LOCUS       sequence-464174-        5921 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Heterologous protein expression of middle transcription regulatory 
            protein motA in Escherichia coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5921)
  TITLE     AI Proteins, Inc. plasmids
REFERENCE   2  (bases 1 to 5921)
  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_1KAF
FEATURES             Location/Qualifiers
     source          1..5921
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     37..56
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     terminator      complement(180..227)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     223..241
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(294..311)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(960..989)
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     RBS             1012..1034
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    1049..1073
                     /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(1073..1092)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1074..1092)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1180..1199)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        1405..1482
                     /gene="lacI"
                     /label=lacI promoter
     CDS             1483..2565
                     /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(1502..1521)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             3374..3565
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(3559..3581)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3667..3806
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3722..3741
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3821..3838)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3992..4580)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4072..4091)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(4691..4710)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             4702..5517
                     /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             4702..5517
                     /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(4775..4794)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      complement(join(5609..5921,1..143))
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5746..5767)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 ttgacgggga aagccggcga acgtggcgag aaaggaaggg aagaaagcga aaggagcggg
       61 cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta accaccacac ccgccgcgct
      121 taatgcgccg ctacagggcg cgtcccattc gccaatccgg atatagttcc tcctttcagc
      181 aaaaaacccc tcaagacccg tttagaggcc ccaaggggtt atgctagtta ttgctcagcg
      241 gtggcagcag ccaactcagc ttcctttcgg gctttgttag cagccggatc tcagtggtgg
      301 tggtggtggt gctcgagtca ttacgaaccc agctcgctcg cgacggtaat aactgcctca
      361 atgttttccg cagagcgttt gatgtccaaa taaacattgc cgtttttagc gatcttgcag
      421 gacataccga tatcggtgaa tttctggata tggtgctcca tcattttgta accaaaaata
      481 cgcatgttgc cgttgttgtt gatctcgaaa ttacgaatgc cgttcgtgcg cttttccaga
      541 atggctaagt aattgctacg atagatttcc accttcttca acacaaaacc atttttatcc
      601 aggagcttca gcatcaggtc cttatcttcc tccatgtctg atgttatttc cattccagag
      661 ccgccaatct gttcacggtg ggcttcaatg atgtcgttgt cttccatgtc caggtcttcc
      721 ggcgcttgat ctgcctggat gcgaataccg tcatacagga aacgcaggct atccatttcc
      781 ttgccctgac gtttggcaaa cgcttccatc aggcgacgca gcggcgtggt ctttttgatc
      841 ttaaagaaaa tttcagagct gccgtcacta actttcaggt taatatgggt ttccggtttc
      901 acttccggct tgacttccgg ctttgcttct tggttgactt ccgaatcttg caggctaccg
      961 tggtgatggt gatggtgatg atggtgatga atcttcgcca tcatatgtat atctccttct
     1021 taaagttaaa caaaattatt tctagagggg aattgttatc cgctcacaat tcccctatag
     1081 tgagtcgtat taatttcgcg ggatcgagat cgatctcgat cctctacgcc ggacgcatcg
     1141 tggccggcat caccggcgcc acaggtgcgg ttgctggcgc ctatatcgcc gacatcaccg
     1201 atggggaaga tcgggctcgc cacttcgggc tcatgagcgc ttgtttcggc gtgggtatgg
     1261 tggcaggccc cgtggccggg ggactgttgg gcgccatctc cttgcatgca ccattccttg
     1321 cggcggcggt gctcaacggc ctcaacctac tactgggctg cttcctaatg caggagtcgc
     1381 ataagggaga gcgtcgagat cccggacacc atcgaatggc gcaaaacctt tcgcggtatg
     1441 gcatgatagc gcccggaaga gagtcaattc agggtggtga atgtgaaacc agtaacgtta
     1501 tacgatgtcg cagagtatgc cggtgtctct tatcagaccg tttcccgcgt ggtgaaccag
     1561 gccagccacg tttctgcgaa aacgcgggaa aaagtggaag cggcgatggc ggagctgaat
     1621 tacattccca accgcgtggc acaacaactg gcgggcaaac agtcgttgct gattggcgtt
     1681 gccacctcca gtctggccct gcacgcgccg tcgcaaattg tcgcggcgat taaatctcgc
     1741 gccgatcaac tgggtgccag cgtggtggtg tcgatggtag aacgaagcgg cgtcgaagcc
     1801 tgtaaagcgg cggtgcacaa tcttctcgcg caacgcgtca gtgggctgat cattaactat
     1861 ccgctggatg accaggatgc cattgctgtg gaagctgcct gcactaatgt tccggcgtta
