LOCUS       sequence-456312-        8372 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express N-terminal mCherry fused to human diacylglycerol kinase 
            delta isoform 2 in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8372)
  TITLE     PHOSPHO2 is PA phosphatase and PHOSPHO1 is PLC
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 8372)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pAcGFP-C1-DGKd2
FEATURES             Location/Qualifiers
     source          1..8372
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      607..616
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             613..1329
                     /codon_start=1
                     /product="Aequorea coerulescens GFP"
                     /label=AcGFP1
                     /note="mammalian codon-optimized"
                     /translation="MVSKGAELFTGIVPILIELNGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLSYGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFEDD
                     GNYKSRAEVKFEGDTLVNRIELTGTDFKEDGNILGNKMEYNYNAHNVYIMTDKAKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMIYF
                     GFVTAAAITHGMDELYK"
     polyA_signal    5160..5281
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5197..5216)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5251..5270
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(5288..5743)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5425..5446)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5637..5656
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        5770..5874
                     /gene="bla"
                     /label=AmpR promoter
     promoter        5876..6233
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        5876..6067
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(5899..5919)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      6084..6219
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             6268..7062
                     /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             6268..7062
                     /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(6322..6341)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6932..6951
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(7250..7269)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    7294..7341
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      7670..8258
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8159..8178
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggtcgcca ccatggtgag caagggcgcc gagctgttca ccggcatcgt gcccatcctg
      661 atcgagctga atggcgatgt gaatggccac aagttcagcg tgagcggcga gggcgagggc
      721 gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcctgtg
      781 ccctggccca ccctggtgac caccctgagc tacggcgtgc agtgcttctc acgctacccc
      841 gatcacatga agcagcacga cttcttcaag agcgccatgc ctgagggcta catccaggag
      901 cgcaccatct tcttcgagga tgacggcaac tacaagtcgc gcgccgaggt gaagttcgag
      961 ggcgataccc tggtgaatcg catcgagctg accggcaccg atttcaagga ggatggcaac
     1021 atcctgggca ataagatgga gtacaactac aacgcccaca atgtgtacat catgaccgac
     1081 aaggccaaga atggcatcaa ggtgaacttc aagatccgcc acaacatcga ggatggcagc
     1141 gtgcagctgg ccgaccacta ccagcagaat acccccatcg gcgatggccc tgtgctgctg
     1201 cccgataacc actacctgtc cacccagagc gccctgtcca aggaccccaa cgagaagcgc
     1261 gatcacatga tctacttcgg cttcgtgacc gccgccgcca tcacccacgg catggatgag
     1321 ctgtacaagt ccggactcag atctcgagct caagcttcga attcaatggc ggcggcggcg
     1381 ggcgcccctc cgccgggtcc cccgcaaccg cctccgccgc cgccgcccga ggagtcgtcc
     1441 gacagcgagc ccgaggcgga gcccggctcc ccacagaagc tcatccgcaa ggtgtccacg
     1501 tcgggtcaga tccgacagaa gaccatcatc aaagagggga tgctgaccaa acagaacaat
     1561 tcattccagc gatcaaaaag gagatacttt aagcttcgag ggcgaacgct ttactatgcc
     1621 aaaacggcaa agtcaatcat atttgatgag gtggatctga cagatgccag cgtagctgaa
     1681 tccagtacca aaaacgtcaa caacagtttt acggtcataa ctccatgcag gaagctcatc
     1741 ttgtgtgctg ataacagaaa agaaatggaa gattggattg cagcattaaa gactgtgcag
