LOCUS       sequence-208595-        7285 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of PHGDH C234S in bacteria.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7285)
  AUTHORS   Pacold ME, Brimacombe KR, Chan SH, Rohde JM, Lewis CA, Swier LJ, 
            Possemato R, Chen WW, Sullivan LB, Fiske BP, Cho S, Freinkman E, 
            Birsoy K, Abu-Remaileh M, Shaul YD, Liu CM, Zhou M, Koh MJ, Chung H,
            Davidson SM, Luengo A, Wang AQ, Xu X, Yasgar A, Liu L, Rai G, 
            Westover KD, Vander Heiden MG, Shen M, Gray NS, Boxer MB, Sabatini 
            DM
  TITLE     A PHGDH inhibitor reveals coordination of serine synthesis and 
            one-carbon unit fate.
  JOURNAL   Nat Chem Biol. 2016 Jun;12(6):452-8. doi: 10.1038/nchembio.2070. 
            Epub 2016 Apr 25.
  PUBMED    27110680
REFERENCE   2  (bases 1 to 7285)
  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"; doi: 
            "10.1038/nchembio.2070"; journalName: "Nat Chem Biol"; date: 
            "2016-06"; volume: "12"; issue: "6"; pages: "452-8"
COMMENT     Sequence Label: pET30-2-PHGDH-C234S
FEATURES             Location/Qualifiers
     source          1..7285
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(26..73)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     69..87
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(1755..1772)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     protein_bind    1821..1845
                     /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     complement(1845..1864)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1846..1864)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1948..1967)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        2173..2250
                     /gene="lacI"
                     /label=lacI promoter
     CDS             2251..3333
                     /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(2270..2289)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             4645..4836
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(4830..4852)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4938..5077
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4993..5012
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(5092..5109)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5263..5851)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5343..5362)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6022..6813,6814..6882))
                     /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     6645..6664
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6883..6987)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7055..7073
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 atccggatat agttcctcct ttcagcaaaa aacccctcaa gacccgttta gaggccccaa
       61 ggggttatgc tagttattgc tcagcggtgg cagcagccaa ctcagcttcc tttcgggctt
      121 tgttagcagc cggatcctta gaagtggaac tggaaggctt cagtcacatg ctgcttccac
      181 gcttccaggc tgggcagcaa ggaggagatg cccatgacgt gccaggtctc cccatctgac
      241 accagtgaag tctggtagga cagcagccgc acgcctgcct ctgccaggag gccaatcatg
      301 gtaggcagca ttgcagggtc agaggtctga gtccggaata ggagcagggg caggtccctg
      361 cggagaggca cttctggcct gaagacagct ccattgagcc cctgcagtac aggtgtagtg
      421 ccttggacca agcccacagc ctggtaaggg gcgcctgcca gggccacggc caggaggcat
      481 tccccgaagc cttgctcccc tggtgcagca gggctgtggg aggtggtgac attgaggcca
      541 gcctctttca ccagcagctt agcgttcacc aagttcacat ccgcctgctt ggaagcctct
      601 ttcaggaggc cgacaatgac tgcggggctt aggcagttcc cagcattctt cagggatgtt
      661 ccctgtgtta tcacctggat ggtccctttg ggggacccag cccaggctcg catcagtgtc
      721 cccagagctt ctgccagacc aatccaaggc ttggtgtgtg gagagaaggc actggtaagg
      781 gcctgggcat tcacaacccc cgtgagagat ttccccttca ccatgtccac gaactgaaca
      841 gcaatttcct ccccacagcg gctctgagcc tccttggtgc tggcacccag gtggggacag
