LOCUS       sequence-464201-        6185 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Heterologous protein expression of peptidyl-tRNA hydrolase (PTH) in 
            Escherichia coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6185)
  TITLE     AI Proteins, Inc. plasmids
REFERENCE   2  (bases 1 to 6185)
  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_6IYE
FEATURES             Location/Qualifiers
     source          1..6185
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     188..207
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     terminator      complement(331..378)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     374..392
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(445..462)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(1375..1404)
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     RBS             1427..1449
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    1464..1488
                     /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(1488..1507)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1489..1507)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1595..1614)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        1820..1897
                     /gene="lacI"
                     /label=lacI promoter
     CDS             1898..2980
                     /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"
     CDS             3789..3980
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(3974..3996)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4082..4221
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4137..4156
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4236..4253)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4407..4995)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4487..4506)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(5106..5125)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             5117..5932
                     /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             5117..5932
                     /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(5190..5209)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      complement(join(6024..6185,1..294))
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6161..6182)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 taaagaacgt ggactccaac gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac
       61 tacgtgaacc atcaccctaa tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc
      121 ggaaccctaa agggagcccc cgatttagag cttgacgggg aaagccggcg aacgtggcga
      181 gaaaggaagg gaagaaagcg aaaggagcgg gcgctagggc gctggcaagt gtagcggtca
      241 cgctgcgcgt aaccaccaca cccgccgcgc ttaatgcgcc gctacagggc gcgtcccatt
      301 cgccaatccg gatatagttc ctcctttcag caaaaaaccc ctcaagaccc gtttagaggc
      361 cccaaggggt tatgctagtt attgctcagc ggtggcagca gccaactcag cttcctttcg
      421 ggctttgtta gcagccggat ctcagtggtg gtggtggtgg tgctcgagtc attacgaacc
      481 tgccggttta tacgcgttga tctgattcat tgcctgcgga acttgaccct gaaccaacag
      541 tttaactttg ctcagggcgt ggtcgatcgc tccatccatc agcgattgct cattgctcgg
      601 agctttaccc agaacatgac cgctgacgcg ctccttagaa cccgggtggc cgataccaat
      661 gcgcaaacga tgaaaatttg ggccaatatg tggcacgatg tcgcgcaggc cgttgtggcc
      721 gccgtgaccg ccaccggtct taagacgaat cacccccgga ttcatatcca gctcgtcatg
      781 cgcaatcagg atcgcttccg gagcaatctg gtagaacttg gaaaacggga ccacggattg
      841 accgctacgg ttcatatagg tcatcggcag taacaatcta acgtcgtgac cttcaatgtt
      901 accacggccg ctgatgccgt ggaatttcgg atcgttcttc agcgtgatgc cgtacttatc
      961 cgccagttgc tccacgaacc aaaaaccggc gttatgacgg gtctgtgcgt attcagaacc
     1021 cgggttgccc agacccacta ttagagaaat atttgacatg tgacttcctc cagagccgcc
     1081 aatctgttca cggtgggctt caatgatgtc gttgtcttcc atgtccaggt cttccggcgc
     1141 ttgatctgcc tggatgcgaa taccgtcata caggaaacgc aggctatcca tttccttgcc
     1201 ctgacgtttg gcaaacgctt ccatcaggcg acgcagcggc gtggtctttt tgatcttaaa
     1261 gaaaatttca gagctgccgt cactaacttt caggttaata tgggtttccg gtttcacttc
     1321 cggcttgact tccggctttg cttcttggtt gacttccgaa tcttgcaggc taccgtggtg
     1381 atggtgatgg tgatgatggt gatgaatctt cgccatcata tgtatatctc cttcttaaag
     1441 ttaaacaaaa ttatttctag aggggaattg ttatccgctc acaattcccc tatagtgagt
     1501 cgtattaatt tcgcgggatc gagatcgatc tcgatcctct acgccggacg catcgtggcc
     1561 ggcatcaccg gcgccacagg tgcggttgct ggcgcctata tcgccgacat caccgatggg
     1621 gaagatcggg ctcgccactt cgggctcatg agcgcttgtt tcggcgtggg tatggtggca
     1681 ggccccgtgg ccgggggact gttgggcgcc atctccttgc atgcaccatt ccttgcggcg
     1741 gcggtgctca acggcctcaa cctactactg ggctgcttcc taatgcagga gtcgcataag
     1801 ggagagcgtc gagatcccgg acaccatcga atggcgcaaa acctttcgcg gtatggcatg
     1861 atagcgcccg gaagagagtc aattcagggt ggtgaatgtg aaaccagtaa cgttatacga
     1921 tgtcgcagag tatgccggtg tctcttatca gaccgtttcc cgcgtggtga accaggccag
     1981 ccacgtttct gcgaaaacgc gggaaaaagt ggaagcggcg atggcggagc tgaattacat
     2041 tcccaaccgc gtggcacaac aactggcggg caaacagtcg ttgctgattg gcgttgccac
