LOCUS       sequence-477905-        8927 bp    DNA     circular SYN 08-JUL-2025
DEFINITION  Used for transfection of cells for expression of C-FLAG PDZD8-T527A.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8927)
  AUTHORS   Li M, Wang Y, Wei X, Cai WF, Wu J, Zhu M, Wang Y, Liu YH, Xiong J, 
            Qu Q, Chen Y, Tian X, Yao L, Xie R, Li X, Chen S, Huang X, Zhang C, 
            Xie C, Wu Y, Xu Z, Zhang B, Jiang B, Wang ZC, Li Q, Li G, Lin SY, Yu
            L, Piao HL, Deng X, Han J, Zhang CS, Lin SC
  TITLE     AMPK targets PDZD8 to trigger carbon source shift from glucose to 
            glutamine.
  JOURNAL   Cell Res. 2024 Jun 19. doi: 10.1038/s41422-024-00985-6.
  PUBMED    38898113
REFERENCE   2  (bases 1 to 8927)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jul 8, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Cell Res. 
            2024 Jun 19. doi: 10.1038/s41422-024-00985-6."
COMMENT     Sequence Label: pcDNA 3.3 C-FLAG PDZD8-T527A
FEATURES             Location/Qualifiers
     source          1..8927
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        50..429
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        430..633
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     584..604
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     630..654
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             4243..4266
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(4313..4332)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4357..4405
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4607..5035
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4694..4713)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4904..4925
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5044..5064)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5049..5378
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5229..5364
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5291..5310
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             5445..6239
                     /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(5499..5518)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6109..6128
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    6415..6536
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6452..6471)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6506..6525
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6585..6601)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6585..6601)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6598..6620)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6609..6625
                     /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(6633..6639,6640..6657,6658..6663))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6678..6699
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6816..6833)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6987..7575)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(7067..7086)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7746..8537,8538..8606))
                     /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     8369..8388
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8607..8711)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(8786..8805)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gttaggcgtt ttgcgctgct tcgcgatgta cgggccagat atacgcgttg acattgatta
       61 ttgactagtt attaatagta atcaattacg gggtcattag ttcatagccc atatatggag
      121 ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc
      181 ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac tttccattga
      241 cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca agtgtatcat
      301 atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc
      361 cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt agtcatcgct
      421 attaccatgg tgatgcggtt ttggcagtac atcaatgggc gtggatagcg gtttgactca
      481 cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg gcaccaaaat
      541 caacgggact ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat gggcggtagg
      601 cgtgtacggt gggaggtcta tataagcaga gctcgtttag tgaaccgtca gatcgcctgg
      661 agacgccatc cacgctgttt tgacctccat agaagacacc gggaccgatc cagcctccgg
      721 actgagctcc ttaaggttaa ctctagagaa ttcggatcca tggggctgct gctcatgatc
      781 ctggcgtcgg ccgtgctggg ttccttcctc acgctcctcg cccagttctt cctgctgtac
      841 cgcagacagc ccgagccgcc ggcggacgag gccgcccgcg cgggcgaggg cttccgctac
      901 atcaagccag tgccgggcct gctcctaagg gagtaccttt atggcggcgg ccgggatgag
      961 gagccctccg gagcggcccc tgagggcggc gcgaccccca ccgcggcccc cgagaccccc
     1021 gccccgccga cgcgggagac ttgctacttc ctcaacgcca ccatcctatt cctgttccgg