      901 ctgatgacat tctcatggtc caccaaggcc cggtcccgtg gcggctcttc cgtaaacacg
      961 tccagtgcag ccccggcaca ctggccagac tgcagggccc ggagcagggc gccttcgtcc
     1021 acgatccctc cacgggcaga gttcaccaca cgcaccccct tcttgcactg ggcaaaggtg
     1081 ttgtcattca gcaagcctgt cgtggagggc aggagaggag tgtgcacagt gatgaaatca
     1141 cagagaggcc agatctcctc caggggcagc tgctgaacac caaaggaggc cgagacctct
     1201 ggggaaatga tggggtcata ccctatagtc ttcatcccaa aggactgcat ccgggtagct
     1261 acctctctcc caatcctgcc caggccaaga attcccaggg tctttccatt cagctctgtt
     1321 cccatgaact tcttccgctc ccatttgccg tccttcatcg aagccgtcgc ctggggaatc
     1381 tgcctggcca ggcacatgat cattccacaa gtgagttctg cggcactgag gctgttccca
     1441 ttgggggtgt tcataaccaa gatgcccttc cttgttgcgg cctccagatc cacattgtcc
     1501 acacctgtgc cagccctgcc caccacctgg agtttctcag ctgcgttgat gacatcagcg
     1561 gtcaccttgg tggcagagcg aacaataagg ccttcacagt cctgcagctc cgctatcagc
     1621 tcctctttgc taaggttctg cttttccacc acctgcagcc ctccatcttg caagatcttc
     1681 cggcagcaag ggtccaggct gtcactgatg agcactttcc gcagatttgc aaaagccatc
     1741 tcgagcgaat tcacgtgatg atgatgatga tgcatatgta tatctccttc ttaaagttta
     1801 aacaaaatta tttctagagg ggaattgtta tccgctcaca attcccctat agtgagtcgt
     1861 attaatttcg cgggatcgag atctcgatcc tctacgccgg acgcatcgtg gccggcatca
     1921 ccggcgccac aggtgcggtt gctggcgcct atatcgccga catcaccgat ggggaagatc
     1981 gggctcgcca cttcgggctc atgagcgctt gtttcggcgt gggtatggtg gcaggccccg
     2041 tggccggggg actgttgggc gccatctcct tgcatgcacc attccttgcg gcggcggtgc
     2101 tcaacggcct caacctacta ctgggctgct tcctaatgca ggagtcgcat aagggagagc
     2161 gtcgagatcc cggacaccat cgaatggcgc aaaacctttc gcggtatggc atgatagcgc
     2221 ccggaagaga gtcaattcag ggtggtgaat gtgaaaccag taacgttata cgatgtcgca
     2281 gagtatgccg gtgtctctta tcagaccgtt tcccgcgtgg tgaaccaggc cagccacgtt
     2341 tctgcgaaaa cgcgggaaaa agtggaagcg gcgatggcgg agctgaatta cattcccaac
     2401 cgcgtggcac aacaactggc gggcaaacag tcgttgctga ttggcgttgc cacctccagt
     2461 ctggccctgc acgcgccgtc gcaaattgtc gcggcgatta aatctcgcgc cgatcaactg
     2521 ggtgccagcg tggtggtgtc gatggtagaa cgaagcggcg tcgaagcctg taaagcggcg
     2581 gtgcacaatc ttctcgcgca acgcgtcagt gggctgatca ttaactatcc gctggatgac
     2641 caggatgcca ttgctgtgga agctgcctgc actaatgttc cggcgttatt tcttgatgtc
     2701 tctgaccaga cacccatcaa cagtattatt ttctcccatg aagacggtac gcgactgggc
     2761 gtggagcatc tggtcgcatt gggtcaccag caaatcgcgc tgttagcggg cccattaagt
     2821 tctgtctcgg cgcgtctgcg tctggctggc tggcataaat atctcactcg caatcaaatt
     2881 cagccgatag cggaacggga aggcgactgg agtgccatgt ccggttttca acaaaccatg
     2941 caaatgctga atgagggcat cgttcccact gcgatgctgg ttgccaacga tcagatggcg
     3001 ctgggcgcaa tgcgcgccat taccgagtcc gggctgcgcg ttggtgcgga tatctcggta
     3061 gtgggatacg acgataccga agacagctca tgttatatcc cgccgttaac caccatcaaa
     3121 caggattttc gcctgctggg gcaaaccagc gtggaccgct tgctgcaact ctctcagggc
     3181 caggcggtga agggcaatca gctgttgccc gtctcactgg tgaaaagaaa aaccaccctg
     3241 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
     3301 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg taagttagct
     3361 cactcattag gcaccgggat ctcgaccgat gcccttgaga gccttcaacc cagtcagctc
     3421 cttccggtgg gcgcggggca tgactatcgt cgccgcactt atgactgtct tctttatcat
     3481 gcaactcgta ggacaggtgc cggcagcgct ctgggtcatt ttcggcgagg accgctttcg
     3541 ctggagcgcg acgatgatcg gcctgtcgct tgcggtattc ggaatcttgc acgccctcgc
     3601 tcaagccttc gtcactggtc ccgccaccaa acgtttcggc gagaagcagg ccattatcgc
     3661 cggcatggcg gccgacgcgc tgggctacgt cttgctggcg ttcgcgacgc gaggctggat
     3721 ggccttcccc attatgattc ttctcgcttc cggcggcatc gggatgcccg cgttgcaggc
     3781 catgctgtcc aggcaggtag atgacgacca tcagggacag cttcaaggat cgctcgcggc
     3841 tcttaccagc ctaacttcga tcattggacc gctgatcgtc acggcgattt atgccgcctc
     3901 ggcgagcaca tggaacgggt tggcatggat tgtaggcgcc gccctatacc ttgtctgcct
     3961 ccccgcgttg cgtcgcggtg catggagccg ggccacctcg acctgaatgg aagccggcgg
