LOCUS       sequence-464188-        5912 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Heterologous protein expression of iron-sulfur cluster assembly 
            protein CyaY in Escherichia coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5912)
  TITLE     AI Proteins, Inc. plasmids
REFERENCE   2  (bases 1 to 5912)
  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"
COMMENT     Sequence Label: pAIP4_4HS5
FEATURES             Location/Qualifiers
     source          1..5912
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             301..492
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(486..508)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    594..733
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     649..668
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(748..765)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(919..1507)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(999..1018)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(1618..1637)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             1629..2444
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             1629..2444
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(1702..1721)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      complement(2536..2991)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2673..2694)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2885..2904
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     terminator      complement(3028..3075)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     3071..3089
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(3142..3159)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(3799..3828)
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     RBS             3851..3873
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    3888..3912
                     /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(3912..3931)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3913..3931)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4019..4038)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        4244..4321
                     /gene="lacI"
                     /label=lacI promoter
     CDS             4322..5404
                     /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(4341..4360)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 gtttcgtaaa gtctggaaac gcggaagtca gcgccctgca ccattatgtt ccggatctgc
       61 atcgcaggat gctgctggct accctgtgga acacctacat ctgtattaac gaagcgctgg
      121 cattgaccct gagtgatttt tctctggtcc cgccgcatcc ataccgccag ttgtttaccc
      181 tcacaacgtt ccagtaaccg ggcatgttca tcatcagtaa cccgtatcgt gagcatcctc
      241 tctcgtttca tcggtatcat tacccccatg aacagaaatc ccccttacac ggaggcatca
      301 gtgaccaaac aggaaaaaac cgcccttaac atggcccgct ttatcagaag ccagacatta
      361 acgcttctgg agaaactcaa cgagctggac gcggatgaac aggcagacat ctgtgaatcg
      421 cttcacgacc acgctgatga gctttaccgc agctgcctcg cgcgtttcgg tgatgacggt
      481 gaaaacctct gacacatgca gctcccggag acggtcacag cttgtctgta agcggatgcc
      541 gggagcagac aagcccgtca gggcgcgtca gcgggtgttg gcgggtgtcg gggcgcagcc
      601 atgacccagt cacgtagcga tagcggagtg tatactggct taactatgcg gcatcagagc
      661 agattgtact gagagtgcac cattgcggtg tgaaataccg cacagatgcg taaggagaaa
      721 ataccgcatc aggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg
      781 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
      841 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
      901 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
      961 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     1021 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     1081 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     1141 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     1201 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     1261 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     1321 gttcttgaag tggtggccta actacggcta cactagaagg acagtatttg gtatctgcgc
     1381 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     1441 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     1501 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     1561 acgttaaggg attttggtca tgaacaataa aactgtctgc ttacataaac agtaatacaa
     1621 ggggtgttat gagccatatt caacgggaaa cgtcttgctc taggccgcga ttaaattcca
     1681 acatggatgc tgatttatat gggtataaat gggctcgcga taatgtcggg caatcaggtg
     1741 cgacaatcta tcgattgtat gggaagcccg atgcgccaga gttgtttctg aaacatggca
     1801 aaggtagcgt tgccaatgat gttacagatg agatggtcag actaaactgg ctgacggaat
     1861 ttatgcctct tccgaccatc aagcatttta tccgtactcc tgatgatgca tggttactca
     1921 ccactgcgat ccccgggaaa acagcattcc aggtattaga agaatatcct gattcaggtg
     1981 aaaatattgt tgatgcgctg gcagtgttcc tgcgccggtt gcattcgatt cctgtttgta