     1081 gagttgcggg acaccgcgct gacccgccgc tgggtcacca agaagatcaa ggtggagttc
     1141 gaggagctgc tgcagaccaa gacggccggg cgcctgctgg aggggctgag cctgcgggac
     1201 gtgttcctgg gcgagacggt gcccttcatc aagaccatcc ggctcgtgcg gccagtcgtg
     1261 ccctcggcca ccggggagcc cgatggccct gaaggggagg cgctgcccgc cgcctgcccc
     1321 gaggagctgg ccttcgaggc ggaggtggag tacaacgggg gcttccacct ggccatcgac
     1381 gtggacctgg tcttcggcaa gtccgcctac ttgtttgtca agctgtcccg cgtggtggga
     1441 aggctgcgct tggtctttac gcgcgtgccc ttcacccact ggttcttctc cttcgtggaa
     1501 gacccgctga tcgacttcga ggtgcgctcc cagtttgaag ggcggcccat gccccagctc
     1561 acctccatca tcgtcaacca gctcaagaag atcatcaagc gcaagcacac cctaccgaat
     1621 tacaagatca ggtttaagcc gttttttcca taccagacct tgcaaggatt tgaagaagat
     1681 gaagagcata tccatataca acaatgggca cttactgaag gccgtcttaa agttacgttg
     1741 ttagaatgta gcaggttact catttttgga tcctatgaca gagaggcaaa tgttcattgt
     1801 acacttgagt taagcagtag tgtttgggaa gaaaaacaga ggagttctat taagacggtt
     1861 gaattaataa aaggaaattt acaaagtgtt ggacttacac ttcgtcttgt ccagtcaact
     1921 gatgggtatg ctgggcacgt catcattgaa actgtggctc caaactcgcc tgctgcaatt
     1981 gcagatcttc agcggggaga tcgacttatc gccattggag gtgtgaaaat cacatcaaca
     2041 ctgcaagtgt tgaagcttat caagcaggct ggtgaccgag tcctggtgta ctatgaaagg
     2101 cctgttggcc agagtaatca aggtgcagtg ctgcaagata actttggcca gttggaagaa
     2161 aactttttgt caagctcatg ccaatcgggt tatgaagagg aagctgccgg gttgacagta
     2221 gatactgaaa gtagagagct ggattctgaa tttgaagact tggcaagtga tgtcagagca
     2281 caaaatgagt tcaaagatga ggcacaatca ttaagtcata gtcccaaacg tgttccagca
     2341 acactttcta ttaaacccct tggagctata tcaccagttt taaaccgtaa attagctgta
     2401 ggaagtcacc cactaccacc gaaaattcag tccaaagatg gaaataaacc tccaccccta
     2461 aaaacttctg agataacaga cccagcacaa gtgtcaaaac caacccaagg atctgctttc
     2521 aaaccacctg tgccaccacg accacaagcg aaagttcctt tgccttccgc cgatgctcca
     2581 aatcaggcag aaccagatgt tctcgttgaa aagccagaga aggtggtgcc acctcctctt
     2641 gtagataaat ctgctgaaaa gcaagcaaaa aatgtggatg ccatagacga tgcagctgca
     2701 cctaagcaat ttttagcaaa gcaagaagtg gccaaagatg tcacttcaga aacttcctgc
     2761 cctactaagg acagttcgga cgaccgtcaa acatgggaat catcagaaat tctttatcgt
     2821 aataagctag gaaaatggac aagaaccaga gcatcctgtt tgtttgacat agaagcctgt
     2881 cacaggtact taaacattgc attgtggtgc agggatcctt tcaagttggg aggtctcatc
     2941 tgtttggggc atgttagttt aaaacttgaa gatgtggctt taggatgcct agctacatca
     3001 aacacggaat acctttccaa attgagactg gaagccccct cacctaaggc tatagtcact
     3061 agaaccgcac tacgcaatct gagtatgcaa aagggattca atgacaaatt ttgctatggt
     3121 gacattacta ttcacttcaa atatttgaaa gaaggagaat cagaccacca tgtagttact
     3181 aacgtagaaa aagaaaaaga accccatttg gttgaagaag tttctgttct ccctaaagag
     3241 gagcaatttg ttggacagat gggtttaaca gaaaacaaac acagttttca ggatactcag
     3301 ttccagaacc caacatggtg tgactactgt aagaaaaaag tttggactaa agcagcttcc
     3361 cagtgtatgt tttgtgctta tgtttgccat aaaaaatgtc aagaaaagtg tctagctgag
     3421 acttctgttt gtggagcaac tgataggcga atagacagga cactgaaaaa ccttaggctg
     3481 gaaggacagg aaaccctctt aggcctgcct cctcgtgttg atgctgaagc tagcaagtca
     3541 gtcaataaaa caacaggttt gacaaggcat attatcaata ctagttctcg tttattaaat
     3601 ttgcgtcaag tctctaaaac tcgcctttct gaaccaggaa ccgatctcgt agaaccttca
     3661 ccaaaacaca cacccaacac gtcagacaac gaaggcagtg acacggaggt ctgtggtcca
     3721 aacagtcctt ctaaacgggg aaacagcaca ggaataaagt tagtgagaaa agagggtggt
     3781 ctggatgaca gtgttttcat tgcagttaaa gaaattggtc gtgatctgta caggggcttg
     3841 cctacagagg aaaggatcca gaaactagag ttcatgttgg ataagctaca gaatgaaatt
     3901 gatcaggagt tggaacacaa taattccctt gttagagaag aaaaagagac aactgataca
     3961 aggaaaaaat cacttctttc tgctgcctta gctaaatcag gtgaaaggct acaagctcta
     4021 acacttctta tgattcacta cagagcaggc attgaagata tagaaacttt agaaagtctg
     4081 tctttagacc agcactccaa aaaaataagc aagtacacag atgatacaga agaagacctt
     4141 gataatgaaa taagccaact aatagactct cagccattca gcagcatatc agatgactta
     4201 tttggcccat ccgagtctgt gctcgagcca tggaagctta tggactacaa agacgatgac
     4261 gacaaggata tctaactgac tgaaccggtg gtaccaaacg ggggaggcta actgaaacac
     4321 ggaaggagac aataccggaa ggaacccgcg ctatgacggc aataaaaaga cagaataaaa
     4381 cgcacgggtg ttgggtcgtt tgttcataaa cgcggggttc ggtcccaggg ctggcactct
     4441 gtcgataccc caccgagacc ccattggggc caatacgccc gcgtttcttc cttttcccca
     4501 ccccaccccc caagttcggg tgaaggccca gggctcgcag ccaacgtcgg ggcggcaggc
     4561 cctgccatag cagatctgcg cagctggggc tctagggggt atccccacgc gccctgtagc
     4621 ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc
     4681 gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt
     4741 ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac
     4801 ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag
     4861 acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa
