LOCUS       sequence-449669-        5965 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  phospho-null mutant of AMPK-SPARK.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5965)
  AUTHORS   Erdogan YC, Pilic J, Gottschalk B, Yigit EN, Zaki AG, Ozturk G, 
            Eroglu E, Okutan B, Sommer NG, Weinberg AM, Schindl R, Graier WF, 
            Malli R
  TITLE     Development of a Dual Reporter System to Simultaneously Visualize 
            Ca(2+) Signals and AMPK Activity.
  JOURNAL   ACS Sens. 2024 Aug 21. doi: 10.1021/acssensors.4c01058.
  PUBMED    39167044
REFERENCE   2  (bases 1 to 5965)
  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: "ACS Sens. 
            2024 Aug 21. doi: 10.1021/acssensors.4c01058."
COMMENT     Sequence Label: AMPK(T>A)-SPARK
FEATURES             Location/Qualifiers
     source          1..5965
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      85..131
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      161..297
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    342..413
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(639..1227)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(719..738)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             join(1973..2041,2042..2833)
                     /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(2191..2210)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      2846..2982
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    3134..3205
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(3210..3256)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    3274..3322
                     /note="synthetic polyadenylation signal"
     enhancer        3403..3706
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3708..3910
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     3861..3881
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      4007..4016
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             5069..5122
                     /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"
     polyA_signal    5814..5935
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5825..5844)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5879..5898
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
ORIGIN
        1 caggttcagg gggaggtgtg ggaggttttt taaagcaagt aaaacctcta caaatgtggt
       61 attggcccat ctctatcggt atcgtagcat aaccccttgg ggcctctaaa cgggtcttga
      121 ggggtttttt gtgcccctcg ggccggattg ctatctaccg gcattggcgc agaaaaaaat
      181 gcctgatgcg acgctgcgcg tcttatactc ccacatatgc cagattcagc aacggatacg
      241 gcttccccaa cttgcccact tccatacgtg tcctccttac cagaaattta tccttaaggt
      301 cgtcagctat cctgcaggcg atctctcgat ttcgatcaag acattccttt aatggtcttt
      361 tctggacacc actaggggtc agaagtagtt catcaaactt tcttccctcc ctaatctcat
      421 tggttacctt gggctatcga aacttaatta accagtcaag tcagctactt ggcgagatcg
      481 acttgtctgg gtttcgacta cgctcagaat tgcgtcagtc aagttcgatc tggtccttgc
      541 tattgcaccc gttctccgat tacgagtttc atttaaatca tgtgagcaaa aggccagcaa
      601 aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct
      661 gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa
      721 agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg
      781 cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca
      841 cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa
      901 ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg
      961 gtaagacacg acttatcgcc actggcagca gccactggta acaggattag cagagcgagg
     1021 tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga
     1081 acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc
     1141 tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag
     1201 attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac
     1261 gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc
     1321 ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag
     1381 taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt
     1441 ctatttcgtt catccatagt tgcatttaaa tttccgaact ctccaaggcc ctcgtcggaa
     1501 aatcttcaaa cctttcgtcc gatccatctt gcaggctacc tctcgaacga actatcgcaa
     1561 gtctcttggc cggccttgcg ccttggctat tgcttggcag cgcctatcgc caggtattac
     1621 tccaatcccg aatatccgag atcgggatca cccgagagaa gttcaaccta catcctcaat
     1681 cccgatctat ccgagatccg aggaatatcg aaatcggggc gcgcctggtg taccgagaac
     1741 gatcctctca gtgcgagtct cgacgatcca tatcgttgct tggcagtcag ccagtcggaa
     1801 tccagcttgg gacccaggaa gtccaatcgt cagatattgt actcaagcct ggtcacggca
     1861 gcgtaccgat ctgtttaaac ctagatattg atagtctgat cggtcaacgt ataatcgagt
     1921 cctagctttt gcaaacatct atcaagagac aggatcagca ggaggctttc gcatgagtat
     1981 tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc
     2041 tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg cgcgagtggg
