LOCUS       sequence-464172-        5999 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Heterologous protein expression of soluble secreted antigen MPT53 in
            Escherichia coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5999)
  TITLE     AI Proteins, Inc. plasmids
REFERENCE   2  (bases 1 to 5999)
  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_1LU4
FEATURES             Location/Qualifiers
     source          1..5999
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(153..608)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(290..311)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     502..521
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     terminator      complement(645..692)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     688..706
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(759..776)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(1503..1532)
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     RBS             1555..1577
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    1592..1616
                     /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(1616..1635)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1617..1635)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1723..1742)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        1948..2025
                     /gene="lacI"
                     /label=lacI promoter
     CDS             2026..3108
                     /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(2045..2064)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             3917..4108
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(4102..4124)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4210..4349
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4265..4284
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4364..4381)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4535..5123)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4615..4634)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(5234..5253)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             join(5245..5999,1..61)
                     /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             join(5245..5999,1..61)
                     /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(5318..5337)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
ORIGIN
        1 tattgataat cctgatatga ataaattgca gtttcatttg atgctcgatg agtttttcta
       61 agaattaatt catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg
      121 ttccgcgcac atttccccga aaagtgccac ctaaattgta agcgttaata ttttgttaaa
      181 attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa
      241 aatcccttat aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa
      301 caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca
      361 gggcgatggc ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg
      421 taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc
      481 ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc
      541 aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca
      601 gggcgcgtcc cattcgccaa tccggatata gttcctcctt tcagcaaaaa acccctcaag
      661 acccgtttag aggccccaag gggttatgct agttattgct cagcggtggc agcagccaac
      721 tcagcttcct ttcgggcttt gttagcagcc ggatctcagt ggtggtggtg gtggtgctcg
      781 agtcattacg aacccagcgc ggcaacacgg ccggacagct catcttggct catggctgcg
      841 gtcgggttgt tcacgaaggt gctcgtgccg tccgcgcggt aaaacacaaa cgccggctgc
      901 cacggaacgt tataacgcgc ccagatcacg ccgtctgcat cattcaagtt ggtgaagttc
      961 aggttgtatt tggaaacaaa gctctgcata gcaccaacgt ccgcacgcgt cgcaataccc
     1021 acgaaggtga ccgctggatt agcagcggcg acttgcgaca agctcggtgc ttctgcattg
     1081 cagaacggac accacggggt ccaaaaccac agcaccgccg gcttaccctg taagctcgcg
     1141 ccatcgaacg gagcaccaga cagagtagtt gctgtaaatt gtagcctttc atcagctcca
     1201 gagccgccaa tctgttcacg gtgggcttca atgatgtcgt tgtcttccat gtccaggtct
     1261 tccggcgctt gatctgcctg gatgcgaata ccgtcataca ggaaacgcag gctatccatt
     1321 tccttgccct gacgtttggc aaacgcttcc atcaggcgac gcagcggcgt ggtctttttg
     1381 atcttaaaga aaatttcaga gctgccgtca ctaactttca ggttaatatg ggtttccggt
     1441 ttcacttccg gcttgacttc cggctttgct tcttggttga cttccgaatc ttgcaggcta
     1501 ccgtggtgat ggtgatggtg atgatggtga tgaatcttcg ccatcatatg tatatctcct
     1561 tcttaaagtt aaacaaaatt atttctagag gggaattgtt atccgctcac aattccccta
     1621 tagtgagtcg tattaatttc gcgggatcga gatcgatctc gatcctctac gccggacgca
     1681 tcgtggccgg catcaccggc gccacaggtg cggttgctgg cgcctatatc gccgacatca
     1741 ccgatgggga agatcgggct cgccacttcg ggctcatgag cgcttgtttc ggcgtgggta
     1801 tggtggcagg ccccgtggcc gggggactgt tgggcgccat ctccttgcat gcaccattcc
     1861 ttgcggcggc ggtgctcaac ggcctcaacc tactactggg ctgcttccta atgcaggagt
     1921 cgcataaggg agagcgtcga gatcccggac accatcgaat ggcgcaaaac ctttcgcggt
     1981 atggcatgat agcgcccgga agagagtcaa ttcagggtgg tgaatgtgaa accagtaacg
