LOCUS       sequence-434438-        6954 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Study the interaction of FeoB in P. aeruginosa, specifically 
            focusing on the role of the outer loop 4 (OL4) by examining the 
            variant lacking OL4 in its interaction with exogenous peptides..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6954)
  AUTHORS   Sharifian Gh M, Norouzi F, Sorci M, Zaidi TS, Pier GB, Achimovich A,
            Ongwae GM, Liang B, Ryan M, Lemke M, Belfort G, Gadjeva M, Gahlmann 
            A, Pires MM, Venter H, Harris TE, Laurie GW
  TITLE     Lacritin cleavage-potentiated targeting of iron - respiratory 
            reciprocity promotes bacterial death.
  JOURNAL   J Biol Chem. 2025 May;301(5):108455. doi: 10.1016/j.jbc.2025.108455.
            Epub 2025 Mar 26.
  PUBMED    40154612
REFERENCE   2  (bases 1 to 6954)
  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: "J Biol 
            Chem."; date: "2025-05"; volume: "301(5):108455. doi"; pages: " 
            10.1016/j.jbc.2025.108455. Epub 2025 Mar 26"
COMMENT     Sequence Label: pET41-FeoB-OL4del
FEATURES             Location/Qualifiers
     source          1..6954
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     RBS             795..817
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    832..856
                     /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(856..875)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(857..875)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(963..982)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        1188..1265
                     /gene="lacI"
                     /label=lacI promoter
     CDS             1266..2348
                     /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(1285..1304)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             2920..3111
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(3105..3127)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3213..3352
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3268..3287
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3367..3384)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3538..4126)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3618..3637)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(4237..4256)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             4248..5063
                     /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             4248..5063
                     /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(4321..4340)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      complement(5155..5610)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5292..5313)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5504..5523
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     terminator      complement(5647..5694)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     5690..5708
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(5771..5794)
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     CDS             complement(5819..5836)
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
ORIGIN
        1 gagcctgctg gtggtcgcag accgccgcac agatctcgcc gatcagccgg tagcgcgcat
       61 cgacgatcgc cagctcgggc tcctcgccgc agccccggcg cgcctgttcc agggtggcgg
      121 gcggcaagcc cagcgccggg ccgttgaaga tgtcgccttc cagcgcctgc aacgccagcc
      181 agcgcggctc gatggccgag gccgccggag cgcgcgtttc cagaaggtag ccgacctgcg
      241 cctggatggc cggtgggtaa tccaccgcca gcgccgcctg cgggagctgt agggagtcga
      301 tggccgcctt cagttcgtcg atgccatcgg cgcgggtgga taccagcggc accaccgggc
      361 agccgagacg ccgcgccaga ccgtcgatat cgatgcggat gcgctggcta cgggcgatgt
      421 cgagcatgtt cagcgcgacg atgcagggga tgcccatctc ccgcagttgc accgtgaggt
      481 acaggttgcg ctccaggttc gccgcatcca ccacgttgac cagcacgtcc acttcgccgc
      541 tggcgatgta gcggcaggcg atctgctcgt ccagcgaggc ctgggcggaa acgctggtca
