LOCUS       sequence-464192-        6014 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Heterologous protein expression of tumor susceptibility gene 101 
            protein in Escherichia coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6014)
  TITLE     AI Proteins, Inc. plasmids
REFERENCE   2  (bases 1 to 6014)
  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_3OBS
FEATURES             Location/Qualifiers
     source          1..6014
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(61..82)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     273..292
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     terminator      complement(416..463)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     459..477
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(530..547)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(1289..1318)
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     RBS             1341..1363
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    1378..1402
                     /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(1402..1421)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1403..1421)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1509..1528)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        1734..1811
                     /gene="lacI"
                     /label=lacI promoter
     CDS             1812..2894
                     /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(1831..1850)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             3703..3894
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(3888..3910)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3996..4135
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4051..4070
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4150..4167)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4321..4909)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4401..4420)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(5020..5039)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             5031..5846
                     /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             5031..5846
                     /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(5104..5123)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      complement(join(5938..6014,1..379))
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
ORIGIN
        1 gaaatcggca aaatccctta taaatcaaaa gaatagaccg agatagggtt gagtgttgtt
       61 ccagtttgga acaagagtcc actattaaag aacgtggact ccaacgtcaa agggcgaaaa
      121 accgtctatc agggcgatgg cccactacgt gaaccatcac cctaatcaag ttttttgggg
      181 tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga gcccccgatt tagagcttga
      241 cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct
      301 agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat
      361 gcgccgctac agggcgcgtc ccattcgcca atccggatat agttcctcct ttcagcaaaa
      421 aacccctcaa gacccgttta gaggccccaa ggggttatgc tagttattgc tcagcggtgg
      481 cagcagccaa ctcagcttcc tttcgggctt tgttagcagc cggatctcag tggtggtggt
      541 ggtggtgctc gagtcattac gaaccgctga acaccggcgg ttcgtcgccg aacaccacga
      601 tcataacctg gatcaaacct aagagatcgc tttgtgggtg tttccactca tgcaggtacg
      661 gcagatagat tttgccgttc gcgtccacgt gcttacccgt tttaatggtc atagaggagg
      721 tcggcttaac aaaacaaatc ggcgggttgt acgggtaagt gtcaagcaac cacaggcaaa
      781 tcgggatatt gtaggtgtta ccgcgatacg gaactggaat ggtgcccgtc aaattcatca
      841 gctcacggct accggtgcca tagctgtcca gaaccggctt cagatcttta tacagcgtga
      901 tgacgttcac ggtttcacga acggtcagat cacggtactt gtatttgctc accattttct
      961 ttagttgact ttctgataca gctccagagc cgccaatctg ttcacggtgg gcttcaatga
     1021 tgtcgttgtc ttccatgtcc aggtcttccg gcgcttgatc tgcctggatg cgaataccgt
     1081 catacaggaa acgcaggcta tccatttcct tgccctgacg tttggcaaac gcttccatca
     1141 ggcgacgcag cggcgtggtc tttttgatct taaagaaaat ttcagagctg ccgtcactaa
     1201 ctttcaggtt aatatgggtt tccggtttca cttccggctt gacttccggc tttgcttctt
     1261 ggttgacttc cgaatcttgc aggctaccgt ggtgatggtg atggtgatga tggtgatgaa
     1321 tcttcgccat catatgtata tctccttctt aaagttaaac aaaattattt ctagagggga
     1381 attgttatcc gctcacaatt cccctatagt gagtcgtatt aatttcgcgg gatcgagatc
     1441 gatctcgatc ctctacgccg gacgcatcgt ggccggcatc accggcgcca caggtgcggt
     1501 tgctggcgcc tatatcgccg acatcaccga tggggaagat cgggctcgcc acttcgggct
     1561 catgagcgct tgtttcggcg tgggtatggt ggcaggcccc gtggccgggg gactgttggg
     1621 cgccatctcc ttgcatgcac cattccttgc ggcggcggtg ctcaacggcc tcaacctact
     1681 actgggctgc ttcctaatgc aggagtcgca taagggagag cgtcgagatc ccggacacca
     1741 tcgaatggcg caaaaccttt cgcggtatgg catgatagcg cccggaagag agtcaattca
     1801 gggtggtgaa tgtgaaacca gtaacgttat acgatgtcgc agagtatgcc ggtgtctctt
     1861 atcagaccgt ttcccgcgtg gtgaaccagg ccagccacgt ttctgcgaaa acgcgggaaa
     1921 aagtggaagc ggcgatggcg gagctgaatt acattcccaa ccgcgtggca caacaactgg
