LOCUS       sequence-477865-        8443 bp    DNA     circular SYN 08-JUL-2025
DEFINITION  To express in E. coli PDZD8-T527A.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8443)
  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 8443)
  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: pGEX4T-1 PDZD8-T527A
FEATURES             Location/Qualifiers
     source          1..8443
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(182..187,188..203,204..210)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    218..234
                     /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)."
     CDS             257..910
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     868..890
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             917..934
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     primer_bind     complement(4492..4514)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4659..4677)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4745..4849
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4850..4918,4919..5710)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(5068..5087)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5881..6469
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6370..6389
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6623..6640
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        6713..6790
                     /gene="lacI"
                     /label=lacI promoter
     CDS             6791..7873
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(6810..6829)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(7922..7927,7928..7945,7946..7952)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7960..7976
                     /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     7984..8000
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7984..8000
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(7996..8443,1..80)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(8016..8033)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(8016..8032)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(8025..8047)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 gcttatcgac tgcacggtgc accaatgctt ctggcgtcag gcagccatcg gaagctgtgg
       61 tatggctgtg caggtcgtaa atcactgcat aattcgtgtc gctcaaggcg cactcccgtt
      121 ctggataatg ttttttgcgc cgacatcata acggttctgg caaatattct gaaatgagct
      181 gttgacaatt aatcatcggc tcgtataatg tgtggaattg tgagcggata acaatttcac
      241 acaggaaaca gtattcatgt cccctatact aggttattgg aaaattaagg gccttgtgca
      301 acccactcga cttcttttgg aatatcttga agaaaaatat gaagagcatt tgtatgagcg
      361 cgatgaaggt gataaatggc gaaacaaaaa gtttgaattg ggtttggagt ttcccaatct
      421 tccttattat attgatggtg atgttaaatt aacacagtct atggccatca tacgttatat
      481 agctgacaag cacaacatgt tgggtggttg tccaaaagag cgtgcagaga tttcaatgct
      541 tgaaggagcg gttttggata ttagatacgg tgtttcgaga attgcatata gtaaagactt
      601 tgaaactctc aaagttgatt ttcttagcaa gctacctgaa atgctgaaaa tgttcgaaga
      661 tcgtttatgt cataaaacat atttaaatgg tgatcatgta acccatcctg acttcatgtt
      721 gtatgacgct cttgatgttg ttttatacat ggacccaatg tgcctggatg cgttcccaaa
      781 attagtttgt tttaaaaaac gtattgaagc tatcccacaa attgataagt acttgaaatc
      841 cagcaagtat atagcatggc ctttgcaggg ctggcaagcc acgtttggtg gtggcgacca
      901 tcctccaaaa tcggatctgg ttccgcgtgg atcgagagaa ttcggatcca tggggctgct
      961 gctcatgatc ctggcgtcgg ccgtgctggg ttccttcctc acgctcctcg cccagttctt
     1021 cctgctgtac cgcagacagc ccgagccgcc ggcggacgag gccgcccgcg cgggcgaggg
     1081 cttccgctac atcaagccag tgccgggcct gctcctaagg gagtaccttt atggcggcgg
     1141 ccgggatgag gagccctccg gagcggcccc tgagggcggc gcgaccccca ccgcggcccc
     1201 cgagaccccc gccccgccga cgcgggagac ttgctacttc ctcaacgcca ccatcctatt
     1261 cctgttccgg gagttgcggg acaccgcgct gacccgccgc tgggtcacca agaagatcaa
     1321 ggtggagttc gaggagctgc tgcagaccaa gacggccggg cgcctgctgg aggggctgag
     1381 cctgcgggac gtgttcctgg gcgagacggt gcccttcatc aagaccatcc ggctcgtgcg
     1441 gccagtcgtg ccctcggcca ccggggagcc cgatggccct gaaggggagg cgctgcccgc
     1501 cgcctgcccc gaggagctgg ccttcgaggc ggaggtggag tacaacgggg gcttccacct