      601 acgaataggt gccgggcagg tcgaccaggc gcaccgcgtg acggacggta tggaacgcac
      661 cctccttgcg ctccacggtg acgccggccc agttgccgac ccgctggcgc gagccggtga
      721 gctggttgaa cagggtggtc ttgccggagt tgggattgcc gatcaggccg agggtcaatg
      781 cggtcatatg tatatctcct tcttaaagtt aaacaaaatt atttctagag gggaattgtt
      841 atccgctcac aattccccta tagtgagtcg tattaatttc gcgggatcga gatcgatctc
      901 gatcctctac gccggacgca tcgtggccgg catcaccggc gccacaggtg cggttgctgg
      961 cgcctatatc gccgacatca ccgatgggga agatcgggct cgccacttcg ggctcatgag
     1021 cgcttgtttc ggcgtgggta tggtggcagg ccccgtggcc gggggactgt tgggcgccat
     1081 ctccttgcat gcaccattcc ttgcggcggc ggtgctcaac ggcctcaacc tactactggg
     1141 ctgcttccta atgcaggagt cgcataaggg agagcgtcga gatcccggac accatcgaat
     1201 ggcgcaaaac ctttcgcggt atggcatgat agcgcccgga agagagtcaa ttcagggtgg
     1261 tgaatgtgaa accagtaacg ttatacgatg tcgcagagta tgccggtgtc tcttatcaga
     1321 ccgtttcccg cgtggtgaac caggccagcc acgtttctgc gaaaacgcgg gaaaaagtgg
     1381 aagcggcgat ggcggagctg aattacattc ccaaccgcgt ggcacaacaa ctggcgggca
     1441 aacagtcgtt gctgattggc gttgccacct ccagtctggc cctgcacgcg ccgtcgcaaa
     1501 ttgtcgcggc gattaaatct cgcgccgatc aactgggtgc cagcgtggtg gtgtcgatgg
     1561 tagaacgaag cggcgtcgaa gcctgtaaag cggcggtgca caatcttctc gcgcaacgcg
     1621 tcagtgggct gatcattaac tatccgctgg atgaccagga tgccattgct gtggaagctg
     1681 cctgcactaa tgttccggcg ttatttcttg atgtctctga ccagacaccc atcaacagta
     1741 ttattttctc ccatgaagac ggtacgcgac tgggcgtgga gcatctggtc gcattgggtc
     1801 accagcaaat cgcgctgtta gcgggcccat taagttctgt ctcggcgcgt ctgcgtctgg
     1861 ctggctggca taaatatctc actcgcaatc aaattcagcc gatagcggaa cgggaaggcg
     1921 actggagtgc catgtccggt tttcaacaaa ccatgcaaat gctgaatgag ggcatcgttc
     1981 ccactgcgat gctggttgcc aacgatcaga tggcgctggg cgcaatgcgc gccattaccg
     2041 agtccgggct gcgcgttggt gcggacatct cggtagtggg atacgacgat accgaagaca
     2101 gctcatgtta tatcccgccg ttaaccacca tcaaacagga ttttcgcctg ctggggcaaa
     2161 ccagcgtgga ccgcttgctg caactctctc agggccaggc ggtgaagggc aatcagctgt
     2221 tgcccgtctc actggtgaaa agaaaaacca ccctggcgcc caatacgcaa accgcctctc
     2281 cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca ggtttcccga ctggaaagcg
     2341 ggcagtgagc gcaacgcaat taatgtaagt tagctcactc attaggcacc gggatctcga
     2401 ccgatgccct tgagagcctt caacccagtc agctccttcc ggtgggcgcg gggcatgact
     2461 agcatgatcg tgctcctgtc gttgaggacc cggctaggct ggcggggttg ccttactggt
     2521 tagcagaatg aatcaccgat acgcgagcga acgtgaagcg actgctgctg caaaacgtct
     2581 gcgacctgag caacaacatg aatggtcttc ggtttccgtg tttcgtaaag tctggaaacg
     2641 cggaagtcag cgccctgcac cattatgttc cggatctgca tcgcaggatg ctgctggcta
     2701 ccctgtggaa cacctacatc tgtattaacg aagcgctggc attgaccctg agtgattttt
     2761 ctctggtccc gccgcatcca taccgccagt tgtttaccct cacaacgttc cagtaaccgg
     2821 gcatgttcat catcagtaac ccgtatcgtg agcatcctct ctcgtttcat cggtatcatt
     2881 acccccatga acagaaatcc cccttacacg gaggcatcag tgaccaaaca ggaaaaaacc
     2941 gcccttaaca tggcccgctt tatcagaagc cagacattaa cgcttctgga gaaactcaac
     3001 gagctggacg cggatgaaca ggcagacatc tgtgaatcgc ttcacgacca cgctgatgag
     3061 ctttaccgca gctgcctcgc gcgtttcggt gatgacggtg aaaacctctg acacatgcag
     3121 ctcccggaga cggtcacagc ttgtctgtaa gcggatgccg ggagcagaca agcccgtcag
     3181 ggcgcgtcag cgggtgttgg cgggtgtcgg ggcgcagcca tgacccagtc acgtagcgat
     3241 agcggagtgt atactggctt aactatgcgg catcagagca gattgtactg agagtgcacc
     3301 attgcggtgt gaaataccgc acagatgcgt aaggagaaaa taccgcatca ggcgctcttc
     3361 cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc
     3421 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat
     3481 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
     3541 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