     1801 aacagggagc actttgagcc cacccagtac agcatggacc acttctcagg gatgcacaat
     1861 tggtacgcct gttcccacgc gaggccgacc tactgcaatg tgtgccgtga ggctctgtct
     1921 ggggtcacgt cgcacgggct gtcctgcgag gtgtgcaaat ttaaggccca caagcgctgt
     1981 gctgtgcgtg caaccaataa ctgcaagtgg accacactgg cctcgatcgg gaaggacatc
     2041 attgaagatg cagatgggat tgcaatgccc caccagtggt tggaaggaaa cctacctgtg
     2101 agcgccaagt gcactgtgtg cgacaagacc tgtggcagtg tgctgcgcct gcaggactgg
     2161 cgctgcctct ggtgcaaggc catggttcac acatcgtgta aagaatcctt gctgaccaag
     2221 tgcccacttg gcctgtgcaa agtgtcagtc atcccaccca cggctctcaa cagcatcgac
     2281 tccgatgggt tctggaaggc cagctgtcct ccttcttgca caagcccact gttggtcttc
     2341 gtcaattcaa aaagtgggga caaccagggt gtgaagttcc tcagaagatt caaacagcta
     2401 ctaaaccccg cccaggtctt cgacctcatg aacggaggcc cacacctcgg cttacggtta
     2461 ttccagaagt ttgacacatt ccggattctg gtttgtggcg gggatggaag tgttggctgg
     2521 gtcctctccg aaatcgacag cctcaacctt cataaacagt gtcagctggg agtgctgccg
     2581 ctcggcacag ggaacgactt ggcccgagta ctgggctggg gctcagcctg cgatgacgac
     2641 acccagctcc cccagatctt ggagaagttg gagagagcca gcaccaagat gctggacagg
     2701 tggagcgtca tggcatacga ggccaagctc ccccggcagg cctcctcctc taccgtcacc
     2761 gaagacttca gcgaggattc cgaggtacag cagattctct tctatgaaga ctcggttgca
     2821 gcccaccttt ctaaaatcct cacctcggac cagcactcgg tggtcatctc ctcggccaaa
     2881 gtgctctgtg agacggtgaa ggacttcgtg gcacgggtgg ggaaggccta tgagaagacg
     2941 accgagagct cggaggagtc agaggtcatg gccaagaagt gctctgtcct gaaagagaag
     3001 ctggattccc ttctcaagac cttggacgat gagtcccagg cctcgtcctc tctgcccaac
     3061 ccgcccccca ccattgccga ggaggctgaa gatggagatg ggtcgggcag catctgcggt
     3121 tccaccggag accgcttggt ggcatcagct tgcccggccc ggccgcagat attccggcct
     3181 cgagaacagc tcatgctgag agccaacagc ctgaagaaag caattcgtca gatcatagaa
     3241 cacacagaaa aagctgtcga tgagcagaat gcccagaccc aggagcagga gggcttcgtc
     3301 ctgggcctct ctgagtcaga ggagaagatg gaccacagag tgtgcccacc actgtcccac
     3361 agcgagagct tcggggtccc caaggggagg agccagcgca aagtgtcgaa atctccgtgt
     3421 gaaaagctga tcagcaaagg gagtctgtcc ctaggcagtt ctgcttccct tccgccccag
     3481 ccgggaagcc gggacggcct gcctgcgctc aacaccaaga tcctgtaccc aaatgtccgg
     3541 gctggaatgt ctggttcctt acccggtggc tcagtcatca gtcgcctgtt aattaatgct
     3601 gatcccttca actctgaacc agaaacccta gagtattaca cggagaaatg tgtcatgaac
     3661 aactattttg gcattggcct ggatgcgaag atatccctgg actttaacaa caagcgcgat
     3721 gagcacccag agaagtgcag gagccgaacc aagaacatga tgtggtatgg agttcttgga
     3781 accaaagagt tgctgcacag aacctacaag aacctggagc aaaaggtctt gctggagtgt
     3841 gacgggcgac ccatcccact ccccagtctt cagggaattg ctgtccttaa cattcccagc
     3901 tatgccggag gaaccaactt ctgggggggt accaaggaag atgatacttt cgcagctcca
     3961 tcattcgatg acaagattct ggaggtggtc gccgtgttcg gcagcatgca gatggccgtc
     4021 tctcgagtca tcaggctaca gcatcatcgg atcgcccagt gtcgcacggt gaagatctcc
     4081 atccttgggg atgagggcgt gcctgtgcag gtggacggag aggcctgggt ccagccgcca
     4141 gggtacattc ggattgtcca caagaaccgg gcacagacac tgaccagaga cagggcattt
     4201 gagagcaccc tgaagtcctg ggaagacaag cagaagtgcg agctgccccg ccctccatcc
     4261 tgttccctgc acccggagat gctgtccgag gaggaggcca cccagatgga ccagtttggg
     4321 caggcagcag gggtcctcat tcacagtatc cgagaaatag ctcagtctca ccgggacatg
     4381 gagcaggaac tggcccacgc cgtcaatgcc agctccaagt ccatggaccg tgtgtatggc
     4441 aagcccagaa ccacagaggg gctcaactgc agcttcgtcc tggaaatggt gaataacttc
     4501 agagctctgc gcagtgagac ggagctgctg ctgtctggga agatggccct gcagctggat
     4561 ccgcctcaga aggagcagct ggggagtgct cttgccgaga tggaccgaca gctcaggagg
     4621 ctggcagaca ccccgtggct ctgccagtcc gcagagcccg gcgacgaaga gagtgtgatg
     4681 ctggatcttg ccaagcgcag tcgcagtggt aaattccgcc tcgtgaccaa gtttaaaaag
     4741 gagaaaaaca acaagaacaa agaagctcac agtagcctgg gagccccggt tcacctctgg
     4801 gggacagagg aggttgctgc ctggctggag cacctcagtc tctgtgagta taaggacatc
     4861 ttcacacggc acgacatccg gggctctgag ctcctgcacc tggagcggag ggacctcaag
     4921 gacctgggcg tgaccaaggt gggccacatg aagaggatcc tgtgtggcat caaggagctg
     4981 agccgcagcg cccccgccgt cgaggcctag tcgacggtac cgcgggcccg ggatccaccg
