LOCUS       sequence-464183-        5999 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Heterologous protein expression of rho GDP-dissociation inhibitor 1 
            (Rho GDI alpha) 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_2JHX
FEATURES             Location/Qualifiers
     source          1..5999
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(63..80)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(807..836)
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     RBS             859..881
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    896..920
                     /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(920..939)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(921..939)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1027..1046)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        1252..1329
                     /gene="lacI"
                     /label=lacI promoter
     CDS             1330..2412
                     /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(1349..1368)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             3221..3412
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(3406..3428)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3514..3653
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3569..3588
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3668..3685)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3839..4427)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3919..3938)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(4538..4557)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             4549..5364
                     /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             4549..5364
                     /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(4622..4641)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      complement(5456..5911)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5593..5614)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5805..5824
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     terminator      complement(5948..5995)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     join(5991..5999,1..10)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
ORIGIN
        1 tgctcagcgg tggcagcagc caactcagct tcctttcggg ctttgttagc agccggatct
       61 cagtggtggt ggtggtggtg ctcgagtcat tacgaacccc aatctttctt aatcgtcaga
      121 ttccattccc aggacaggtg gtcggtctta tcgtcgtcgg taaatctgga cttaatgctg
      181 taagagccac gcgccagcat gcccttcgga gcctcctcaa ccggagtcag aaaatgatag
      241 tgttccgcac gtgggccgta agaaccaacc atatagtccg tcttgtcgat tttcacgcct
      301 ttacggtagg tatgctggat gtatttcata ccgctaacaa tttcacggtt cacgcggaag
      361 ctgatcttga tgcgatactc gacaccttct ttcagcacga agctttgctt tttgaacgac
      421 tccaaatcac cggttaaatc cagctccaac ggacccggtg cgctgctgca gactagtgtt
      481 aatccagtta ctacaacatt tccagagccg ccaatctgtt cacggtgggc ttcaatgatg
      541 tcgttgtctt ccatgtccag gtcttccggc gcttgatctg cctggatgcg aataccgtca
      601 tacaggaaac gcaggctatc catttccttg ccctgacgtt tggcaaacgc ttccatcagg
      661 cgacgcagcg gcgtggtctt tttgatctta aagaaaattt cagagctgcc gtcactaact
      721 ttcaggttaa tatgggtttc cggtttcact tccggcttga cttccggctt tgcttcttgg
      781 ttgacttccg aatcttgcag gctaccgtgg tgatggtgat ggtgatgatg gtgatgaatc
      841 ttcgccatca tatgtatatc tccttcttaa agttaaacaa aattatttct agaggggaat
      901 tgttatccgc tcacaattcc cctatagtga gtcgtattaa tttcgcggga tcgagatcga
      961 tctcgatcct ctacgccgga cgcatcgtgg ccggcatcac cggcgccaca ggtgcggttg
     1021 ctggcgccta tatcgccgac atcaccgatg gggaagatcg ggctcgccac ttcgggctca
     1081 tgagcgcttg tttcggcgtg ggtatggtgg caggccccgt ggccggggga ctgttgggcg
     1141 ccatctcctt gcatgcacca ttccttgcgg cggcggtgct caacggcctc aacctactac
     1201 tgggctgctt cctaatgcag gagtcgcata agggagagcg tcgagatccc ggacaccatc
     1261 gaatggcgca aaacctttcg cggtatggca tgatagcgcc cggaagagag tcaattcagg
     1321 gtggtgaatg tgaaaccagt aacgttatac gatgtcgcag agtatgccgg tgtctcttat
     1381 cagaccgttt cccgcgtggt gaaccaggcc agccacgttt ctgcgaaaac gcgggaaaaa
     1441 gtggaagcgg cgatggcgga gctgaattac attcccaacc gcgtggcaca acaactggcg
     1501 ggcaaacagt cgttgctgat tggcgttgcc acctccagtc tggccctgca cgcgccgtcg
     1561 caaattgtcg cggcgattaa atctcgcgcc gatcaactgg gtgccagcgt ggtggtgtcg
     1621 atggtagaac gaagcggcgt cgaagcctgt aaagcggcgg tgcacaatct tctcgcgcaa
     1681 cgcgtcagtg ggctgatcat taactatccg ctggatgacc aggatgccat tgctgtggaa
     1741 gctgcctgca ctaatgttcc ggcgttattt cttgatgtct ctgaccagac acccatcaac
     1801 agtattattt tctcccatga agacggtacg cgactgggcg tggagcatct ggtcgcattg
     1861 ggtcaccagc aaatcgcgct gttagcgggc ccattaagtt ctgtctcggc gcgtctgcgt
     1921 ctggctggct ggcataaata tctcactcgc aatcaaattc agccgatagc ggaacgggaa
     1981 ggcgactgga gtgccatgtc cggttttcaa caaaccatgc aaatgctgaa tgagggcatc