     1561 ggccatcgac gtggacctgg tcttcggcaa gtccgcctac ttgtttgtca agctgtcccg
     1621 cgtggtggga aggctgcgct tggtctttac gcgcgtgccc ttcacccact ggttcttctc
     1681 cttcgtggaa gacccgctga tcgacttcga ggtgcgctcc cagtttgaag ggcggcccat
     1741 gccccagctc acctccatca tcgtcaacca gctcaagaag atcatcaagc gcaagcacac
     1801 cctaccgaat tacaagatca ggtttaagcc gttttttcca taccagacct tgcaaggatt
     1861 tgaagaagat gaagagcata tccatataca acaatgggca cttactgaag gccgtcttaa
     1921 agttacgttg ttagaatgta gcaggttact catttttgga tcctatgaca gagaggcaaa
     1981 tgttcattgt acacttgagt taagcagtag tgtttgggaa gaaaaacaga ggagttctat
     2041 taagacggtt gaattaataa aaggaaattt acaaagtgtt ggacttacac ttcgtcttgt
     2101 ccagtcaact gatgggtatg ctgggcacgt catcattgaa actgtggctc caaactcgcc
     2161 tgctgcaatt gcagatcttc agcggggaga tcgacttatc gccattggag gtgtgaaaat
     2221 cacatcaaca ctgcaagtgt tgaagcttat caagcaggct ggtgaccgag tcctggtgta
     2281 ctatgaaagg cctgttggcc agagtaatca aggtgcagtg ctgcaagata actttggcca
     2341 gttggaagaa aactttttgt caagctcatg ccaatcgggt tatgaagagg aagctgccgg
     2401 gttgacagta gatactgaaa gtagagagct ggattctgaa tttgaagact tggcaagtga
     2461 tgtcagagca caaaatgagt tcaaagatga ggcacaatca ttaagtcata gtcccaaacg
     2521 tgttccagca acactttcta ttaaacccct tggagctata tcaccagttt taaaccgtaa
     2581 attagctgta ggaagtcacc cactaccacc gaaaattcag tccaaagatg gaaataaacc
     2641 tccaccccta aaaacttctg agataacaga cccagcacaa gtgtcaaaac caacccaagg
     2701 atctgctttc aaaccacctg tgccaccacg accacaagcg aaagttcctt tgccttccgc
     2761 cgatgctcca aatcaggcag aaccagatgt tctcgttgaa aagccagaga aggtggtgcc
     2821 acctcctctt gtagataaat ctgctgaaaa gcaagcaaaa aatgtggatg ccatagacga
     2881 tgcagctgca cctaagcaat ttttagcaaa gcaagaagtg gccaaagatg tcacttcaga
     2941 aacttcctgc cctactaagg acagttcgga cgaccgtcaa acatgggaat catcagaaat
     3001 tctttatcgt aataagctag gaaaatggac aagaaccaga gcatcctgtt tgtttgacat
     3061 agaagcctgt cacaggtact taaacattgc attgtggtgc agggatcctt tcaagttggg
     3121 aggtctcatc tgtttggggc atgttagttt aaaacttgaa gatgtggctt taggatgcct
     3181 agctacatca aacacggaat acctttccaa attgagactg gaagccccct cacctaaggc
     3241 tatagtcact agaaccgcac tacgcaatct gagtatgcaa aagggattca atgacaaatt
     3301 ttgctatggt gacattacta ttcacttcaa atatttgaaa gaaggagaat cagaccacca
     3361 tgtagttact aacgtagaaa aagaaaaaga accccatttg gttgaagaag tttctgttct
     3421 ccctaaagag gagcaatttg ttggacagat gggtttaaca gaaaacaaac acagttttca
     3481 ggatactcag ttccagaacc caacatggtg tgactactgt aagaaaaaag tttggactaa
     3541 agcagcttcc cagtgtatgt tttgtgctta tgtttgccat aaaaaatgtc aagaaaagtg
     3601 tctagctgag acttctgttt gtggagcaac tgataggcga atagacagga cactgaaaaa
     3661 ccttaggctg gaaggacagg aaaccctctt aggcctgcct cctcgtgttg atgctgaagc
     3721 tagcaagtca gtcaataaaa caacaggttt gacaaggcat attatcaata ctagttctcg
     3781 tttattaaat ttgcgtcaag tctctaaaac tcgcctttct gaaccaggaa ccgatctcgt
     3841 agaaccttca ccaaaacaca cacccaacac gtcagacaac gaaggcagtg acacggaggt
     3901 ctgtggtcca aacagtcctt ctaaacgggg aaacagcaca ggaataaagt tagtgagaaa
     3961 agagggtggt ctggatgaca gtgttttcat tgcagttaaa gaaattggtc gtgatctgta
     4021 caggggcttg cctacagagg aaaggatcca gaaactagag ttcatgttgg ataagctaca
     4081 gaatgaaatt gatcaggagt tggaacacaa taattccctt gttagagaag aaaaagagac
     4141 aactgataca aggaaaaaat cacttctttc tgctgcctta gctaaatcag gtgaaaggct
     4201 acaagctcta acacttctta tgattcacta cagagcaggc attgaagata tagaaacttt
     4261 agaaagtctg tctttagacc agcactccaa aaaaataagc aagtacacag atgatacaga
     4321 agaagacctt gataatgaaa taagccaact aatagactct cagccattca gcagcatatc
     4381 agatgactta tttggcccat ccgagtctgt gctcgagcca tggaaggtcg agcggccgca
     4441 tcgtgactga ctgacgatct gcctcgcgcg tttcggtgat gacggtgaaa acctctgaca
     4501 catgcagctc ccggagacgg tcacagcttg tctgtaagcg gatgccggga gcagacaagc
     4561 ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc gcagccatga cccagtcacg
     4621 tagcgatagc ggagtgtata attcttgaag acgaaagggc ctcgtgatac gcctattttt
     4681 ataggttaat gtcatgataa taatggtttc ttagacgtca ggtggcactt ttcggggaaa
     4741 tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat
     4801 gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca
     4861 acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca
