LOCUS       sequence-268614-        6270 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of recombinant Arabidopsis thaliana 
            ferrochelatase1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6270)
  AUTHORS   Shimizu T, Kacprzak SM, Mochizuki N, Nagatani A, Watanabe S, Shimada
            T, Tanaka K, Hayashi Y, Arai M, Leister D, Okamoto H, Terry MJ, 
            Masuda T
  TITLE     The retrograde signaling protein GUN1 regulates tetrapyrrole 
            biosynthesis.
  JOURNAL   Proc Natl Acad Sci U S A. 2019 Dec 3;116(49):24900-24906. doi: 
            10.1073/pnas.1911251116. Epub 2019 Nov 15.
  PUBMED    31732672
REFERENCE   2  (bases 1 to 6270)
  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: 
            "10.1073/pnas.1911251116"; journalName: "Proc Natl Acad Sci U S A"; 
            date: "2019-12-3- 3"; volume: "116"; issue: "49"; pages: 
            "24900-24906"
COMMENT     Sequence Label: pGEX2T AtFC1
FEATURES             Location/Qualifiers
     source          1..6270
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(1..6,7..22,23..29)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    37..53
                     /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             76..729
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     687..709
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             736..753
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     RBS             1500..1508
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     primer_bind     complement(2138..2160)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2305..2323)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2391..2495
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2496..2564,2565..3356)
                     /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(2714..2733)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3527..4115
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4016..4035
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4269..4286
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        4359..4436
                     /gene="lacI"
                     /label=lacI promoter
     CDS             4437..5519
                     /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(4456..4475)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(5568..5573,5574..5591,5592..5598)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5606..5622
                     /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     5630..5646
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5630..5646
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             5642..6169
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(5662..5679)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5662..5678)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5671..5693)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 ttgacaatta atcatcggct cgtataatgt gtggaattgt gagcggataa caatttcaca
       61 caggaaacag tattcatgtc ccctatacta ggttattgga aaattaaggg ccttgtgcaa
      121 cccactcgac ttcttttgga atatcttgaa gaaaaatatg aagagcattt gtatgagcgc
      181 gatgaaggtg ataaatggcg aaacaaaaag tttgaattgg gtttggagtt tcccaatctt
      241 ccttattata ttgatggtga tgttaaatta acacagtcta tggccatcat acgttatata
      301 gctgacaagc acaacatgtt gggtggttgt ccaaaagagc gtgcagagat ttcaatgctt
      361 gaaggagcgg ttttggatat tagatacggt gtttcgagaa ttgcatatag taaagacttt
      421 gaaactctca aagttgattt tcttagcaag ctacctgaaa tgctgaaaat gttcgaagat
      481 cgtttatgtc ataaaacata tttaaatggt gatcatgtaa cccatcctga cttcatgttg
      541 tatgacgctc ttgatgttgt tttatacatg gacccaatgt gcctggatgc gttcccaaaa
      601 ttagtttgtt ttaaaaaacg tattgaagct atcccacaaa ttgataagta cttgaaatcc
      661 agcaagtata tagcatggcc tttgcagggc tggcaagcca cgtttggtgg tggcgaccat
      721 cctccaaaat cggatctggt tccgcgtgga tcctctcaga gaaattctct gtctttgatt
      781 caatgcgata taaaagagag atctttcgga gagtctatga cgatcacgaa tcgtggattg
      841 agttttaaga cgaatgtgtt tgagcaagct cgttctgtga ctggagactg ttcttatgat
      901 gaaacttcag ctaaagcacg ttctcatgtt gttgcagaag ataagattgg tgtcttgctt
      961 ttgaatttag gtggtcctga aactcttaac gatgttcaac ctttcttgta taatctcttt
     1021 gctgatccgg atattataag gcttcctaga ccatttcagt ttcttcaagg gactatagct
     1081 aagtttatat ctgttgttcg tgctccgaaa tctaaagaag ggtatgctgc tattggtggt
     1141 ggctctcctt tgcgtaaaat aactgatgag caagcggatg ctattaagat gtctttgcaa
     1201 gcgaagaaca ttgctgctaa tgtctatgtt ggtatgcggt attggtatcc gttcactgag
     1261 gaggctgttc agcagataaa gaaggacaaa attactagac ttgttgtact gccattgtat
     1321 cctcagtatt ctatctctac aacgggttca agcatacgcg ttctccaaga tttattcagg
     1381 aaagatccgt acctagctgg agtgccggta gctattataa agtcctggta ccaaaggcga
     1441 ggctatgtca attctatggc tgacctcatt gagaaggagc ttcaaacttt ctctgatcct
     1501 aaggaggtta tgatattctt cagtgcccat ggtgttccgg tcagctacgt agagaatgct
     1561 ggagatccgt accagaagca gatggaagag tgtattgact tgataatgga agagctaaaa
     1621 gccagagggg ttcttaacga ccataaattg gcataccaga gtcgtgttgg ccctgttcaa
     1681 tggctgaagc catacaccga tgaggttctt gtcgaccttg gtaagagtgg tgttaagagt
     1741 ctactagccg ttccagtcag tttcgtgagt gagcacattg agacacttga ggagatagac
     1801 atggagtata gggaattagc tcttgagtca ggggtagaga actggggacg ggtacccgcg