     2041 gttcccactg cgatgctggt tgccaacgat cagatggcgc tgggcgcaat gcgcgccatt
     2101 accgagtccg ggctgcgcgt tggtgcggac atctcggtag tgggatacga cgataccgaa
     2161 gacagctcat gttatatccc gccgttaacc accatcaaac aggattttcg cctgctgggg
     2221 caaaccagcg tggaccgctt gctgcaactc tctcagggcc aggcggtgaa gggcaatcag
     2281 ctgttgcccg tctcactggt gaaaagaaaa accaccctgg cgcccaatac gcaaaccgcc
     2341 tctccccgcg cgttggccga ttcattaatg cagctggcac gacaggtttc ccgactggaa
     2401 agcgggcagt gagcgcaacg caattaatgt aagttagctc actcattagg caccgggatc
     2461 tcgaccgatg cccttgagag ccttcaaccc agtcagctcc ttccggtggg cgcggggcat
     2521 gactatcgtc gccgcactta tgactgtctt ctttatcatg caactcgtag gacaggtgcc
     2581 ggcagcgctc tgggtcattt tcggcgagga ccgctttcgc tggagcgcga cgatgatcgg
     2641 cctgtcgctt gcggtattcg gaatcttgca cgccctcgct caagccttcg tcactggtcc
     2701 cgccaccaaa cgtttcggcg agaagcaggc cattatcgcc ggcatggcgg ccccacgggt
     2761 gcgcatgatc gtgctcctgt cgttgaggac ccggctaggc tggcggggtt gccttactgg
     2821 ttagcagaat gaatcaccga tacgcgagcg aacgtgaagc gactgctgct gcaaaacgtc
     2881 tgcgacctga gcaacaacat gaatggtctt cggtttccgt gtttcgtaaa gtctggaaac
     2941 gcggaagtca gcgccctgca ccattatgtt ccggatctgc atcgcaggat gctgctggct
     3001 accctgtgga acacctacat ctgtattaac gaagcgctgg cattgaccct gagtgatttt
     3061 tctctggtcc cgccgcatcc ataccgccag ttgtttaccc tcacaacgtt ccagtaaccg
     3121 ggcatgttca tcatcagtaa cccgtatcgt gagcatcctc tctcgtttca tcggtatcat
     3181 tacccccatg aacagaaatc ccccttacac ggaggcatca gtgaccaaac aggaaaaaac
     3241 cgcccttaac atggcccgct ttatcagaag ccagacatta acgcttctgg agaaactcaa
     3301 cgagctggac gcggatgaac aggcagacat ctgtgaatcg cttcacgacc acgctgatga
     3361 gctttaccgc agctgcctcg cgcgtttcgg tgatgacggt gaaaacctct gacacatgca
     3421 gctcccggag acggtcacag cttgtctgta agcggatgcc gggagcagac aagcccgtca
     3481 gggcgcgtca gcgggtgttg gcgggtgtcg gggcgcagcc atgacccagt cacgtagcga
     3541 tagcggagtg tatactggct taactatgcg gcatcagagc agattgtact gagagtgcac
     3601 cattgcggtg tgaaataccg cacagatgcg taaggagaaa ataccgcatc aggcgctctt
     3661 ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag
     3721 ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca
     3781 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
     3841 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
     3901 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
     3961 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
     4021 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
     4081 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
     4141 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
     4201 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta
     4261 actacggcta cactagaagg acagtatttg gtatctgcgc tctgctgaag ccagttacct
     4321 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
     4381 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
     4441 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca
     4501 tgaacaataa aactgtctgc ttacataaac agtaatacaa ggggtgttat gagccatatt
     4561 caacgggaaa cgtcttgctc taggccgcga ttaaattcca acatggatgc tgatttatat
     4621 gggtataaat gggctcgcga taatgtcggg caatcaggtg cgacaatcta tcgattgtat
     4681 gggaagcccg atgcgccaga gttgtttctg aaacatggca aaggtagcgt tgccaatgat
     4741 gttacagatg agatggtcag actaaactgg ctgacggaat ttatgcctct tccgaccatc
     4801 aagcatttta tccgtactcc tgatgatgca tggttactca ccactgcgat ccccgggaaa
     4861 acagcattcc aggtattaga agaatatcct gattcaggtg aaaatattgt tgatgcgctg
     4921 gcagtgttcc tgcgccggtt gcattcgatt cctgtttgta attgtccttt taacagcgat
     4981 cgcgtatttc gtctcgctca ggcgcaatca cgaatgaata acggtttggt tgatgcgagt
     5041 gattttgatg acgagcgtaa tggctggcct gttgaacaag tctggaaaga aatgcataaa
     5101 cttttgccat tctcaccgga ttcagtcgtc actcatggtg atttctcact tgataacctt
     5161 atttttgacg aggggaaatt aataggttgt attgatgttg gacgagtcgg aatcgcagac
     5221 cgataccagg atcttgccat cctatggaac tgcctcggtg agttttctcc ttcattacag
     5281 aaacggcttt ttcaaaaata tggtattgat aatcctgata tgaataaatt gcagtttcat
     5341 ttgatgctcg atgagttttt ctaagaatta attcatgagc ggatacatat ttgaatgtat
     5401 ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctaaatt
     5461 gtaagcgtta atattttgtt aaaattcgcg ttaaattttt gttaaatcag ctcatttttt
     5521 aaccaatagg ccgaaatcgg caaaatccct tataaatcaa aagaatagac cgagataggg
     5581 ttgagtgttg ttccagtttg gaacaagagt ccactattaa agaacgtgga ctccaacgtc
     5641 aaagggcgaa aaaccgtcta tcagggcgat ggcccactac gtgaaccatc accctaatca
     5701 agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg gagcccccga
     5761 tttagagctt gacggggaaa gccggcgaac gtggcgagaa aggaagggaa gaaagcgaaa
     5821 ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac caccacaccc
     5881 gccgcgctta atgcgccgct acagggcgcg tcccattcgc caatccggat atagttcctc
     5941 ctttcagcaa aaaacccctc aagacccgtt tagaggcccc aaggggttat gctagttat
//
