LOCUS       sequence-477890-        5773 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  To express in E. coli AMPK alpha-KD(27-290aa).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5773)
  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 5773)
  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: "Cell Res. 
            2024 Jun 19. doi: 10.1038/s41422-024-00985-6."
COMMENT     Sequence Label: pGEX4T-1 AMPK alpha-KD(27-290aa)
FEATURES             Location/Qualifiers
     source          1..5773
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(181..186,187..202,203..209)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    217..233
                     /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             256..909
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     867..889
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             916..933
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     primer_bind     complement(1821..1843)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1988..2006)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2074..2178
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2179..2247,2248..3039)
                     /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(2397..2416)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3210..3798
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3699..3718
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3952..3969
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        4042..4119
                     /gene="lacI"
                     /label=lacI promoter
     CDS             4120..5202
                     /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(4139..4158)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(5251..5256,5257..5274,5275..5281)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5289..5305
                     /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     5313..5329
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5313..5329
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(5325..5773,1..79)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(5345..5362)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5345..5361)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5354..5376)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 cttatcgact gcacggtgca ccaatgcttc tggcgtcagg cagccatcgg aagctgtggt
       61 atggctgtgc aggtcgtaaa tcactgcata attcgtgtcg ctcaaggcgc actcccgttc
      121 tggataatgt tttttgcgcc gacatcataa cggttctggc aaatattctg aaatgagctg
      181 ttgacaatta atcatcggct cgtataatgt gtggaattgt gagcggataa caatttcaca
      241 caggaaacag tattcatgtc ccctatacta ggttattgga aaattaaggg ccttgtgcaa
      301 cccactcgac ttcttttgga atatcttgaa gaaaaatatg aagagcattt gtatgagcgc
      361 gatgaaggtg ataaatggcg aaacaaaaag tttgaattgg gtttggagtt tcccaatctt
      421 ccttattata ttgatggtga tgttaaatta acacagtcta tggccatcat acgttatata
      481 gctgacaagc acaacatgtt gggtggttgt ccaaaagagc gtgcagagat ttcaatgctt
      541 gaaggagcgg ttttggatat tagatacggt gtttcgagaa ttgcatatag taaagacttt
      601 gaaactctca aagttgattt tcttagcaag ctacctgaaa tgctgaaaat gttcgaagat
      661 cgtttatgtc ataaaacata tttaaatggt gatcatgtaa cccatcctga cttcatgttg
      721 tatgacgctc ttgatgttgt tttatacatg gacccaatgt gcctggatgc gttcccaaaa
      781 ttagtttgtt ttaaaaaacg tattgaagct atcccacaaa ttgataagta cttgaaatcc
      841 agcaagtata tagcatggcc tttgcagggc tggcaagcca cgtttggtgg tggcgaccat
      901 cctccaaaat cggatctggt tccgcgtgga tcgagagaat tcggatccta catcctgggg
      961 gacacgcttg gtgtcggcac cttcgggaaa gtgaaggtgg gcaagcacga gttgaccgga
     1021 cataaagtgg ctgtgaagat actcaaccgg cagaagattc ggagccttga cgtggtggga
     1081 aaaatccgcc gggagattca gaacctgaag ctgttcaggc accctcacat catcaaactg
     1141 taccaggtca tcagtacacc atctgatatt ttcatggtga tggaatatgt ctctggagga
     1201 gagctatttg attatatctg taaaaatgga aggttggacg aaaaggaaag ccgccgtctg
     1261 ttccagcaga tcctttccgg tgtggattat tgtcacaggc atatggtggt ccacagagat
     1321 ttgaaacctg agaacgtcct gcttgatgca cacatgaatg caaagatagc cgactttggt
     1381 ctttcaaaca tgatgtcaga tggtgaattt ttaagaacaa gctgtggctc acccaattat
     1441 gccgcaccag aagtcatttc aggaagattg tacgcaggcc ccgaggtgga catctggagc
     1501 agcggggtca ttctctatgc tttgctgtgt ggaaccctcc cttttgatga tgaccatgtg
     1561 ccaactcttt ttaagaagat atgtgatggg atcttttata cccctcagta cttaaaccct
     1621 tcagtaatca gccttttgaa acatatgctg caggtggatc ccatgaagag ggccacgata
     1681 aaagatatca gggaacacga gtggtttaaa caggaccttc cgaagtatct ctttcctgag
     1741 ctcgagccat ggaaggtcga gcggccgcat cgtgactgac tgacgatctg cctcgcgcgt
     1801 ttcggtgatg acggtgaaaa cctctgacac atgcagctcc cggagacggt cacagcttgt