     4021 cacctcgcta acggattcac cactccaaga attggagcca atcaattctt gcggagaact
     4081 gtgaatgcgc aaaccaaccc ttggcagaac atatccatcg cgtccgccat ctccagcagc
     4141 cgcacgcggc gcatctcggg cagcgttggg tcctggccac gggtgcgcat gatcgtgctc
     4201 ctgtcgttga ggacccggct aggctggcgg ggttgcctta ctggttagca gaatgaatca
     4261 ccgatacgcg agcgaacgtg aagcgactgc tgctgcaaaa cgtctgcgac ctgagcaaca
     4321 acatgaatgg tcttcggttt ccgtgtttcg taaagtctgg aaacgcggaa gtcagcgccc
     4381 tgcaccatta tgttccggat ctgcatcgca ggatgctgct ggctaccctg tggaacacct
     4441 acatctgtat taacgaagcg ctggcattga ccctgagtga tttttctctg gtcccgccgc
     4501 atccataccg ccagttgttt accctcacaa cgttccagta accgggcatg ttcatcatca
     4561 gtaacccgta tcgtgagcat cctctctcgt ttcatcggta tcattacccc catgaacaga
     4621 aatccccctt acacggaggc atcagtgacc aaacaggaaa aaaccgccct taacatggcc
     4681 cgctttatca gaagccagac attaacgctt ctggagaaac tcaacgagct ggacgcggat
     4741 gaacaggcag acatctgtga atcgcttcac gaccacgctg atgagcttta ccgcagctgc
     4801 ctcgcgcgtt tcggtgatga cggtgaaaac ctctgacaca tgcagctccc ggagacggtc
     4861 acagcttgtc tgtaagcgga tgccgggagc agacaagccc gtcagggcgc gtcagcgggt
     4921 gttggcgggt gtcggggcgc agccatgacc cagtcacgta gcgatagcgg agtgtatact
     4981 ggcttaacta tgcggcatca gagcagattg tactgagagt gcaccattgc ggtgtgaaat
     5041 accgcacaga tgcgtaagga gaaaataccg catcaggcgc tcttccgctt cctcgctcac
     5101 tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt
     5161 aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca
     5221 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
     5281 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
     5341 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
     5401 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag
     5461 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca
     5521 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa
     5581 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc
     5641 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag
     5701 aaggacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg
     5761 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca
     5821 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc
     5881 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag
     5941 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata
     6001 tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat
     6061 ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg
     6121 ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc
     6181 tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc
     6241 aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc
     6301 gccagttaat agtttgcgca acgttgttgc cattgctgca ggcatcgtgg tgtcacgctc
     6361 gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc
     6421 ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa
     6481 gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat
     6541 gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata
     6601 gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaaca cgggataata ccgcgccaca
     6661 tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag
     6721 gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc
     6781 agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc
     6841 aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata
     6901 ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta
     6961 gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacgtcta
     7021 agaaaccatt attatcatga cattaaccta taaaaatagg cgtatcacga ggccctttcg
     7081 tcttcaagaa ttaattctca tgtttgacag cttatcatcg ataagctagc tttaatgcgg
     7141 tagtttatca cagttaaatt gctaacgcag tcaggcaccg tgtatgaaat ctaacaatgc
     7201 gctcatcgtc atcctcggca ccgtcaccct ggatgctgta ggcataggct tggttatgcc
     7261 ggtactgccg ggcctcttgc gggat
//