     2101 ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg
     2161 ctttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga
     2221 cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta
     2281 ttcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc
     2341 tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga ttggaggacc
     2401 gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg
     2461 ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc
     2521 aatggcaaca accttgcgta aactattaac tggcgaacta cttactctag cttcccggca
     2581 acagttgata gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct
     2641 tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat
     2701 cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg
     2761 gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat
     2821 taagcattgg taaccgattc taggtgcatt ggcgcagaaa aaaatgcctg atgcgacgct
     2881 gcgcgtctta tactcccaca tatgccagat tcagcaacgg atacggcttc cccaacttgc
     2941 ccacttccat acgtgtcctc cttaccagaa atttatcctt aagatcccga atcgtttaaa
     3001 ctcgactctg gctctatcga atctccgtcg tttcgagctt acgcgaacag ccgtggcgct
     3061 catttgctcg tcgggcatcg aatctcgtca gctatcgtca gcttaccttt ttggcagcga
     3121 tcgcggctcc cgacatcttg gaccattagc tccacaggta tcttcttccc tctagtggtc
     3181 ataacagcag cttcagctac ctctcaattc aaaaaacccc tcaagacccg tttagaggcc
     3241 ccaaggggtt atgctatcaa tcgttgcgtt acacacacaa aaaaccaaca cacatccatc
     3301 ttcgatggat agcgatttta ttatctaact gctgatcgag tgtagccaga tctagtaatc
     3361 aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt
     3421 aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta
     3481 tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg
     3541 gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga
     3601 cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt
     3661 tcctacttgg cagtacatct acgtattagt catcgctatt accatgctga tgcggttttg
     3721 gcagtacatc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa gtctccaccc
     3781 cattgacgtc aatgggagtt tgttttggca ccaaaatcaa cgggactttc caaaatgtcg
     3841 taacaactcc gccccattga cgcaaatggg cggtaggcgt gtacggtggg aggtctatat
     3901 aagcagagct ggtttagtga accgtcagat cagatctttg tcgatcctac catccactcg
     3961 acacacccgc cagcggccgc tgccaagctt ccgagctctc gaattcgcca ccatgggagg
     4021 agggggctca ggcagcggag aagggagcac aaagatgcgg agggtggccg ctctcgtgga
     4081 tttgggaacc ggcgggtctg agctcgacgg agggctggag tccggcttgc gctctgggtc
     4141 tggtggtggc gatattatgg tgtccaaggg tgaggagctg ttcacaggcg tcgtgccaat
     4201 cctggtggag ctggacgggg acgtgaatgg ccacaaattc tcagtgagtg gtgaggggga
     4261 aggggacgct acatatggga agctgaccct gaagttcatc tgcacaactg gaaaacttcc
     4321 tgtcccgtgg cccacactgg tgaccacact cacctacggc gtgcaatgtt tttccagata
     4381 tcccgaccac atgaaacagc atgacttttt taaatcagct atgcccgaag ggtatgtgca
     4441 ggagcgcact atcttcttta aagacgacgg taattacaaa acacgagccg aagtgaagtt
     4501 cgagggagat actctggtta acaggattga actcaagggt atcgatttca aagaagatgg
     4561 gaacatattg gggcataagc ttgagtataa ttacaattct cataacgtct acataatggc
     4621 cgacaaacaa aagaacggca tcaaggcaaa ctttaaaatt cggcacaaca ttgaagatgg
     4681 gtctgtacag ctggccgatc attaccaaca gaataccccc atcggcgacg gtcctgttct
     4741 gcttccggac aaccactact tgagcaccca gagtgcattg tctaaggatc caaacgaaaa
     4801 aagggaccac atggtgctgc tggagtttgt aacggcggcc gggatcaccc tgggcatgga
     4861 tgagctgtac aagggcagcg gtagcgcggg cggatccgct gggggaagtg ccggtggaag
     4921 tgcaggtgga agcgctggag gctctgcagg aggctccgcc ggaggatcca ggggcgagat
     4981 cgctaaatcc ctgaaggaaa ttgctaaaag tctgaaagaa atagcctgga gtctcaagga
     5041 aatcgccaaa tcactgaagg gcggaacgga agggagggga tcccttctta catgtggcga
     5101 cgtcgaagag aatccaggcc caaaactggc gaccatgcac aagttcagtc aggagcagat
     5161 tggcgagaac atcgtctgcc gcgttatttg caccaccgga cagatcccta ttagagacct
     5221 ttccgccgac atctcacagg tcttgaaaga gaaacggtct ataaagaagg tttggacttt
     5281 tggacgaaac cctgcatgcg attatcatct gggcaacatt agcagactct caaacaagca
     5341 cttccagata ctcctgggcg aggatggcaa tcttctgctg aatgacatta gcactaacgg
     5401 gacttggctc aatggccaaa aggtggagaa gaacagtaat cagctgctca gccagggcga
     5461 tgagattacg gtgggagttg gagtagagtc cgatattctc agcctggtta tctttattaa
     5521 tgataagttc aagcagtgtc tggagcaaaa taaggtcgat cgaggctcag gctctgccgg
     5581 gggatctgct ggagggagcg ctgggggcag tgctggtggg tctgcaggcg gttccgcagg
     5641 tggttccgcc gggggcagca gaaccctgcg ggagatcgaa gaacttttgc gcaagatcat
     5701 cgaggacagc gtaagatcag tggccgaact cgaagacatc gagaaatggc tgaaaaagat
     5761 atgagctagc ttgactgact gagatacagc gtaccttcag ctcacagaca tgataagata
     5821 cattgatgag tttggacaaa ccacaactag aatgcagtga aaaaaatgct ttatttgtga
     5881 aatttgtgat gctattgctt tatttgtaac cattataagc tgcaataaac aagttaacaa
     5941 caacaattgc attcatttta tgttt
//