     1861 ctgtaagcgg atgccgggag cagacaagcc cgtcagggcg cgtcagcggg tgttggcggg
     1921 tgtcggggcg cagccatgac ccagtcacgt agcgatagcg gagtgtataa ttcttgaaga
     1981 cgaaagggcc tcgtgatacg cctattttta taggttaatg tcatgataat aatggtttct
     2041 tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc
     2101 taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa
     2161 tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt
     2221 gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct
     2281 gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc
     2341 cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta
     2401 tgtggcgcgg tattatcccg tgttgacgcc gggcaagagc aactcggtcg ccgcatacac
     2461 tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc
     2521 atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac
     2581 ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg
     2641 gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac
     2701 gagcgtgaca ccacgatgcc tgcagcaatg gcaacaacgt tgcgcaaact attaactggc
     2761 gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt
     2821 gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga
     2881 gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc
     2941 cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag
     3001 atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca
     3061 tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc
     3121 ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca
     3181 gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc
     3241 tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta
     3301 ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgtcctt
     3361 ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc
     3421 gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg
     3481 ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg
     3541 tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag
     3601 ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc
     3661 agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat
     3721 agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg
     3781 gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc
     3841 tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt
     3901 accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca
     3961 gtgagcgagg aagcggaaga gcgcctgatg cggtattttc tccttacgca tctgtgcggt
     4021 atttcacacc gcataaattc cgacaccatc gaatggcgca aaacctttcg cggtatggca
     4081 tgatagcgcc cggaagagag tcaattcagg gtggtgaatg tgaaaccagt aacgttatac
     4141 gatgtcgcag agtatgccgg tgtctcttat cagaccgttt cccgcgtggt gaaccaggcc
     4201 agccacgttt ctgcgaaaac gcgggaaaaa gtggaagcgg cgatggcgga gctgaattac
     4261 attcccaacc gcgtggcaca acaactggcg ggcaaacagt cgttgctgat tggcgttgcc
     4321 acctccagtc tggccctgca cgcgccgtcg caaattgtcg cggcgattaa atctcgcgcc
     4381 gatcaactgg gtgccagcgt ggtggtgtcg atggtagaac gaagcggcgt cgaagcctgt
     4441 aaagcggcgg tgcacaatct tctcgcgcaa cgcgtcagtg ggctgatcat taactatccg
     4501 ctggatgacc aggatgccat tgctgtggaa gctgcctgca ctaatgttcc ggcgttattt
     4561 cttgatgtct ctgaccagac acccatcaac agtattattt tctcccatga agacggtacg
     4621 cgactgggcg tggagcatct ggtcgcattg ggtcaccagc aaatcgcgct gttagcgggc
     4681 ccattaagtt ctgtctcggc gcgtctgcgt ctggctggct ggcataaata tctcactcgc
     4741 aatcaaattc agccgatagc ggaacgggaa ggcgactgga gtgccatgtc cggttttcaa
     4801 caaaccatgc aaatgctgaa tgagggcatc gttcccactg cgatgctggt tgccaacgat
     4861 cagatggcgc tgggcgcaat gcgcgccatt accgagtccg ggctgcgcgt tggtgcggat
     4921 atctcggtag tgggatacga cgataccgaa gacagctcat gttatatccc gccgttaacc
     4981 accatcaaac aggattttcg cctgctgggg caaaccagcg tggaccgctt gctgcaactc
     5041 tctcagggcc aggcggtgaa gggcaatcag ctgttgcccg tctcactggt gaaaagaaaa
     5101 accaccctgg cgcccaatac gcaaaccgcc tctccccgcg cgttggccga ttcattaatg
     5161 cagctggcac gacaggtttc ccgactggaa agcgggcagt gagcgcaacg caattaatgt
     5221 aagttagctc actcattagg caccccaggc tttacacttt atgcttccgg ctcgtatgtt
     5281 gtgtggaatt gtgagcggat aacaatttca cacaggaaac agctatgacc atgattacgg
     5341 attcactggc cgtcgtttta caacgtcgtg actgggaaaa ccctggcgtt acccaactta
     5401 atcgccttgc agcacatccc cctttcgcca gctggcgtaa tagcgaagag gcccgcaccg
     5461 atcgcccttc ccaacagttg cgcagcctga atggcgaatg gcgctttgcc tggtttccgg
     5521 caccagaagc ggtgccggaa agctggctgg agtgcgatct tcctgaggcc gatactgtcg
     5581 tcgtcccctc aaactggcag atgcacggtt acgatgcgcc catctacacc aacgtgacct
     5641 atcccattac ggtcaatccg ccgtttgttc ccacggagaa tccgacgggt tgttactcgc
     5701 tcacatttaa tgttgatgaa agctggctac aggaaggcca gacgcgaatt atttttgatg
     5761 gcgttggaat tag
//
