LOCUS       sequence-398227-        7855 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  express Thr to Ala mutant ATMSPARK2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7855)
  AUTHORS   Li X, Chung CI, Yang J, Chaudhuri S, Munster PN, Shu X
  TITLE     ATM-SPARK: A GFP phase separation-based activity reporter of ATM.
  JOURNAL   Sci Adv. 2023 Mar;9(9):eade3760. doi: 10.1126/sciadv.ade3760. Epub 
            2023 Mar 1.
  PUBMED    36857446
REFERENCE   2  (bases 1 to 7855)
  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: "Sci Adv."; 
            date: "2023-03"; volume: "9(9):eade3760. doi"; pages: " 
            10.1126/sciadv.ade3760. Epub 2023 Mar 1"
COMMENT     Sequence Label: pcDNA-mutATMSPARK2
FEATURES             Location/Qualifiers
     source          1..7855
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             991..1017
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             join(1018..1020,1021..1023,1024..1734)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1063..1084)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1324..1343)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1671..1692
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1930..1983
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             2467..2493
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     primer_bind     complement(3446..3463)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3452..3676
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3722..4150
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3809..3828)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4019..4040
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4159..4179)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4164..4493
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4344..4479
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4406..4425
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4560..5354
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4614..4633)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5224..5243
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5528..5649
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5565..5584)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5619..5638
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5698..5714)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5698..5714)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5711..5733)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5722..5738
                     /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(5746..5752,5753..5770,5771..5776))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5791..5812
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5929..5946)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6100..6688)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6180..6199)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6859..7650,7651..7719))
                     /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     7482..7501
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7720..7824)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gccaccatgt tggagacggt gagcgcgcag gagttgtata gcataccgct cgagtccgga
      961 ctcagatctg gcagcggtgg aggcgatatc cccgccgcca agcgggtgaa gctggacatg
     1021 gtgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtcga gctggacggc
     1081 gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc cacctacggc
     1141 aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc
     1201 gtgaccaccc tgacctacgg cgtgcagtgc ttcagccgct accccgacca catgaagcag
     1261 cacgacttct tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac catcttcttc
     1321 aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga caccctggtg
     1381 aaccgcatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct ggggcacaag
     1441 ctggagtaca actacaacag ccacaacgtc tatatcatgg ccgacaagca gaagaacggc
     1501 atcaaggcca acttcaagat ccgccacaac atcgaggacg gcagcgtgca gctcgccgac
     1561 cactaccagc agaacacccc catcggcgac ggccccgtgc tgctgcccga caaccactac
     1621 ctgagcaccc agtccgccct gagcaaagac cccaacgaga agcgcgatca catggtcctg
     1681 ctggagttcg tgaccgccgc cgggatcact ctcggcatgg acgagctgta caagggatcc
     1741 ggcagtgctg gtggtagtgc tggtggtagt gctggtggta gtgctggtgg cagtgctggt
     1801 ggtagtgctg gtggtagtgc tggtggctct agaggagaaa tcgcaaagag tctgaaagaa
     1861 attgcaaaat ccttgaagga aatagcgtgg tcactgaaag aaattgccaa gtccctcaaa
     1921 gggggtaccg agggcagagg aagtctgcta acatgcggtg acgtcgagga gaatcctggc
     1981 ccaaagcttg ccaccatgca taagttttct caagaacaga tcggcgaaaa cattgtgtgc
     2041 agggtcattt gtaccacggg tcaaattccc atccgagatt tgtcagctga tatttcacaa
     2101 gtgcttaagg aaaaacgatc cataaagaaa gtttggacat ttggtagaaa cccagcctgt
     2161 gactatcatt taggaaacat ttcaagactg tcaaataagc atttccaaat actactagga
     2221 gaagacggta accttttatt gaatgacatt tccactaatg ggacctggtt aaatgggcaa
     2281 aaagtcgaga agaacagcaa tcagttactg tctcaaggtg atgaaataac cgttggtgta
     2341 ggcgtggaat cagatatttt atctctggtc attttcataa acgacaaatt taagcagtgc
     2401 ctcgaacaga acaaagttga tcgcctcgag tctggtctgc gttctggttc tggtggtggt
     2461 atcgatccag cagctaaaag agtcaaattg gatatggttt ctaaaggtga agaactgttt
     2521 actggtgttg taccgatcct ggttgaactg gacggcgacg taaatggtca taaattctcc
     2581 gtttctggtg agggcgaggg tgatgcaact tatggcaagc tgaccctgaa attcatttgc
     2641 accaccggaa aactgccggt tccgtggccg accctggtga ccactctgac ctttggcgta
     2701 cagtgcttct ctcgctaccc ggatcacatg aaacagcacg actttttcaa atctgcgatg
     2761 ccggaaggtt acgtacaaga gcgtaccatc ttcttcaaag atgacggtaa ttacaaaaca
     2821 agagcggaag ttaaattcga gggtgacacc ctggtgaacc gcatcgaact gaaaggcatc
     2881 gacttcaaag aggatggcaa catcctgggc cacaaactgg aatacaacta caactcccac