     2041 attgtccttt taacagcgat cgcgtatttc gtctcgctca ggcgcaatca cgaatgaata
     2101 acggtttggt tgatgcgagt gattttgatg acgagcgtaa tggctggcct gttgaacaag
     2161 tctggaaaga aatgcataaa cttttgccat tctcaccgga ttcagtcgtc actcatggtg
     2221 atttctcact tgataacctt atttttgacg aggggaaatt aataggttgt attgatgttg
     2281 gacgagtcgg aatcgcagac cgataccagg atcttgccat cctatggaac tgcctcggtg
     2341 agttttctcc ttcattacag aaacggcttt ttcaaaaata tggtattgat aatcctgata
     2401 tgaataaatt gcagtttcat ttgatgctcg atgagttttt ctaagaatta attcatgagc
     2461 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc
     2521 cgaaaagtgc cacctaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt
     2581 gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa
     2641 aagaatagac cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa
     2701 agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac
     2761 gtgaaccatc accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga
     2821 accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa
     2881 aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc
     2941 tgcgcgtaac caccacaccc gccgcgctta atgcgccgct acagggcgcg tcccattcgc
     3001 caatccggat atagttcctc ctttcagcaa aaaacccctc aagacccgtt tagaggcccc
     3061 aaggggttat gctagttatt gctcagcggt ggcagcagcc aactcagctt cctttcgggc
     3121 tttgttagca gccggatctc agtggtggtg gtggtggtgc tcgagtcatt acgaaccttc
     3181 ggtgaagtcg acggtttctt tgctttgctt aaaaattgcc gcgctcagaa acgtcaagaa
     3241 ttcatcacca gaacgatcat caatccattt accgttattg tagtcatagt ggtgaccgtt
     3301 ctcgagggta gccagccaaa cctgatgcag cggctgctgg cggttgataa tcagcttggt
     3361 atgattctcg aactctaaag tcagcaagct gccgttttgg tcgtagtcca catccgcgcc
     3421 ggactcttcg attttttcct cgatcttctg atacagttgg tcagctagtt gtataaattc
     3481 tgaatcattc attccagagc cgccaatctg ttcacggtgg gcttcaatga tgtcgttgtc
     3541 ttccatgtcc aggtcttccg gcgcttgatc tgcctggatg cgaataccgt catacaggaa
     3601 acgcaggcta tccatttcct tgccctgacg tttggcaaac gcttccatca ggcgacgcag
     3661 cggcgtggtc tttttgatct taaagaaaat ttcagagctg ccgtcactaa ctttcaggtt
     3721 aatatgggtt tccggtttca cttccggctt gacttccggc tttgcttctt ggttgacttc
     3781 cgaatcttgc aggctaccgt ggtgatggtg atggtgatga tggtgatgaa tcttcgccat
     3841 catatgtata tctccttctt aaagttaaac aaaattattt ctagagggga attgttatcc
     3901 gctcacaatt cccctatagt gagtcgtatt aatttcgcgg gatcgagatc gatctcgatc
     3961 ctctacgccg gacgcatcgt ggccggcatc accggcgcca caggtgcggt tgctggcgcc
     4021 tatatcgccg acatcaccga tggggaagat cgggctcgcc acttcgggct catgagcgct
     4081 tgtttcggcg tgggtatggt ggcaggcccc gtggccgggg gactgttggg cgccatctcc
     4141 ttgcatgcac cattccttgc ggcggcggtg ctcaacggcc tcaacctact actgggctgc
     4201 ttcctaatgc aggagtcgca taagggagag cgtcgagatc ccggacacca tcgaatggcg
     4261 caaaaccttt cgcggtatgg catgatagcg cccggaagag agtcaattca gggtggtgaa
     4321 tgtgaaacca gtaacgttat acgatgtcgc agagtatgcc ggtgtctctt atcagaccgt
     4381 ttcccgcgtg gtgaaccagg ccagccacgt ttctgcgaaa acgcgggaaa aagtggaagc
     4441 ggcgatggcg gagctgaatt acattcccaa ccgcgtggca caacaactgg cgggcaaaca
     4501 gtcgttgctg attggcgttg ccacctccag tctggccctg cacgcgccgt cgcaaattgt
     4561 cgcggcgatt aaatctcgcg ccgatcaact gggtgccagc gtggtggtgt cgatggtaga
     4621 acgaagcggc gtcgaagcct gtaaagcggc ggtgcacaat cttctcgcgc aacgcgtcag
     4681 tgggctgatc attaactatc cgctggatga ccaggatgcc attgctgtgg aagctgcctg
     4741 cactaatgtt ccggcgttat ttcttgatgt ctctgaccag acacccatca acagtattat
     4801 tttctcccat gaagacggta cgcgactggg cgtggagcat ctggtcgcat tgggtcacca
     4861 gcaaatcgcg ctgttagcgg gcccattaag ttctgtctcg gcgcgtctgc gtctggctgg
     4921 ctggcataaa tatctcactc gcaatcaaat tcagccgata gcggaacggg aaggcgactg
     4981 gagtgccatg tccggttttc aacaaaccat gcaaatgctg aatgagggca tcgttcccac
     5041 tgcgatgctg gttgccaacg atcagatggc gctgggcgca atgcgcgcca ttaccgagtc
     5101 cgggctgcgc gttggtgcgg acatctcggt agtgggatac gacgataccg aagacagctc
     5161 atgttatatc ccgccgttaa ccaccatcaa acaggatttt cgcctgctgg ggcaaaccag
     5221 cgtggaccgc ttgctgcaac tctctcaggg ccaggcggtg aagggcaatc agctgttgcc
     5281 cgtctcactg gtgaaaagaa aaaccaccct ggcgcccaat acgcaaaccg cctctccccg
     5341 cgcgttggcc gattcattaa tgcagctggc acgacaggtt tcccgactgg aaagcgggca
     5401 gtgagcgcaa cgcaattaat gtaagttagc tcactcatta ggcaccggga tctcgaccga
     5461 tgcccttgag agccttcaac ccagtcagct ccttccggtg ggcgcggggc atgactatcg
     5521 tcgccgcact tatgactgtc ttctttatca tgcaactcgt aggacaggtg ccggcagcgc
     5581 tctgggtcat tttcggcgag gaccgctttc gctggagcgc gacgatgatc ggcctgtcgc
     5641 ttgcggtatt cggaatcttg cacgccctcg ctcaagcctt cgtcactggt cccgccacca
     5701 aacgtttcgg cgagaagcag gccattatcg ccggcatggc ggccccacgg gtgcgcatga
     5761 tcgtgctcct gtcgttgagg acccggctag gctggcgggg ttgccttact ggttagcaga
     5821 atgaatcacc gatacgcgag cgaacgtgaa gcgactgctg ctgcaaaacg tctgcgacct
     5881 gagcaacaac atgaatggtc ttcggtttcc gt
//
