LOCUS       sequence-464145-        5915 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 5915)
  TITLE     AI Proteins, Inc. plasmids
REFERENCE   2  (bases 1 to 5915)
  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_1EW4
FEATURES             Location/Qualifiers
     source          1..5915
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(55..102)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     98..116
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(169..186)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(829..858)
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     RBS             881..903
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    918..942
                     /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(942..961)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(943..961)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1049..1068)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        1274..1351
                     /gene="lacI"
                     /label=lacI promoter
     CDS             1352..2434
                     /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(1371..1390)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             3243..3434
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(3428..3450)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3536..3675
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3591..3610
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3690..3707)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3861..4449)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3941..3960)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(4560..4579)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             4571..5386
                     /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             4571..5386
                     /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(4644..4663)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      complement(join(5478..5915,1..18))
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5615..5636)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5827..5846
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 cgccgctaca gggcgcgtcc cattcgccaa tccggatata gttcctcctt tcagcaaaaa
       61 acccctcaag acccgtttag aggccccaag gggttatgct agttattgct cagcggtggc
      121 agcagccaac tcagcttcct ttcgggcttt gttagcagcc ggatctcagt ggtggtggtg
      181 gtggtgctcg agtcattacg aaccacgaaa gctgacggtt tcaccagctt gctgggttgc
      241 cgcctgttcc agcaggtccc agaaagtctc accagaacga tcgcaaatcc actcgtcacc
      301 ctttaaatca aaatggtagc caccttgttt ggtcgccaac caaacctggt gcagcggctc
      361 ttgacgatta atgatgattt tggaaccgtt ctcgaaggtg atggtgagca cgccgccgtt
      421 aatttcacag tcaatatcgc tgtcgccgtc ccaatcatcc aggcgctctt cgatcgtcaa
      481 ccacagctgg tcagctagcc tgtgaaattc tgaatcattc attccagagc cgccaatctg
      541 ttcacggtgg gcttcaatga tgtcgttgtc ttccatgtcc aggtcttccg gcgcttgatc
      601 tgcctggatg cgaataccgt catacaggaa acgcaggcta tccatttcct tgccctgacg
      661 tttggcaaac gcttccatca ggcgacgcag cggcgtggtc tttttgatct taaagaaaat
      721 ttcagagctg ccgtcactaa ctttcaggtt aatatgggtt tccggtttca cttccggctt
      781 gacttccggc tttgcttctt ggttgacttc cgaatcttgc aggctaccgt ggtgatggtg
      841 atggtgatga tggtgatgaa tcttcgccat catatgtata tctccttctt aaagttaaac
      901 aaaattattt ctagagggga attgttatcc gctcacaatt cccctatagt gagtcgtatt
      961 aatttcgcgg gatcgagatc gatctcgatc ctctacgccg gacgcatcgt ggccggcatc
     1021 accggcgcca caggtgcggt tgctggcgcc tatatcgccg acatcaccga tggggaagat
     1081 cgggctcgcc acttcgggct catgagcgct tgtttcggcg tgggtatggt ggcaggcccc
     1141 gtggccgggg gactgttggg cgccatctcc ttgcatgcac cattccttgc ggcggcggtg
     1201 ctcaacggcc tcaacctact actgggctgc ttcctaatgc aggagtcgca taagggagag
     1261 cgtcgagatc ccggacacca tcgaatggcg caaaaccttt cgcggtatgg catgatagcg
     1321 cccggaagag agtcaattca gggtggtgaa tgtgaaacca gtaacgttat acgatgtcgc
     1381 agagtatgcc ggtgtctctt atcagaccgt ttcccgcgtg gtgaaccagg ccagccacgt
     1441 ttctgcgaaa acgcgggaaa aagtggaagc ggcgatggcg gagctgaatt acattcccaa
     1501 ccgcgtggca caacaactgg cgggcaaaca gtcgttgctg attggcgttg ccacctccag
     1561 tctggccctg cacgcgccgt cgcaaattgt cgcggcgatt aaatctcgcg ccgatcaact
     1621 gggtgccagc gtggtggtgt cgatggtaga acgaagcggc gtcgaagcct gtaaagcggc
     1681 ggtgcacaat cttctcgcgc aacgcgtcag tgggctgatc attaactatc cgctggatga
     1741 ccaggatgcc attgctgtgg aagctgcctg cactaatgtt ccggcgttat ttcttgatgt
     1801 ctctgaccag acacccatca acagtattat tttctcccat gaagacggta cgcgactggg
     1861 cgtggagcat ctggtcgcat tgggtcacca gcaaatcgcg ctgttagcgg gcccattaag
     1921 ttctgtctcg gcgcgtctgc gtctggctgg ctggcataaa tatctcactc gcaatcaaat
     1981 tcagccgata gcggaacggg aaggcgactg gagtgccatg tccggttttc aacaaaccat