     3601 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     3661 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     3721 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     3781 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     3841 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     3901 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     3961 ctacggctac actagaagga cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     4021 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     4081 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     4141 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     4201 gaacaataaa actgtctgct tacataaaca gtaatacaag gggtgttatg agccatattc
     4261 aacgggaaac gtcttgctct aggccgcgat taaattccaa catggatgct gatttatatg
     4321 ggtataaatg ggctcgcgat aatgtcgggc aatcaggtgc gacaatctat cgattgtatg
     4381 ggaagcccga tgcgccagag ttgtttctga aacatggcaa aggtagcgtt gccaatgatg
     4441 ttacagatga gatggtcaga ctaaactggc tgacggaatt tatgcctctt ccgaccatca
     4501 agcattttat ccgtactcct gatgatgcat ggttactcac cactgcgatc cccgggaaaa
     4561 cagcattcca ggtattagaa gaatatcctg attcaggtga aaatattgtt gatgcgctgg
     4621 cagtgttcct gcgccggttg cattcgattc ctgtttgtaa ttgtcctttt aacagcgatc
     4681 gcgtatttcg tctcgctcag gcgcaatcac gaatgaataa cggtttggtt gatgcgagtg
     4741 attttgatga cgagcgtaat ggctggcctg ttgaacaagt ctggaaagaa atgcataaac
     4801 ttttgccatt ctcaccggat tcagtcgtca ctcatggtga tttctcactt gataacctta
     4861 tttttgacga ggggaaatta ataggttgta ttgatgttgg acgagtcgga atcgcagacc
     4921 gataccagga tcttgccatc ctatggaact gcctcggtga gttttctcct tcattacaga
     4981 aacggctttt tcaaaaatat ggtattgata atcctgatat gaataaattg cagtttcatt
     5041 tgatgctcga tgagtttttc taagaattaa ttcatgagcg gatacatatt tgaatgtatt
     5101 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctaaattg
     5161 taagcgttaa tattttgtta aaattcgcgt taaatttttg ttaaatcagc tcatttttta
     5221 accaataggc cgaaatcggc aaaatccctt ataaatcaaa agaatagacc gagatagggt
     5281 tgagtgttgt tccagtttgg aacaagagtc cactattaaa gaacgtggac tccaacgtca
     5341 aagggcgaaa aaccgtctat cagggcgatg gcccactacg tgaaccatca ccctaatcaa
     5401 gttttttggg gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg agcccccgat
     5461 ttagagcttg acggggaaag ccggcgaacg tggcgagaaa ggaagggaag aaagcgaaag
     5521 gagcgggcgc tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc accacacccg
     5581 ccgcgcttaa tgcgccgcta cagggcgcgt cccattcgcc aatccggata tagttcctcc
     5641 tttcagcaaa aaacccctca agacccgttt agaggcccca aggggttatg ctagttattg
     5701 ctcagcggtg gcagcagcca actcagcttc ctttcgggct ttgtttagca gcctaggtat
     5761 taatcaatta gtggtggtgg tggtggtggt ggtgctcgag tgcggccgca agcttcccac
     5821 taccgcgtgg caccaggtga catcctccct gcccgctcgg ggcatcggcc attcggcctg
     5881 gcagttgcag tacctgctca cggccgaaca gacgcaggca ggtcatcagg atcaggttga
     5941 agagcagcac cacggcgatg gtcaggacgc tacgctcggg gtgcgcggcg aaggtggcga
     6001 cctggtagca gagggtcgcc agcgagtagg cgacgttcag cccccagagc accgagaaag
     6061 ccatccagcc cttgctgctt tcccgggcga tcgcgcccat ggcggtcacg cagggcacgt
     6121 agagcaggac gaagaccaga tagctgtagg ccgcgatgct gttcaggccg ccgatcacca
     6181 ggctgaccag gatgatcacc ttgccggccc gcaccacgaa gccgcgcagg cgcgaccagg
     6241 tctgcagcag caggctctta aggtgtggca catggtacag cggcagttcc atgacgaacg
     6301 gcgaggcctc gccgcgcatc aaggtgtgct tgagcaacag cccggtgagg atcgccacga
     6361 cgatgccgag caggtacagg gagaagatca ccagcgcgcc gccctggccg aagaaggccc
     6421 cggcgaatac ggcgaagatc gccaggcgcg cgccgcagga catgaacggc gccatcatga
     6481 tggtaatcag ccgctcgcgc tgcgcgtcca gggtccgcgc gcccatgatc gacggtacgt
     6541 tgcagccgaa accgacgatc agcggcacga acgacttgcc cggcaggccc agcgcctgca
     6601 tcaggcggtc catgacgaag gccgcgcgtg ccatgtagcc ggaatcctcg agcagcgaga
     6661 ggaacaggta catcaagccg atctgcggca ccagcggcaa caccgtgttg accccgccac
     6721 cgatgccctg ggcgaggaac accgtgagcc aatcgggcag gccgaagcgg atacccagcc
     6781 actggatgcc gtcgatgaag atcgccgacg agcccttgtc gaagatcggc tgtagcgccc
     6841 cgccgatgtt gatggcgaag aagaacatca ggtacatcac cagcaggaag atcggcagac
     6901 ccagccagcg attcaggacg acccggtcga gccattgggt cagccggtgc ggct
//
