LOCUS       sequence-444357-        6928 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  IPTG-inducible expression of C. elegans protein GSPD-1(D60N) 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_D60N
FEATURES             Location/Qualifiers
     source          1..6928
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     RBS             1365..1385
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    1401..1425
                     /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(1425..1444)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1426..1444)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1528..1547)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        1753..1830
                     /gene="lacI"
                     /label=lacI promoter
     CDS             1831..2913
                     /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(1850..1869)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             3722..3913
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(3907..3929)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4015..4154
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4070..4089
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4169..4186)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4340..4928)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4420..4439)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5099..5890,5891..5959))
                     /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     5722..5741
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5960..6064)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(6090..6545)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6227..6248)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6439..6458
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     terminator      complement(6582..6629)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     6625..6643
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(6696..6713)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
ORIGIN
        1 atacttcgag aaataggcgt tcataagcat caggaagacg gacttctttg aatctattat
       61 tataagtcaa atcaagttca gtttcttcaa caccgaatcc cattccaggt ttcttggtca
      121 tcagtttcat gtaaactgcc tcatttggtt gcactctcat cactaattcg gatcgcttta
      181 attcaccaga cggatagata tctccactga cttctttgaa ttgtatacga acttctgctt
      241 tcttctcgtt gagagctttt ccacaacgca agaagaatgg aacaccctcc caacgttcat
      301 tattgatgtg aacaacagca agagcatacg ttggagtagt cgaatcggct ggaactgatt
      361 tgtcatcttt gtatccttgt gatgcttctg ggtgatcaaa ttcaggactg gcaatgtatt
      421 gtccaactac tacatccttt aattctacaa ctttagcagc tttcaacact ttgacctttt
      481 catcacgaat atcttcagca ttcaaacttg ctggtttctc catggcgact aaagttagaa
      541 tttgcatcaa atgattctgc ataacatcac ggataattcc agctgtatca aaatatccag
      601 cacggccacc agttccaaaa tcttctttaa atgagatcat aactgatgca atgtgatcgc
      661 ggttccaaga tggagcgaga attcggtttc caaatcgcat aaccatcaaa ttctgaacca
      721 tttcttttcc gagataatga tcaatacggt agatttgatc ttctttaaaa agtttcgcca
      781 aatgagttga aagttcacaa gatgatttca aatcatgtcc aaatggtttt tctatgataa
      841 cacgtgtcca tgaatcacca tgatccatac aattcttctt taattccgta gaaaccacgt
      901 tgaaaactga tggaggaaga gcgaggtagt agagacgatt gacggcttga ttattgcttt
      961 ctttttggaa atcatcaatt gaagattgaa ggcgttggaa tccttcagaa gtgtcgtatt
     1021 ggccctgaac gtatgagcac ttcttgatga agtcatcaaa agcacatttc tcattttctc
     1081 gaactttaca gtttttctca aaagattcac gaagcttgaa aactgtcaga tcggaacggg
     1141 catatccaat aaatttgatg ttgactggca aaaggttatt acggaacaac caccacagtg
     1201 ttgggtatat tttctttttt gcgagatctc ccgatgctcc aaatatcacg aaaacgtacg
     1261 gagtctcaaa tttgagatct ctttgcattg attcacgaag acactcgacg agatccttcg
     1321 aaatgggatc ggacactgaa tgacgtttgc atgccattct agacctcctt cttaaagtta
     1381 aacaaaatta ttactagagg ggaattgtta tccgctcaca attcccctat agtgagtcgt
     1441 attaatttcg cgggatcgag atctcgatcc tctacgccgg acgcatcgtg gccggcatca
     1501 ccggcgccac aggtgcggtt gctggcgcct atatcgccga catcaccgat ggggaagatc
     1561 gggctcgcca cttcgggctc atgagcgctt gtttcggcgt gggtatggtg gcaggccccg
     1621 tggccggggg actgttgggc gccatctcct tgcatgcacc attccttgcg gcggcggtgc
     1681 tcaacggcct caacctacta ctgggctgct tcctaatgca ggagtcgcat aagggagagc
     1741 gtcgagatcc cggacaccat cgaatggcgc aaaacctttc gcggtatggc atgatagcgc
     1801 ccggaagaga gtcaattcag ggtggtgaat gtgaaaccag taacgttata cgatgtcgca
     1861 gagtatgccg gtgtctctta tcagaccgtt tcccgcgtgg tgaaccaggc cagccacgtt
     1921 tctgcgaaaa cgcgggaaaa agtggaagcg gcgatggcgg agctgaatta cattcccaac
     1981 cgcgtggcac aacaactggc gggcaaacag tcgttgctga ttggcgttgc cacctccagt
     2041 ctggccctgc acgcgccgtc gcaaattgtc gcggcgatta aatctcgcgc cgatcaactg
     2101 ggtgccagcg tggtggtgtc gatggtagaa cgaagcggcg tcgaagcctg taaagcggcg
     2161 gtgcacaatc ttctcgcgca acgcgtcagt gggctgatca ttaactatcc gctggatgac
     2221 caggatgcca ttgctgtgga agctgcctgc actaatgttc cggcgttatt tcttgatgtc
     2281 tctgaccaga cacccatcaa cagtattatt ttctcccatg aagacggtac gcgactgggc
     2341 gtggagcatc tggtcgcatt gggtcaccag caaatcgcgc tgttagcggg cccattaagt
     2401 tctgtctcgg cgcgtctgcg tctggctggc tggcataaat atctcactcg caatcaaatt
     2461 cagccgatag cggaacggga aggcgactgg agtgccatgt ccggttttca acaaaccatg