     2041 gcaaatgctg aatgagggca tcgttcccac tgcgatgctg gttgccaacg atcagatggc
     2101 gctgggcgca atgcgcgcca ttaccgagtc cgggctgcgc gttggtgcgg acatctcggt
     2161 agtgggatac gacgataccg aagacagctc atgttatatc ccgccgttaa ccaccatcaa
     2221 acaggatttt cgcctgctgg ggcaaaccag cgtggaccgc ttgctgcaac tctctcaggg
     2281 ccaggcggtg aagggcaatc agctgttgcc cgtctcactg gtgaaaagaa aaaccaccct
     2341 ggcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
     2401 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtaagttagc
     2461 tcactcatta ggcaccggga tctcgaccga tgcccttgag agccttcaac ccagtcagct
     2521 ccttccggtg ggcgcggggc atgactatcg tcgccgcact tatgactgtc ttctttatca
     2581 tgcaactcgt aggacaggtg ccggcagcgc tctgggtcat tttcggcgag gaccgctttc
     2641 gctggagcgc gacgatgatc ggcctgtcgc ttgcggtatt cggaatcttg cacgccctcg
     2701 ctcaagcctt cgtcactggt cccgccacca aacgtttcgg cgagaagcag gccattatcg
     2761 ccggcatggc ggccccacgg gtgcgcatga tcgtgctcct gtcgttgagg acccggctag
     2821 gctggcgggg ttgccttact ggttagcaga atgaatcacc gatacgcgag cgaacgtgaa
     2881 gcgactgctg ctgcaaaacg tctgcgacct gagcaacaac atgaatggtc ttcggtttcc
     2941 gtgtttcgta aagtctggaa acgcggaagt cagcgccctg caccattatg ttccggatct
     3001 gcatcgcagg atgctgctgg ctaccctgtg gaacacctac atctgtatta acgaagcgct
     3061 ggcattgacc ctgagtgatt tttctctggt cccgccgcat ccataccgcc agttgtttac
     3121 cctcacaacg ttccagtaac cgggcatgtt catcatcagt aacccgtatc gtgagcatcc
     3181 tctctcgttt catcggtatc attaccccca tgaacagaaa tcccccttac acggaggcat
     3241 cagtgaccaa acaggaaaaa accgccctta acatggcccg ctttatcaga agccagacat
     3301 taacgcttct ggagaaactc aacgagctgg acgcggatga acaggcagac atctgtgaat
     3361 cgcttcacga ccacgctgat gagctttacc gcagctgcct cgcgcgtttc ggtgatgacg
     3421 gtgaaaacct ctgacacatg cagctcccgg agacggtcac agcttgtctg taagcggatg
     3481 ccgggagcag acaagcccgt cagggcgcgt cagcgggtgt tggcgggtgt cggggcgcag
     3541 ccatgaccca gtcacgtagc gatagcggag tgtatactgg cttaactatg cggcatcaga
     3601 gcagattgta ctgagagtgc accattgcgg tgtgaaatac cgcacagatg cgtaaggaga
     3661 aaataccgca tcaggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt
     3721 cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca
     3781 ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa
     3841 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat
     3901 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc
     3961 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc
     4021 gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt
     4081 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac
     4141 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg
     4201 ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca
     4261 gagttcttga agtggtggcc taactacggc tacactagaa ggacagtatt tggtatctgc
     4321 gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa
     4381 accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa
     4441 ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac
     4501 tcacgttaag ggattttggt catgaacaat aaaactgtct gcttacataa acagtaatac
     4561 aaggggtgtt atgagccata ttcaacggga aacgtcttgc tctaggccgc gattaaattc
     4621 caacatggat gctgatttat atgggtataa atgggctcgc gataatgtcg ggcaatcagg
     4681 tgcgacaatc tatcgattgt atgggaagcc cgatgcgcca gagttgtttc tgaaacatgg
     4741 caaaggtagc gttgccaatg atgttacaga tgagatggtc agactaaact ggctgacgga
     4801 atttatgcct cttccgacca tcaagcattt tatccgtact cctgatgatg catggttact
     4861 caccactgcg atccccggga aaacagcatt ccaggtatta gaagaatatc ctgattcagg
     4921 tgaaaatatt gttgatgcgc tggcagtgtt cctgcgccgg ttgcattcga ttcctgtttg
     4981 taattgtcct tttaacagcg atcgcgtatt tcgtctcgct caggcgcaat cacgaatgaa
     5041 taacggtttg gttgatgcga gtgattttga tgacgagcgt aatggctggc ctgttgaaca
     5101 agtctggaaa gaaatgcata aacttttgcc attctcaccg gattcagtcg tcactcatgg
     5161 tgatttctca cttgataacc ttatttttga cgaggggaaa ttaataggtt gtattgatgt
     5221 tggacgagtc ggaatcgcag accgatacca ggatcttgcc atcctatgga actgcctcgg
     5281 tgagttttct ccttcattac agaaacggct ttttcaaaaa tatggtattg ataatcctga
     5341 tatgaataaa ttgcagtttc atttgatgct cgatgagttt ttctaagaat taattcatga
     5401 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc
     5461 cccgaaaagt gccacctaaa ttgtaagcgt taatattttg ttaaaattcg cgttaaattt
     5521 ttgttaaatc agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc
     5581 aaaagaatag accgagatag ggttgagtgt tgttccagtt tggaacaaga gtccactatt
     5641 aaagaacgtg gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg atggcccact
     5701 acgtgaacca tcaccctaat caagtttttt ggggtcgagg tgccgtaaag cactaaatcg
     5761 gaaccctaaa gggagccccc gatttagagc ttgacgggga aagccggcga acgtggcgag
     5821 aaaggaaggg aagaaagcga aaggagcggg cgctagggcg ctggcaagtg tagcggtcac
     5881 gctgcgcgta accaccacac ccgccgcgct taatg
//