     1861 ctaggtctca caccatcctt catcaccgac ttagctgatg cagtgataga atcacttcct
     1921 tcagcagaag caatgtcaaa cccaaatgca gtggttgact cagaagatag cgagtcgtca
     1981 gatgctttca gttacattgt caagatgttc ttcggttcga ttctggcttt cgtcctactt
     2041 ctctccccaa agatgttcca tgcgttccgg aacctatagg aattcatcgt gactgactga
     2101 cgatctgcct cgcgcgtttc ggtgatgacg gtgaaaacct ctgacacatg cagctcccgg
     2161 agacggtcac agcttgtctg taagcggatg ccgggagcag acaagcccgt cagggcgcgt
     2221 cagcgggtgt tggcgggtgt cggggcgcag ccatgaccca gtcacgtagc gatagcggag
     2281 tgtataattc ttgaagacga aagggcctcg tgatacgcct atttttatag gttaatgtca
     2341 tgataataat ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc
     2401 ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct
     2461 gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg
     2521 cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg
     2581 tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc
     2641 tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca
     2701 cttttaaagt tctgctatgt ggcgcggtat tatcccgtgt tgacgccggg caagagcaac
     2761 tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa
     2821 agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg
     2881 ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt
     2941 ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg
     3001 aagccatacc aaacgacgag cgtgacacca cgatgcctgc agcaatggca acaacgttgc
     3061 gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga
     3121 tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta
     3181 ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc
     3241 cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg
     3301 atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt
     3361 cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa
     3421 ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt
     3481 cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt
     3541 ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt
     3601 tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga
     3661 taccaaatac tgtccttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag
     3721 caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata
     3781 agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg
     3841 gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga
     3901 gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca
     3961 ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa
     4021 acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt
     4081 tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac
     4141 ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt
     4201 ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga
     4261 ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cctgatgcgg tattttctcc
     4321 ttacgcatct gtgcggtatt tcacaccgca taaattccga caccatcgaa tggcgcaaaa
     4381 cctttcgcgg tatggcatga tagcgcccgg aagagagtca attcagggtg gtgaatgtga
     4441 aaccagtaac gttatacgat gtcgcagagt atgccggtgt ctcttatcag accgtttccc
     4501 gcgtggtgaa ccaggccagc cacgtttctg cgaaaacgcg ggaaaaagtg gaagcggcga
     4561 tggcggagct gaattacatt cccaaccgcg tggcacaaca actggcgggc aaacagtcgt
     4621 tgctgattgg cgttgccacc tccagtctgg ccctgcacgc gccgtcgcaa attgtcgcgg
     4681 cgattaaatc tcgcgccgat caactgggtg ccagcgtggt ggtgtcgatg gtagaacgaa
     4741 gcggcgtcga agcctgtaaa gcggcggtgc acaatcttct cgcgcaacgc gtcagtgggc
     4801 tgatcattaa ctatccgctg gatgaccagg atgccattgc tgtggaagct gcctgcacta
     4861 atgttccggc gttatttctt gatgtctctg accagacacc catcaacagt attattttct
     4921 cccatgaaga cggtacgcga ctgggcgtgg agcatctggt cgcattgggt caccagcaaa
     4981 tcgcgctgtt agcgggccca ttaagttctg tctcggcgcg tctgcgtctg gctggctggc
     5041 ataaatatct cactcgcaat caaattcagc cgatagcgga acgggaaggc gactggagtg
     5101 ccatgtccgg ttttcaacaa accatgcaaa tgctgaatga gggcatcgtt cccactgcga
     5161 tgctggttgc caacgatcag atggcgctgg gcgcaatgcg cgccattacc gagtccgggc
     5221 tgcgcgttgg tgcggatatc tcggtagtgg gatacgacga taccgaagac agctcatgtt
     5281 atatcccgcc gttaaccacc atcaaacagg attttcgcct gctggggcaa accagcgtgg
     5341 accgcttgct gcaactctct cagggccagg cggtgaaggg caatcagctg ttgcccgtct
     5401 cactggtgaa aagaaaaacc accctggcgc ccaatacgca aaccgcctct ccccgcgcgt
     5461 tggccgattc attaatgcag ctggcacgac aggtttcccg actggaaagc gggcagtgag
     5521 cgcaacgcaa ttaatgtaag ttagctcact cattaggcac cccaggcttt acactttatg
     5581 cttccggctc gtatgttgtg tggaattgtg agcggataac aatttcacac aggaaacagc
     5641 tatgaccatg attacggatt cactggccgt cgttttacaa cgtcgtgact gggaaaaccc
     5701 tggcgttacc caacttaatc gccttgcagc acatccccct ttcgccagct ggcgtaatag
     5761 cgaagaggcc cgcaccgatc gcccttccca acagttgcgc agcctgaatg gcgaatggcg
     5821 ctttgcctgg tttccggcac cagaagcggt gccggaaagc tggctggagt gcgatcttcc
     5881 tgaggccgat actgtcgtcg tcccctcaaa ctggcagatg cacggttacg atgcgcccat
     5941 ctacaccaac gtgacctatc ccattacggt caatccgccg tttgttccca cggagaatcc
     6001 gacgggttgt tactcgctca catttaatgt tgatgaaagc tggctacagg aaggccagac
     6061 gcgaattatt tttgatggcg ttggaattag cttatcgact gcacggtgca ccaatgcttc
     6121 tggcgtcagg cagccatcgg aagctgtggt atggctgtgc aggtcgtaaa tcactgcata
     6181 attcgtgtcg ctcaaggcgc actcccgttc tggataatgt tttttgcgcc gacatcataa
     6241 cggttctggc aaatattctg aaatgagctg
//
