LOCUS       sequence-444360-        6928 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  IPTG-inducible expression of C. elegans protein GSPD-1(R252L) with a
            His tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6928)
  AUTHORS   Loges LN, Walstrom KM
  TITLE     Modeling human glucose-6-phosphate dehydrogenase mutations using C. 
            elegans GSPD-1.
  JOURNAL   MicroPubl Biol. 2021 Sep 10;2021:10.17912/micropub.biology.000451. 
            doi: 10.17912/micropub.biology.000451. eCollection 2021.
  PUBMED    34532700
REFERENCE   2  (bases 1 to 6928)
  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: "MicroPubl 
            Biol."; date: "2021-09-10"; volume: 
            "2021:10.17912/micropub.biology.000451. doi"; pages: " 
            10.17912/micropub.biology.000451. eCollection 2021"
COMMENT     Sequence Label: GSPD1_R252L
FEATURES             Location/Qualifiers
     source          1..6928
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     82..101
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(121..198)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     404..423
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     507..526
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        507..525
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    526..550
                     /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)."
     RBS             566..586
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             2166..2183
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2236..2254)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2250..2297
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2334..2789
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2421..2440)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2631..2652
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2815..2919
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2920..2988,2989..3780)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(3138..3157)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3951..4539
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4440..4459
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4693..4710
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4725..4864
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4790..4809)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4950..4972
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4966..5157)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(5966..6928,1..120))
                     /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"
ORIGIN
        1 cgcttccact ttttcccgcg ttttcgcaga aacgtggctg gcctggttca ccacgcggga
       61 aacggtctga taagagacac cggcatactc tgcgacatcg tataacgtta ctggtttcac
      121 attcaccacc ctgaattgac tctcttccgg gcgctatcat gccataccgc gaaaggtttt
      181 gcgccattcg atggtgtccg ggatctcgac gctctccctt atgcgactcc tgcattagga
      241 agcagcccag tagtaggttg aggccgttga gcaccgccgc cgcaaggaat ggtgcatgca
      301 aggagatggc gcccaacagt cccccggcca cggggcctgc caccataccc acgccgaaac
      361 aagcgctcat gagcccgaag tggcgagccc gatcttcccc atcggtgatg tcggcgatat
      421 aggcgccagc aaccgcacct gtggcgccgg tgatgccggc cacgatgcgt ccggcgtaga
      481 ggatcgagat ctcgatcccg cgaaattaat acgactcact ataggggaat tgtgagcgga
      541 taacaattcc cctctagtaa taattttgtt taactttaag aaggaggtct agaatggcat
      601 gcaaacgtca ttcagtgtcc gatcccattt cgaaggatct cgtcgagtgt cttcgtgaat
      661 caatgcaaag agatctcaaa tttgagactc cgtacgtttt cgtgatattt ggagcatcgg
      721 gagatctcgc aaaaaagaaa atatacccaa cactgtggtg gttgttccgt gacaaccttt
      781 tgccagtcaa catcaaattt attggatatg cccgttccga tctgacagtt ttcaagcttc
      841 gtgaatcttt tgagaaaaac tgtaaagttc gagaaaatga gaaatgtgct tttgatgact
      901 tcatcaagaa gtgctcatac gttcagggcc aatacgacac ttctgaagga ttccaacgcc
      961 ttcaatcttc aattgatgat ttccaaaaag aaagcaataa tcaagccgtc aatcgtctct
     1021 actacctcgc tcttcctcca tcagttttca acgtggtttc tacggaatta aagaagaatt
     1081 gtatggatca tggtgattca tggacacgtg ttatcataga aaaaccattt ggacatgatt
     1141 tgaaatcatc ttgtgaactt tcaactcatt tggcgaaact ttttaaagaa gatcaaatct
     1201 accgtattga tcattatctc ggaaaagaaa tggttcagaa tttgatggtt atgcgatttg
     1261 gaaaccgaat tctcgctcca tcttggaacc gcgatcacat tgcatcagtt atgatctcat
     1321 ttaaagaaga ttttggaact ggtggccttg ctggatattt tgatacagct ggaattatcc
     1381 gtgatgttat gcagaatcat ttgatgcaaa ttctaacttt agtcgccatg gagaaaccag
     1441 caagtttgaa tgctgaagat attcgtgatg aaaaggtcaa agtgttgaaa gctgctaaag
     1501 ttgtagaatt aaaggatgta gtagttggac aatacattgc cagtcctgaa tttgatcacc
     1561 cagaagcatc acaaggatac aaagatgaca aatcagttcc agccgattcg actactccaa
     1621 cgtatgctct tgctgttgtt cacatcaata atgaacgttg ggagggtgtt ccattcttct
     1681 tgcgttgtgg aaaagctctc aacgagaaga aagcagaagt tcgtatacaa ttcaaagaag
     1741 tcagtggaga tatctatccg tctggtgaat taaagcgatc cgaattagtg atgagagtgc
     1801 aaccaaatga ggcagtttac atgaaactga tgaccaagaa acctggaatg ggattcggtg
     1861 ttgaagaaac tgaacttgat ttgacttata ataatagatt caaagaagtc cgtcttcctg
     1921 atgcttatga acgcctattt ctcgaagtat tcatgggatc tcaaattaat ttcgttcgaa
     1981 ctgatgagct tgaatacgct tggcgtattt tgacgccagt tctcgaagag cttaaaaaga
     2041 agaaagttca accggttcaa tataaatttg gaagtcgtgg tccaactgaa ggtgacgaat
     2101 tgatgaaaaa gtacggtttc atcttcactg gaacatacaa atgggttgct ccaaagctac
     2161 tcgagcacca ccaccaccac cactgagatc cggctgctaa caaagcccga aaggaagctg
     2221 agttggctgc tgccaccgct gagcaataac tagcataacc ccttggggcc tctaaacggg