     4921 cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta
     4981 catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt
     5041 tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc gtgttgacgc
     5101 cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc
     5161 accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc
     5221 cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa
     5281 ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga
     5341 accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgcagcaat
     5401 ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca
     5461 attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc
     5521 ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat
     5581 tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag
     5641 tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa
     5701 gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca
     5761 tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc
     5821 ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
     5881 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
     5941 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
     6001 cagcagagcg cagataccaa atactgtcct tctagtgtag ccgtagttag gccaccactt
     6061 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
     6121 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
     6181 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
     6241 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
     6301 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
     6361 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
     6421 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
     6481 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
     6541 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
     6601 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcctgat
     6661 gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcataaatt ccgacaccat
     6721 cgaatggcgc aaaacctttc gcggtatggc atgatagcgc ccggaagaga gtcaattcag
     6781 ggtggtgaat gtgaaaccag taacgttata cgatgtcgca gagtatgccg gtgtctctta
     6841 tcagaccgtt tcccgcgtgg tgaaccaggc cagccacgtt tctgcgaaaa cgcgggaaaa
     6901 agtggaagcg gcgatggcgg agctgaatta cattcccaac cgcgtggcac aacaactggc
     6961 gggcaaacag tcgttgctga ttggcgttgc cacctccagt ctggccctgc acgcgccgtc
     7021 gcaaattgtc gcggcgatta aatctcgcgc cgatcaactg ggtgccagcg tggtggtgtc
     7081 gatggtagaa cgaagcggcg tcgaagcctg taaagcggcg gtgcacaatc ttctcgcgca
     7141 acgcgtcagt gggctgatca ttaactatcc gctggatgac caggatgcca ttgctgtgga
     7201 agctgcctgc actaatgttc cggcgttatt tcttgatgtc tctgaccaga cacccatcaa
     7261 cagtattatt ttctcccatg aagacggtac gcgactgggc gtggagcatc tggtcgcatt
     7321 gggtcaccag caaatcgcgc tgttagcggg cccattaagt tctgtctcgg cgcgtctgcg
     7381 tctggctggc tggcataaat atctcactcg caatcaaatt cagccgatag cggaacggga
     7441 aggcgactgg agtgccatgt ccggttttca acaaaccatg caaatgctga atgagggcat
     7501 cgttcccact gcgatgctgg ttgccaacga tcagatggcg ctgggcgcaa tgcgcgccat
     7561 taccgagtcc gggctgcgcg ttggtgcgga tatctcggta gtgggatacg acgataccga
     7621 agacagctca tgttatatcc cgccgttaac caccatcaaa caggattttc gcctgctggg
     7681 gcaaaccagc gtggaccgct tgctgcaact ctctcagggc caggcggtga agggcaatca
     7741 gctgttgccc gtctcactgg tgaaaagaaa aaccaccctg gcgcccaata cgcaaaccgc
     7801 ctctccccgc gcgttggccg attcattaat gcagctggca cgacaggttt cccgactgga
     7861 aagcgggcag tgagcgcaac gcaattaatg taagttagct cactcattag gcaccccagg
     7921 ctttacactt tatgcttccg gctcgtatgt tgtgtggaat tgtgagcgga taacaatttc
     7981 acacaggaaa cagctatgac catgattacg gattcactgg ccgtcgtttt acaacgtcgt
     8041 gactgggaaa accctggcgt tacccaactt aatcgccttg cagcacatcc ccctttcgcc
     8101 agctggcgta atagcgaaga ggcccgcacc gatcgccctt cccaacagtt gcgcagcctg
     8161 aatggcgaat ggcgctttgc ctggtttccg gcaccagaag cggtgccgga aagctggctg
     8221 gagtgcgatc ttcctgaggc cgatactgtc gtcgtcccct caaactggca gatgcacggt
     8281 tacgatgcgc ccatctacac caacgtgacc tatcccatta cggtcaatcc gccgtttgtt
     8341 cccacggaga atccgacggg ttgttactcg ctcacattta atgttgatga aagctggcta
     8401 caggaaggcc agacgcgaat tatttttgat ggcgttggaa tta
//