     2521 caaatgctga atgagggcat cgttcccact gcgatgctgg ttgccaacga tcagatggcg
     2581 ctgggcgcaa tgcgcgccat taccgagtcc gggctgcgcg ttggtgcgga tatctcggta
     2641 gtgggatacg acgataccga agacagctca tgttatatcc cgccgttaac caccatcaaa
     2701 caggattttc gcctgctggg gcaaaccagc gtggaccgct tgctgcaact ctctcagggc
     2761 caggcggtga agggcaatca gctgttgccc gtctcactgg tgaaaagaaa aaccaccctg
     2821 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
     2881 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg taagttagct
     2941 cactcattag gcaccgggat ctcgaccgat gcccttgaga gccttcaacc cagtcagctc
     3001 cttccggtgg gcgcggggca tgactatcgt cgccgcactt atgactgtct tctttatcat
     3061 gcaactcgta ggacaggtgc cggcagcgct ctgggtcatt ttcggcgagg accgctttcg
     3121 ctggagcgcg acgatgatcg gcctgtcgct tgcggtattc ggaatcttgc acgccctcgc
     3181 tcaagccttc gtcactggtc ccgccaccaa acgtttcggc gagaagcagg ccattatcgc
     3241 cggcatggcg gccccacggg tgcgcatgat cgtgctcctg tcgttgagga cccggctagg
     3301 ctggcggggt tgccttactg gttagcagaa tgaatcaccg atacgcgagc gaacgtgaag
     3361 cgactgctgc tgcaaaacgt ctgcgacctg agcaacaaca tgaatggtct tcggtttccg
     3421 tgtttcgtaa agtctggaaa cgcggaagtc agcgccctgc accattatgt tccggatctg
     3481 catcgcagga tgctgctggc taccctgtgg aacacctaca tctgtattaa cgaagcgctg
     3541 gcattgaccc tgagtgattt ttctctggtc ccgccgcatc cataccgcca gttgtttacc
     3601 ctcacaacgt tccagtaacc gggcatgttc atcatcagta acccgtatcg tgagcatcct
     3661 ctctcgtttc atcggtatca ttacccccat gaacagaaat cccccttaca cggaggcatc
     3721 agtgaccaaa caggaaaaaa ccgcccttaa catggcccgc tttatcagaa gccagacatt
     3781 aacgcttctg gagaaactca acgagctgga cgcggatgaa caggcagaca tctgtgaatc
     3841 gcttcacgac cacgctgatg agctttaccg cagctgcctc gcgcgtttcg gtgatgacgg
     3901 tgaaaacctc tgacacatgc agctcccgga gacggtcaca gcttgtctgt aagcggatgc
     3961 cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc ggggcgcagc
     4021 catgacccag tcacgtagcg atagcggagt gtatactggc ttaactatgc ggcatcagag
     4081 cagattgtac tgagagtgca ccattgcggt gtgaaatacc gcacagatgc gtaaggagaa
     4141 aataccgcat caggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
     4201 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag
     4261 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
     4321 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
     4381 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     4441 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     4501 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
     4561 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     4621 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     4681 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     4741 agttcttgaa gtggtggcct aactacggct acactagaag gacagtattt ggtatctgcg
     4801 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     4861 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     4921 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     4981 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa
     5041 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
     5101 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
     5161 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
     5221 gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc
     5281 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt
     5341 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg
     5401 ttgttgccat tgctgcaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca
     5461 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg
     5521 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca
     5581 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg
     5641 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct
     5701 cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca
     5761 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca
     5821 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg
     5881 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac
     5941 ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt
     6001 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc
     6061 cgcgcacatt tccccgaaaa gtgccaccta aattgtaagc gttaatattt tgttaaaatt
     6121 cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat
     6181 cccttataaa tcaaaagaat agaccgagat agggttgagt gttgttccag tttggaacaa
     6241 gagtccacta ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg
     6301 cgatggccca ctacgtgaac catcacccta atcaagtttt ttggggtcga ggtgccgtaa
     6361 agcactaaat cggaacccta aagggagccc ccgatttaga gcttgacggg gaaagccggc
     6421 gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag
     6481 tgtagcggtc acgctgcgcg taaccaccac acccgccgcg cttaatgcgc cgctacaggg
     6541 cgcgtcccat tcgccaatcc ggatatagtt cctcctttca gcaaaaaacc cctcaagacc
     6601 cgtttagagg ccccaagggg ttatgctagt tattgctcag cggtggcagc agccaactca
     6661 gcttcctttc gggctttgtt agcagccgga tctcagtggt ggtggtggtg gtgctcgagt
     6721 agctttggag caacccattt gtatgttcca gtgaagatga aaccgtactt tttcatcaat
     6781 tcgtcacctt cagttggacc acgacttcca aatttatatt gaaccggttg aactttcttc
     6841 tttttaagct cttcgagaac tggcgtcaaa atacgccaag cgtattcaag ctcatcagtt
     6901 cgaacgaaat taatttgaga tcccatga
//