     2281 tcttgagggg ttttttgctg aaaggaggaa ctatatccgg attggcgaat gggacgcgcc
     2341 ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact
     2401 tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc
     2461 cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt
     2521 acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc
     2581 ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt
     2641 gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat
     2701 tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa
     2761 ttttaacaaa atattaacgc ttacaattta ggtggcactt ttcggggaaa tgtgcgcgga
     2821 acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa
     2881 ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt
     2941 gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg
     3001 ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg
     3061 gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg
     3121 agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag
     3181 caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca
     3241 gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg
     3301 agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc
     3361 gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg
     3421 aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgcagcaat ggcaacaacg
     3481 ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca attaatagac
     3541 tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg
     3601 tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg
     3661 gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact
     3721 atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa
     3781 ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca tttttaattt
     3841 aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag
     3901 ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct
     3961 ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt
     4021 tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg
     4081 cagataccaa atactgtcct tctagtgtag ccgtagttag gccaccactt caagaactct
     4141 gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc
     4201 gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg
     4261 tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa
     4321 ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg
     4381 gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg
     4441 ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga
     4501 tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt
     4561 ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct
     4621 gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga
     4681 acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcctgat gcggtatttt
     4741 ctccttacgc atctgtgcgg tatttcacac cgcaatggtg cactctcagt acaatctgct
     4801 ctgatgccgc atagttaagc cagtatacac tccgctatcg ctacgtgact gggtcatggc
     4861 tgcgccccga cacccgccaa cacccgctga cgcgccctga cgggcttgtc tgctcccggc
     4921 atccgcttac agacaagctg tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc
     4981 gtcatcaccg aaacgcgcga ggcagctgcg gtaaagctca tcagcgtggt cgtgaagcga
     5041 ttcacagatg tctgcctgtt catccgcgtc cagctcgttg agtttctcca gaagcgttaa
     5101 tgtctggctt ctgataaagc gggccatgtt aagggcggtt ttttcctgtt tggtcactga
     5161 tgcctccgtg taagggggat ttctgttcat gggggtaatg ataccgatga aacgagagag
     5221 gatgctcacg atacgggtta ctgatgatga acatgcccgg ttactggaac gttgtgaggg
     5281 taaacaactg gcggtatgga tgcggcggga ccagagaaaa atcactcagg gtcaatgcca
     5341 gcgcttcgtt aatacagatg taggtgttcc acagggtagc cagcagcatc ctgcgatgca
     5401 gatccggaac ataatggtgc agggcgctga cttccgcgtt tccagacttt acgaaacacg
     5461 gaaaccgaag accattcatg ttgttgctca ggtcgcagac gttttgcagc agcagtcgct
     5521 tcacgttcgc tcgcgtatcg gtgattcatt ctgctaacca gtaaggcaac cccgccagcc
     5581 tagccgggtc ctcaacgaca ggagcacgat catgcgcacc cgtggggccg ccatgccggc
     5641 gataatggcc tgcttctcgc cgaaacgttt ggtggcggga ccagtgacga aggcttgagc
     5701 gagggcgtgc aagattccga ataccgcaag cgacaggccg atcatcgtcg cgctccagcg
     5761 aaagcggtcc tcgccgaaaa tgacccagag cgctgccggc acctgtccta cgagttgcat
     5821 gataaagaag acagtcataa gtgcggcgac gatagtcatg ccccgcgccc accggaagga
     5881 gctgactggg ttgaaggctc tcaagggcat cggtcgagat cccggtgcct aatgagtgag
     5941 ctaacttaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg
     6001 ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgcca
     6061 gggtggtttt tcttttcacc agtgagacgg gcaacagctg attgcccttc accgcctggc
     6121 cctgagagag ttgcagcaag cggtccacgc tggtttgccc cagcaggcga aaatcctgtt
     6181 tgatggtggt taacggcggg atataacatg agctgtcttc ggtatcgtcg tatcccacta
     6241 ccgagatatc cgcaccaacg cgcagcccgg actcggtaat ggcgcgcatt gcgcccagcg
     6301 ccatctgatc gttggcaacc agcatcgcag tgggaacgat gccctcattc agcatttgca
     6361 tggtttgttg aaaaccggac atggcactcc agtcgccttc ccgttccgct atcggctgaa
     6421 tttgattgcg agtgagatat ttatgccagc cagccagacg cagacgcgcc gagacagaac
     6481 ttaatgggcc cgctaacagc gcgatttgct ggtgacccaa tgcgaccaga tgctccacgc
     6541 ccagtcgcgt accgtcttca tgggagaaaa taatactgtt gatgggtgtc tggtcagaga
     6601 catcaagaaa taacgccgga acattagtgc aggcagcttc cacagcaatg gcatcctggt
     6661 catccagcgg atagttaatg atcagcccac tgacgcgttg cgcgagaaga ttgtgcaccg
     6721 ccgctttaca ggcttcgacg ccgcttcgtt ctaccatcga caccaccacg ctggcaccca
     6781 gttgatcggc gcgagattta atcgccgcga caatttgcga cggcgcgtgc agggccagac
     6841 tggaggtggc aacgccaatc agcaacgact gtttgcccgc cagttgttgt gccacgcggt
     6901 tgggaatgta attcagctcc gccatcgc
//