     4921 actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg
     4981 atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattaattc
     5041 tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca ggcagaagta
     5101 tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag
     5161 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa
     5221 ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc catggctgac
     5281 taattttttt tatttatgca gaggccgagg ccgcctctgc ctctgagcta ttccagaagt
     5341 agtgaggagg cttttttgga ggcctaggct tttgcaaaaa gctcccggga gcttgtatat
     5401 ccattttcgg atctgatcaa gagacaggat gaggatcgtt tcgcatgatt gaacaagatg
     5461 gattgcacgc aggttctccg gccgcttggg tggagaggct attcggctat gactgggcac
     5521 aacagacaat cggctgctct gatgccgccg tgttccggct gtcagcgcag gggcgcccgg
     5581 ttctttttgt caagaccgac ctgtccggtg ccctgaatga actgcaggac gaggcagcgc
     5641 ggctatcgtg gctggccacg acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg
     5701 aagcgggaag ggactggctg ctattgggcg aagtgccggg gcaggatctc ctgtcatctc
     5761 accttgctcc tgccgagaaa gtatccatca tggctgatgc aatgcggcgg ctgcatacgc
     5821 ttgatccggc tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta
     5881 ctcggatgga agccggtctt gtcgatcagg atgatctgga cgaagagcat caggggctcg
     5941 cgccagccga actgttcgcc aggctcaagg cgcgcatgcc cgacggcgag gatctcgtcg
     6001 tgacccatgg cgatgcctgc ttgccgaata tcatggtgga aaatggccgc ttttctggat
     6061 tcatcgactg tggccggctg ggtgtggcgg accgctatca ggacatagcg ttggctaccc
     6121 gtgatattgc tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg ctttacggta
     6181 tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct tcttgacgag ttcttctgag
     6241 cgggactctg gggttcgcga aatgaccgac caagcgacgc ccaacctgcc atcacgagat
     6301 ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc
     6361 ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca ccccaacttg
     6421 tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa
     6481 gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat
     6541 gtctgtatac cgtcgacctc tagctagagc ttggcgtaat catggtcata gctgtttcct
     6601 gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt
     6661 aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc
     6721 gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg
     6781 agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg
     6841 gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca
     6901 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac
     6961 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac
     7021 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg
     7081 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac
     7141 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat
     7201 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag
     7261 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac
     7321 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt
     7381 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt
     7441 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc
     7501 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga
     7561 aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac
     7621 gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc
     7681 cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct
     7741 gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca
     7801 tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct
     7861 ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca
     7921 ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc
     7981 atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg
     8041 cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct
     8101 tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa
     8161 aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta
     8221 tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc
     8281 ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg
     8341 agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa
     8401 gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg
     8461 agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc
     8521 accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg
     8581 gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat
     8641 cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata
     8701 ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tcgacggatc gggagatctc
     8761 ccgatcccct atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagt
     8821 atctgctccc tgcttgtgtg ttggaggtcg ctgagtagtg cgcgagcaaa atttaagcta
     8881 caacaaggca aggcttgacc gacaattgca tgaagaatct gcttagg
//