     2941 aacgtgtaca tcatggctga taagcagaag aacggcatca aagcgaactt caaaattcgt
     3001 cacaacatcg aagacggctc tgttcaactg gcggatcact accagcagaa cacgccgatt
     3061 ggcgacggcc cggttctgct gccggataac cactatctgt ccacccagtc tgccctgtct
     3121 aaagacccga acgagaaacg tgaccatatg gtactgctgg aatttgttac cgccgctggt
     3181 atcaccctgg gtatggacga actgtacaag accggtggct ccgcaggtgg ttccgctggt
     3241 ggaagcgccg gtggttctgc gggtggatct gctggaggtt ctgcaggagg aagcgctgga
     3301 ggtgggcgcg ccaccttgag agaaatcgag gaattgttga ggaaaatcat tgaagacagc
     3361 gtgcgcagtg tggccgagct ggaggacatt gagaagtggc tgaagaaaat ctaagaattc
     3421 gggcccgttt aaacccgctg atcagcctcg actgtgcctt ctagttgcca gccatctgtt
     3481 gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtcctttcc
     3541 taataaaatg aggaaattgc atcgcattgt ctgagtaggt gtcattctat tctggggggt
     3601 ggggtggggc aggacagcaa gggggaggat tgggaagaca atagcaggca tgctggggat
     3661 gcggtgggct ctatggcttc tgaggcggaa agaaccagct ggggctctag ggggtatccc
     3721 cacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc
     3781 gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc
     3841 acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt
     3901 agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg
     3961 ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt
     4021 ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta
     4081 taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt
     4141 aacgcgaatt aattctgtgg aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc
     4201 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccagg tgtggaaagt
     4261 ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca
     4321 tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc gcccattctc
     4381 cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc tctgcctctg
     4441 agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaaaagctcc
     4501 cgggagcttg tatatccatt ttcggatctg atcaagagac aggatgagga tcgtttcgca
     4561 tgattgaaca agatggattg cacgcaggtt ctccggccgc ttgggtggag aggctattcg
     4621 gctatgactg ggcacaacag acaatcggct gctctgatgc cgccgtgttc cggctgtcag
     4681 cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc
     4741 aggacgaggc agcgcggcta tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc
     4801 tcgacgttgt cactgaagcg ggaagggact ggctgctatt gggcgaagtg ccggggcagg
     4861 atctcctgtc atctcacctt gctcctgccg agaaagtatc catcatggct gatgcaatgc
     4921 ggcggctgca tacgcttgat ccggctacct gcccattcga ccaccaagcg aaacatcgca
     4981 tcgagcgagc acgtactcgg atggaagccg gtcttgtcga tcaggatgat ctggacgaag
     5041 agcatcaggg gctcgcgcca gccgaactgt tcgccaggct caaggcgcgc atgcccgacg
     5101 gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg gtggaaaatg
     5161 gccgcttttc tggattcatc gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca
     5221 tagcgttggc tacccgtgat attgctgaag agcttggcgg cgaatgggct gaccgcttcc
     5281 tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat cgccttctat cgccttcttg
     5341 acgagttctt ctgagcggga ctctggggtt cgaaatgacc gaccaagcga cgcccaacct
     5401 gccatcacga gatttcgatt ccaccgccgc cttctatgaa aggttgggct tcggaatcgt
     5461 tttccgggac gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc
     5521 ccaccccaac ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa
     5581 tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa
     5641 tgtatcttat catgtctgta taccgtcgac ctctagctag agcttggcgt aatcatggtc
     5701 atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg
     5761 aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt
     5821 gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg
     5881 ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga
     5941 ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat
     6001 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca
     6061 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc
     6121 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata
     6181 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc
     6241 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc
     6301 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga
     6361 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc
     6421 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag
     6481 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag
     6541 aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag
     6601 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca
     6661 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga
     6721 cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat
     6781 cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga
     6841 gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg
     6901 tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga
     6961 gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc
     7021 agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac
     7081 tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc
     7141 agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc
     7201 gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc
     7261 catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt
     7321 ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc
     7381 atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg
     7441 tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag
     7501 cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat
     7561 cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc
     7621 atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa
     7681 aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta
     7741 ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa
     7801 aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtc
//