     2101 ctccagtctg gccctgcacg cgccgtcgca aattgtcgcg gcgattaaat ctcgcgccga
     2161 tcaactgggt gccagcgtgg tggtgtcgat ggtagaacga agcggcgtcg aagcctgtaa
     2221 agcggcggtg cacaatcttc tcgcgcaacg cgtcagtggg ctgatcatta actatccgct
     2281 ggatgaccag gatgccattg ctgtggaagc tgcctgcact aatgttccgg cgttatttct
     2341 tgatgtctct gaccagacac ccatcaacag tattattttc tcccatgaag acggtacgcg
     2401 actgggcgtg gagcatctgg tcgcattggg tcaccagcaa atcgcgctgt tagcgggccc
     2461 attaagttct gtctcggcgc gtctgcgtct ggctggctgg cataaatatc tcactcgcaa
     2521 tcaaattcag ccgatagcgg aacgggaagg cgactggagt gccatgtccg gttttcaaca
     2581 aaccatgcaa atgctgaatg agggcatcgt tcccactgcg atgctggttg ccaacgatca
     2641 gatggcgctg ggcgcaatgc gcgccattac cgagtccggg ctgcgcgttg gtgcggacat
     2701 ctcggtagtg ggatacgacg ataccgaaga cagctcatgt tatatcccgc cgttaaccac
     2761 catcaaacag gattttcgcc tgctggggca aaccagcgtg gaccgcttgc tgcaactctc
     2821 tcagggccag gcggtgaagg gcaatcagct gttgcccgtc tcactggtga aaagaaaaac
     2881 caccctggcg cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt cattaatgca
     2941 gctggcacga caggtttccc gactggaaag cgggcagtga gcgcaacgca attaatgtaa
     3001 gttagctcac tcattaggca ccgggatctc gaccgatgcc cttgagagcc ttcaacccag
     3061 tcagctcctt ccggtgggcg cggggcatga ctatcgtcgc cgcacttatg actgtcttct
     3121 ttatcatgca actcgtagga caggtgccgg cagcgctctg ggtcattttc ggcgaggacc
     3181 gctttcgctg gagcgcgacg atgatcggcc tgtcgcttgc ggtattcgga atcttgcacg
     3241 ccctcgctca agccttcgtc actggtcccg ccaccaaacg tttcggcgag aagcaggcca
     3301 ttatcgccgg catggcggcc ccacgggtgc gcatgatcgt gctcctgtcg ttgaggaccc
     3361 ggctaggctg gcggggttgc cttactggtt agcagaatga atcaccgata cgcgagcgaa
     3421 cgtgaagcga ctgctgctgc aaaacgtctg cgacctgagc aacaacatga atggtcttcg
     3481 gtttccgtgt ttcgtaaagt ctggaaacgc ggaagtcagc gccctgcacc attatgttcc
     3541 ggatctgcat cgcaggatgc tgctggctac cctgtggaac acctacatct gtattaacga
     3601 agcgctggca ttgaccctga gtgatttttc tctggtcccg ccgcatccat accgccagtt
     3661 gtttaccctc acaacgttcc agtaaccggg catgttcatc atcagtaacc cgtatcgtga
     3721 gcatcctctc tcgtttcatc ggtatcatta cccccatgaa cagaaatccc ccttacacgg
     3781 aggcatcagt gaccaaacag gaaaaaaccg cccttaacat ggcccgcttt atcagaagcc
     3841 agacattaac gcttctggag aaactcaacg agctggacgc ggatgaacag gcagacatct
     3901 gtgaatcgct tcacgaccac gctgatgagc tttaccgcag ctgcctcgcg cgtttcggtg
     3961 atgacggtga aaacctctga cacatgcagc tcccggagac ggtcacagct tgtctgtaag
     4021 cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc gggtgtcggg
     4081 gcgcagccat gacccagtca cgtagcgata gcggagtgta tactggctta actatgcggc
     4141 atcagagcag attgtactga gagtgcacca ttgcggtgtg aaataccgca cagatgcgta
     4201 aggagaaaat accgcatcag gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg
     4261 gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca
     4321 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac
     4381 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac
     4441 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg
     4501 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac
     4561 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat
     4621 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag
     4681 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac
     4741 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt
     4801 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaaggac agtatttggt
     4861 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc
     4921 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga
     4981 aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac
     5041 gaaaactcac gttaagggat tttggtcatg aacaataaaa ctgtctgctt acataaacag
     5101 taatacaagg ggtgttatga gccatattca acgggaaacg tcttgctcta ggccgcgatt
     5161 aaattccaac atggatgctg atttatatgg gtataaatgg gctcgcgata atgtcgggca
     5221 atcaggtgcg acaatctatc gattgtatgg gaagcccgat gcgccagagt tgtttctgaa
     5281 acatggcaaa ggtagcgttg ccaatgatgt tacagatgag atggtcagac taaactggct
     5341 gacggaattt atgcctcttc cgaccatcaa gcattttatc cgtactcctg atgatgcatg
     5401 gttactcacc actgcgatcc ccgggaaaac agcattccag gtattagaag aatatcctga
     5461 ttcaggtgaa aatattgttg atgcgctggc agtgttcctg cgccggttgc attcgattcc
     5521 tgtttgtaat tgtcctttta acagcgatcg cgtatttcgt ctcgctcagg cgcaatcacg
     5581 aatgaataac ggtttggttg atgcgagtga ttttgatgac gagcgtaatg gctggcctgt
     5641 tgaacaagtc tggaaagaaa tgcataaact tttgccattc tcaccggatt cagtcgtcac
     5701 tcatggtgat ttctcacttg ataaccttat ttttgacgag gggaaattaa taggttgtat
     5761 tgatgttgga cgagtcggaa tcgcagaccg ataccaggat cttgccatcc tatggaactg
     5821 cctcggtgag ttttctcctt cattacagaa acggcttttt caaaaatatg gtattgataa
     5881 tcctgatatg aataaattgc agtttcattt gatgctcgat gagtttttct aagaattaat
     5941 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     6001 catttccccg aaaagtgcca cctaaattgt aagcgttaat attttgttaa aattcgcgtt
     6061 aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggca aaatccctta
     6121 taaatcaaaa gaatagaccg agatagggtt gagtgttgtt ccagtttgga acaagagtcc
     6181 actat
//