     5041 gatctagata actgatcata atcagccata ccacatttgt agaggtttta cttgctttaa
     5101 aaaacctccc acacctcccc ctgaacctga aacataaaat gaatgcaatt gttgttgtta
     5161 acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa
     5221 ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt
     5281 aacgcgtaaa ttgtaagcgt taatattttg ttaaaattcg cgttaaattt ttgttaaatc
     5341 agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag
     5401 accgagatag ggttgagtgt tgttccagtt tggaacaaga gtccactatt aaagaacgtg
     5461 gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg atggcccact acgtgaacca
     5521 tcaccctaat caagtttttt ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa
     5581 gggagccccc gatttagagc ttgacgggga aagccggcga acgtggcgag aaaggaaggg
     5641 aagaaagcga aaggagcggg cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta
     5701 accaccacac ccgccgcgct taatgcgccg ctacagggcg cgtcaggtgg cacttttcgg
     5761 ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg
     5821 ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtcctgag
     5881 gcggaaagaa ccagctgtgg aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc
     5941 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccagg tgtggaaagt
     6001 ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca
     6061 tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc gcccattctc
     6121 cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc tcggcctctg
     6181 agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaagatcgat
     6241 caagagacag gatgaggatc gtttcgcatg attgaacaag atggattgca cgcaggttct
     6301 ccggccgctt gggtggagag gctattcggc tatgactggg cacaacagac aatcggctgc
     6361 tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc cggttctttt tgtcaagacc
     6421 gacctgtccg gtgccctgaa tgaactgcaa gacgaggcag cgcggctatc gtggctggcc
     6481 acgacgggcg ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg aagggactgg
     6541 ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag
     6601 aaagtatcca tcatggctga tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc
     6661 ccattcgacc accaagcgaa acatcgcatc gagcgagcac gtactcggat ggaagccggt
     6721 cttgtcgatc aggatgatct ggacgaagag catcaggggc tcgcgccagc cgaactgttc
     6781 gccaggctca aggcgagcat gcccgacggc gaggatctcg tcgtgaccca tggcgatgcc
     6841 tgcttgccga atatcatggt ggaaaatggc cgcttttctg gattcatcga ctgtggccgg
     6901 ctgggtgtgg cggaccgcta tcaggacata gcgttggcta cccgtgatat tgctgaagag
     6961 cttggcggcg aatgggctga ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg
     7021 cagcgcatcg ccttctatcg ccttcttgac gagttcttct gagcgggact ctggggttcg
     7081 aaatgaccga ccaagcgacg cccaacctgc catcacgaga tttcgattcc accgccgcct
     7141 tctatgaaag gttgggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc
     7201 gcggggatct catgctggag ttcttcgccc accctagggg gaggctaact gaaacacgga
     7261 aggagacaat accggaagga acccgcgcta tgacggcaat aaaaagacag aataaaacgc
     7321 acggtgttgg gtcgtttgtt cataaacgcg gggttcggtc ccagggctgg cactctgtcg
     7381 ataccccacc gagaccccat tggggccaat acgcccgcgt ttcttccttt tccccacccc
     7441 accccccaag ttcgggtgaa ggcccagggc tcgcagccaa cgtcggggcg gcaggccctg
     7501 ccatagcctc aggttactca tatatacttt agattgattt aaaacttcat ttttaattta
     7561 aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt
     7621 tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt
     7681 tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt
     7741 gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc
     7801 agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg
     7861 tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg
     7921 ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt
     7981 cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac
     8041 tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg
     8101 acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg
     8161 gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat
     8221 ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt
     8281 tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg
     8341 attctgtgga taaccgtatt accgccatgc at
//
