat cagtcaagat atccacatgt gtttttagta aacaaatttt gggacctaat
     3361 gcttcaacta actccagtaa ttccttggtg gtacgaacat ccaatgaagc acacaagttt
     3421 gtttgctttt cgtgcatgat attaaatagc ttggcagcaa caggactagg atgagtagca
     3481 gcacgttcct tatatgtagc tttcgacatg atttatcttc gtttcctgca ggtttttgtt
     3541 ctgtgcagtt gggttaagaa tactgggcaa tttcatgttt cttcaacact acatatgcgt
     3601 atatatacca atctaagtct gtgctccttc cttcgttctt ccttctgttc ggagattacc
     3661 gaatcaaaaa aatttcaagg aaaccgaaat caaaaaaaag aataaaaaaa aaatgatgaa
     3721 ttgaaaaggt ggtatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc
     3781 agccccgaca cccgccaaca cccgctgacg cgccctgacg ggcttgtctg ctcccggcat
     3841 ccgcttacag acaagctgtg accgtctccg ggagctgcat gtgtcagagg ttttcaccgt
     3901 catcaccgaa acgcgcgaga cgaaagggcc tcgtgatacg cctattttta taggttaatg
     3961 tcatgataat aatggtttct tagacggatc gcttgcctgt aacttacacg cgcctcgtat
     4021 cttttaatga tggaataatt tgggaattta ctctgtgttt atttattttt atgttttgta
     4081 tttggatttt agaaagtaaa taaagaaggt agaagagtta cggaatgaag aaaaaaaaat
     4141 aaacaaaggt ttaaaaaatt tcaacaaaaa gcgtacttta catatatatt tattagacaa
     4201 gaaaagcaga ttaaatagat atacattcga ttaacgataa gtaaaatgta aaatcacagg
     4261 attttcgtgt gtggtcttct acacagacaa gatgaaacaa ttcggcatta atacctgaga
     4321 gcaggaagag caagataaaa ggtagtattt gttggcgatc cccctagagt cttttacatc
     4381 ttcggaaaac aaaaactatt ttttctttaa tttctttttt tactttctat ttttaattta
     4441 tatatttata ttaaaaaatt taaattataa ttatttttat agcacgtgat gaaaaggacc
     4501 caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
     4561 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa
     4621 aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat
     4681 tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc
     4741 agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga
     4801 gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg
     4861 cggtattatc ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc
     4921 agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag
     4981 taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc
     5041 tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg
     5101 taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg
     5161 acaccacgat gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac
     5221 ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac
     5281 cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg
     5341 agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg
     5401 tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg
     5461 agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac
     5521 tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg
     5581 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg
     5641 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc
     5701 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc
     5761 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt
     5821 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
     5881 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
     5941 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
     6001 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag
     6061 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg
     6121 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
     6181 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
     6241 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt
     6301 ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct
     6361 ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg
     6421 aggaagcgga agagcgccca atacgcaaac
//