     2041 ttatacgatg tcgcagagta tgccggtgtc tcttatcaga ccgtttcccg cgtggtgaac
     2101 caggccagcc acgtttctgc gaaaacgcgg gaaaaagtgg aagcggcgat ggcggagctg
     2161 aattacattc ccaaccgcgt ggcacaacaa ctggcgggca aacagtcgtt gctgattggc
     2221 gttgccacct ccagtctggc cctgcacgcg ccgtcgcaaa ttgtcgcggc gattaaatct
     2281 cgcgccgatc aactgggtgc cagcgtggtg gtgtcgatgg tagaacgaag cggcgtcgaa
     2341 gcctgtaaag cggcggtgca caatcttctc gcgcaacgcg tcagtgggct gatcattaac
     2401 tatccgctgg atgaccagga tgccattgct gtggaagctg cctgcactaa tgttccggcg
     2461 ttatttcttg atgtctctga ccagacaccc atcaacagta ttattttctc ccatgaagac
     2521 ggtacgcgac tgggcgtgga gcatctggtc gcattgggtc accagcaaat cgcgctgtta
     2581 gcgggcccat taagttctgt ctcggcgcgt ctgcgtctgg ctggctggca taaatatctc
     2641 actcgcaatc aaattcagcc gatagcggaa cgggaaggcg actggagtgc catgtccggt
     2701 tttcaacaaa ccatgcaaat gctgaatgag ggcatcgttc ccactgcgat gctggttgcc
     2761 aacgatcaga tggcgctggg cgcaatgcgc gccattaccg agtccgggct gcgcgttggt
     2821 gcggacatct cggtagtggg atacgacgat accgaagaca gctcatgtta tatcccgccg
     2881 ttaaccacca tcaaacagga ttttcgcctg ctggggcaaa ccagcgtgga ccgcttgctg
     2941 caactctctc agggccaggc ggtgaagggc aatcagctgt tgcccgtctc actggtgaaa
     3001 agaaaaacca ccctggcgcc caatacgcaa accgcctctc cccgcgcgtt ggccgattca
     3061 ttaatgcagc tggcacgaca ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat
     3121 taatgtaagt tagctcactc attaggcacc gggatctcga ccgatgccct tgagagcctt
     3181 caacccagtc agctccttcc ggtgggcgcg gggcatgact atcgtcgccg cacttatgac
     3241 tgtcttcttt atcatgcaac tcgtaggaca ggtgccggca gcgctctggg tcattttcgg
     3301 cgaggaccgc tttcgctgga gcgcgacgat gatcggcctg tcgcttgcgg tattcggaat
     3361 cttgcacgcc ctcgctcaag ccttcgtcac tggtcccgcc accaaacgtt tcggcgagaa
     3421 gcaggccatt atcgccggca tggcggcccc acgggtgcgc atgatcgtgc tcctgtcgtt
     3481 gaggacccgg ctaggctggc ggggttgcct tactggttag cagaatgaat caccgatacg
     3541 cgagcgaacg tgaagcgact gctgctgcaa aacgtctgcg acctgagcaa caacatgaat
     3601 ggtcttcggt ttccgtgttt cgtaaagtct ggaaacgcgg aagtcagcgc cctgcaccat
     3661 tatgttccgg atctgcatcg caggatgctg ctggctaccc tgtggaacac ctacatctgt
     3721 attaacgaag cgctggcatt gaccctgagt gatttttctc tggtcccgcc gcatccatac
     3781 cgccagttgt ttaccctcac aacgttccag taaccgggca tgttcatcat cagtaacccg
     3841 tatcgtgagc atcctctctc gtttcatcgg tatcattacc cccatgaaca gaaatccccc
     3901 ttacacggag gcatcagtga ccaaacagga aaaaaccgcc cttaacatgg cccgctttat
     3961 cagaagccag acattaacgc ttctggagaa actcaacgag ctggacgcgg atgaacaggc
     4021 agacatctgt gaatcgcttc acgaccacgc tgatgagctt taccgcagct gcctcgcgcg
     4081 tttcggtgat gacggtgaaa acctctgaca catgcagctc ccggagacgg tcacagcttg
     4141 tctgtaagcg gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg
     4201 gtgtcggggc gcagccatga cccagtcacg tagcgatagc ggagtgtata ctggcttaac
     4261 tatgcggcat cagagcagat tgtactgaga gtgcaccatt gcggtgtgaa ataccgcaca
     4321 gatgcgtaag gagaaaatac cgcatcaggc gctcttccgc ttcctcgctc actgactcgc
     4381 tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt
     4441 tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg
     4501 ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg
     4561 agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat
     4621 accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta
     4681 ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct
     4741 gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc
     4801 ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa
     4861 gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg
     4921 taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact agaaggacag
     4981 tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt
     5041 gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta
     5101 cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc
     5161 agtggaacga aaactcacgt taagggattt tggtcatgaa caataaaact gtctgcttac
     5221 ataaacagta atacaagggg tgttatgagc catattcaac gggaaacgtc ttgctctagg
     5281 ccgcgattaa attccaacat ggatgctgat ttatatgggt ataaatgggc tcgcgataat
     5341 gtcgggcaat caggtgcgac aatctatcga ttgtatggga agcccgatgc gccagagttg
     5401 tttctgaaac atggcaaagg tagcgttgcc aatgatgtta cagatgagat ggtcagacta
     5461 aactggctga cggaatttat gcctcttccg accatcaagc attttatccg tactcctgat
     5521 gatgcatggt tactcaccac tgcgatcccc gggaaaacag cattccaggt attagaagaa
     5581 tatcctgatt caggtgaaaa tattgttgat gcgctggcag tgttcctgcg ccggttgcat
     5641 tcgattcctg tttgtaattg tccttttaac agcgatcgcg tatttcgtct cgctcaggcg
     5701 caatcacgaa tgaataacgg tttggttgat gcgagtgatt ttgatgacga gcgtaatggc
     5761 tggcctgttg aacaagtctg gaaagaaatg cataaacttt tgccattctc accggattca
     5821 gtcgtcactc atggtgattt ctcacttgat aaccttattt ttgacgaggg gaaattaata
     5881 ggttgtattg atgttggacg agtcggaatc gcagaccgat accaggatct tgccatccta
     5941 tggaactgcc tcggtgagtt ttctccttca ttacagaaac ggctttttca aaaatatgg
//