     1921 tttcttgatg tctctgacca gacacccatc aacagtatta ttttctccca tgaagacggt
     1981 acgcgactgg gcgtggagca tctggtcgca ttgggtcacc agcaaatcgc gctgttagcg
     2041 ggcccattaa gttctgtctc ggcgcgtctg cgtctggctg gctggcataa atatctcact
     2101 cgcaatcaaa ttcagccgat agcggaacgg gaaggcgact ggagtgccat gtccggtttt
     2161 caacaaacca tgcaaatgct gaatgagggc atcgttccca ctgcgatgct ggttgccaac
     2221 gatcagatgg cgctgggcgc aatgcgcgcc attaccgagt ccgggctgcg cgttggtgcg
     2281 gacatctcgg tagtgggata cgacgatacc gaagacagct catgttatat cccgccgtta
     2341 accaccatca aacaggattt tcgcctgctg gggcaaacca gcgtggaccg cttgctgcaa
     2401 ctctctcagg gccaggcggt gaagggcaat cagctgttgc ccgtctcact ggtgaaaaga
     2461 aaaaccaccc tggcgcccaa tacgcaaacc gcctctcccc gcgcgttggc cgattcatta
     2521 atgcagctgg cacgacaggt ttcccgactg gaaagcgggc agtgagcgca acgcaattaa
     2581 tgtaagttag ctcactcatt aggcaccggg atctcgaccg atgcccttga gagccttcaa
     2641 cccagtcagc tccttccggt gggcgcgggg catgactatc gtcgccgcac ttatgactgt
     2701 cttctttatc atgcaactcg taggacaggt gccggcagcg ctctgggtca ttttcggcga
     2761 ggaccgcttt cgctggagcg cgacgatgat cggcctgtcg cttgcggtat tcggaatctt
     2821 gcacgccctc gctcaagcct tcgtcactgg tcccgccacc aaacgtttcg gcgagaagca
     2881 ggccattatc gccggcatgg cggccccacg ggtgcgcatg atcgtgctcc tgtcgttgag
     2941 gacccggcta ggctggcggg gttgccttac tggttagcag aatgaatcac cgatacgcga
     3001 gcgaacgtga agcgactgct gctgcaaaac gtctgcgacc tgagcaacaa catgaatggt
     3061 cttcggtttc cgtgtttcgt aaagtctgga aacgcggaag tcagcgccct gcaccattat
     3121 gttccggatc tgcatcgcag gatgctgctg gctaccctgt ggaacaccta catctgtatt
     3181 aacgaagcgc tggcattgac cctgagtgat ttttctctgg tcccgccgca tccataccgc
     3241 cagttgttta ccctcacaac gttccagtaa ccgggcatgt tcatcatcag taacccgtat
     3301 cgtgagcatc ctctctcgtt tcatcggtat cattaccccc atgaacagaa atccccctta
     3361 cacggaggca tcagtgacca aacaggaaaa aaccgccctt aacatggccc gctttatcag
     3421 aagccagaca ttaacgcttc tggagaaact caacgagctg gacgcggatg aacaggcaga
     3481 catctgtgaa tcgcttcacg accacgctga tgagctttac cgcagctgcc tcgcgcgttt
     3541 cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca cagcttgtct
     3601 gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg ttggcgggtg
     3661 tcggggcgca gccatgaccc agtcacgtag cgatagcgga gtgtatactg gcttaactat
     3721 gcggcatcag agcagattgt actgagagtg caccattgcg gtgtgaaata ccgcacagat
     3781 gcgtaaggag aaaataccgc atcaggcgct cttccgcttc ctcgctcact gactcgctgc
     3841 gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat
     3901 ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca
     3961 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc
     4021 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc
     4081 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg
     4141 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta
     4201 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg
     4261 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac
     4321 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag
     4381 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga aggacagtat
     4441 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat
     4501 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc
     4561 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt
     4621 ggaacgaaaa ctcacgttaa gggattttgg tcatgaacaa taaaactgtc tgcttacata
     4681 aacagtaata caaggggtgt tatgagccat attcaacggg aaacgtcttg ctctaggccg
     4741 cgattaaatt ccaacatgga tgctgattta tatgggtata aatgggctcg cgataatgtc
     4801 gggcaatcag gtgcgacaat ctatcgattg tatgggaagc ccgatgcgcc agagttgttt
     4861 ctgaaacatg gcaaaggtag cgttgccaat gatgttacag atgagatggt cagactaaac
     4921 tggctgacgg aatttatgcc tcttccgacc atcaagcatt ttatccgtac tcctgatgat
     4981 gcatggttac tcaccactgc gatccccggg aaaacagcat tccaggtatt agaagaatat
     5041 cctgattcag gtgaaaatat tgttgatgcg ctggcagtgt tcctgcgccg gttgcattcg
     5101 attcctgttt gtaattgtcc ttttaacagc gatcgcgtat ttcgtctcgc tcaggcgcaa
     5161 tcacgaatga ataacggttt ggttgatgcg agtgattttg atgacgagcg taatggctgg
     5221 cctgttgaac aagtctggaa agaaatgcat aaacttttgc cattctcacc ggattcagtc
     5281 gtcactcatg gtgatttctc acttgataac cttatttttg acgaggggaa attaataggt
     5341 tgtattgatg ttggacgagt cggaatcgca gaccgatacc aggatcttgc catcctatgg
     5401 aactgcctcg gtgagttttc tccttcatta cagaaacggc tttttcaaaa atatggtatt
     5461 gataatcctg atatgaataa attgcagttt catttgatgc tcgatgagtt tttctaagaa
     5521 ttaattcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
     5581 gcgcacattt ccccgaaaag tgccacctaa attgtaagcg ttaatatttt gttaaaattc
     5641 gcgttaaatt tttgttaaat cagctcattt tttaaccaat aggccgaaat cggcaaaatc
     5701 ccttataaat caaaagaata gaccgagata gggttgagtg ttgttccagt ttggaacaag
     5761 agtccactat taaagaacgt ggactccaac gtcaaagggc gaaaaaccgt ctatcagggc
     5821 gatggcccac tacgtgaacc atcaccctaa tcaagttttt tggggtcgag gtgccgtaaa
     5881 gcactaaatc ggaaccctaa agggagcccc cgatttagag c
//