     1981 cgggcaaaca gtcgttgctg attggcgttg ccacctccag tctggccctg cacgcgccgt
     2041 cgcaaattgt cgcggcgatt aaatctcgcg ccgatcaact gggtgccagc gtggtggtgt
     2101 cgatggtaga acgaagcggc gtcgaagcct gtaaagcggc ggtgcacaat cttctcgcgc
     2161 aacgcgtcag tgggctgatc attaactatc cgctggatga ccaggatgcc attgctgtgg
     2221 aagctgcctg cactaatgtt ccggcgttat ttcttgatgt ctctgaccag acacccatca
     2281 acagtattat tttctcccat gaagacggta cgcgactggg cgtggagcat ctggtcgcat
     2341 tgggtcacca gcaaatcgcg ctgttagcgg gcccattaag ttctgtctcg gcgcgtctgc
     2401 gtctggctgg ctggcataaa tatctcactc gcaatcaaat tcagccgata gcggaacggg
     2461 aaggcgactg gagtgccatg tccggttttc aacaaaccat gcaaatgctg aatgagggca
     2521 tcgttcccac tgcgatgctg gttgccaacg atcagatggc gctgggcgca atgcgcgcca
     2581 ttaccgagtc cgggctgcgc gttggtgcgg acatctcggt agtgggatac gacgataccg
     2641 aagacagctc atgttatatc ccgccgttaa ccaccatcaa acaggatttt cgcctgctgg
     2701 ggcaaaccag cgtggaccgc ttgctgcaac tctctcaggg ccaggcggtg aagggcaatc
     2761 agctgttgcc cgtctcactg gtgaaaagaa aaaccaccct ggcgcccaat acgcaaaccg
     2821 cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt tcccgactgg
     2881 aaagcgggca gtgagcgcaa cgcaattaat gtaagttagc tcactcatta ggcaccggga
     2941 tctcgaccga tgcccttgag agccttcaac ccagtcagct ccttccggtg ggcgcggggc
     3001 atgactatcg tcgccgcact tatgactgtc ttctttatca tgcaactcgt aggacaggtg
     3061 ccggcagcgc tctgggtcat tttcggcgag gaccgctttc gctggagcgc gacgatgatc
     3121 ggcctgtcgc ttgcggtatt cggaatcttg cacgccctcg ctcaagcctt cgtcactggt
     3181 cccgccacca aacgtttcgg cgagaagcag gccattatcg ccggcatggc ggccccacgg
     3241 gtgcgcatga tcgtgctcct gtcgttgagg acccggctag gctggcgggg ttgccttact
     3301 ggttagcaga atgaatcacc gatacgcgag cgaacgtgaa gcgactgctg ctgcaaaacg
     3361 tctgcgacct gagcaacaac atgaatggtc ttcggtttcc gtgtttcgta aagtctggaa
     3421 acgcggaagt cagcgccctg caccattatg ttccggatct gcatcgcagg atgctgctgg
     3481 ctaccctgtg gaacacctac atctgtatta acgaagcgct ggcattgacc ctgagtgatt
     3541 tttctctggt cccgccgcat ccataccgcc agttgtttac cctcacaacg ttccagtaac
     3601 cgggcatgtt catcatcagt aacccgtatc gtgagcatcc tctctcgttt catcggtatc
     3661 attaccccca tgaacagaaa tcccccttac acggaggcat cagtgaccaa acaggaaaaa
     3721 accgccctta acatggcccg ctttatcaga agccagacat taacgcttct ggagaaactc
     3781 aacgagctgg acgcggatga acaggcagac atctgtgaat cgcttcacga ccacgctgat
     3841 gagctttacc gcagctgcct cgcgcgtttc ggtgatgacg gtgaaaacct ctgacacatg
     3901 cagctcccgg agacggtcac agcttgtctg taagcggatg ccgggagcag acaagcccgt
     3961 cagggcgcgt cagcgggtgt tggcgggtgt cggggcgcag ccatgaccca gtcacgtagc
     4021 gatagcggag tgtatactgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
     4081 accattgcgg tgtgaaatac cgcacagatg cgtaaggaga aaataccgca tcaggcgctc
     4141 ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc
     4201 agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa
     4261 catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt
     4321 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
     4381 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
     4441 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
     4501 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
     4561 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
     4621 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
     4681 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
     4741 taactacggc tacactagaa ggacagtatt tggtatctgc gctctgctga agccagttac
     4801 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
     4861 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt
     4921 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt
     4981 catgaacaat aaaactgtct gcttacataa acagtaatac aaggggtgtt atgagccata
     5041 ttcaacggga aacgtcttgc tctaggccgc gattaaattc caacatggat gctgatttat
     5101 atgggtataa atgggctcgc gataatgtcg ggcaatcagg tgcgacaatc tatcgattgt
     5161 atgggaagcc cgatgcgcca gagttgtttc tgaaacatgg caaaggtagc gttgccaatg
     5221 atgttacaga tgagatggtc agactaaact ggctgacgga atttatgcct cttccgacca
     5281 tcaagcattt tatccgtact cctgatgatg catggttact caccactgcg atccccggga
     5341 aaacagcatt ccaggtatta gaagaatatc ctgattcagg tgaaaatatt gttgatgcgc
     5401 tggcagtgtt cctgcgccgg ttgcattcga ttcctgtttg taattgtcct tttaacagcg
     5461 atcgcgtatt tcgtctcgct caggcgcaat cacgaatgaa taacggtttg gttgatgcga
     5521 gtgattttga tgacgagcgt aatggctggc ctgttgaaca agtctggaaa gaaatgcata
     5581 aacttttgcc attctcaccg gattcagtcg tcactcatgg tgatttctca cttgataacc
     5641 ttatttttga cgaggggaaa ttaataggtt gtattgatgt tggacgagtc ggaatcgcag
     5701 accgatacca ggatcttgcc atcctatgga actgcctcgg tgagttttct ccttcattac
     5761 agaaacggct ttttcaaaaa tatggtattg ataatcctga tatgaataaa ttgcagtttc
     5821 atttgatgct cgatgagttt ttctaagaat taattcatga gcggatacat atttgaatgt
     5881 atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctaaa
     5941 ttgtaagcgt taatattttg ttaaaattcg cgttaaattt ttgttaaatc agctcatttt
     6001 ttaaccaata ggcc
//
