LOCUS       sequence-464146-        5999 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Heterologous protein expression of cellular retinol-binding protein 
            type II 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_1KQX
FEATURES             Location/Qualifiers
     source          1..5999
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(141..188)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     184..202
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(255..272)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(999..1028)
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     RBS             1051..1073
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    1088..1112
                     /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(1112..1131)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1113..1131)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1219..1238)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        1444..1521
                     /gene="lacI"
                     /label=lacI promoter
     CDS             1522..2604
                     /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(1541..1560)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             3413..3604
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(3598..3620)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3706..3845
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3761..3780
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3860..3877)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4031..4619)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4111..4130)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(4730..4749)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             4741..5556
                     /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             4741..5556
                     /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(4814..4833)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      complement(join(5648..5999,1..104))
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5785..5806)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     join(5997..5999,1..17)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 gaagaaagcg aaaggagcgg gcgctagggc gctggcaagt gtagcggtca cgctgcgcgt
       61 aaccaccaca cccgccgcgc ttaatgcgcc gctacagggc gcgtcccatt cgccaatccg
      121 gatatagttc ctcctttcag caaaaaaccc ctcaagaccc gtttagaggc cccaaggggt
      181 tatgctagtt attgctcagc ggtggcagca gccaactcag cttcctttcg ggctttgtta
      241 gcagccggat ctcagtggtg gtggtggtgg tgctcgagtc attacgaacc gttcttcttt
      301 ttaaaaacct gatgacacac cttatcctgg cagtgaattt ccaggtgcag caggtcacct
      361 tcaatccatt gtttccaacc acggttctct ttctcgcctt tctgcacgca aaccagcttg
      421 tcaccatccc acttgaccaa agttttaacg gtgcggttgt ccagaccctt cgtctgttca
      481 tcaaattcct cgccgatcac gaaattaacc tcgtaattac gaaaggtgct cagggtcttg
      541 gtttcgaact tgtcgccgtt ttgcacgatg accttcgttt gttttaaatg caccgcaatt
      601 ttacgggtag cgaagtcgat gtcgagtgct ttcataacat cctcgaagtt atcattggac
      661 aacatttccc atgttccatt aaaatcagcg ggtccagagc cgccaatctg ttcacggtgg
      721 gcttcaatga tgtcgttgtc ttccatgtcc aggtcttccg gcgcttgatc tgcctggatg
      781 cgaataccgt catacaggaa acgcaggcta tccatttcct tgccctgacg tttggcaaac
      841 gcttccatca ggcgacgcag cggcgtggtc tttttgatct taaagaaaat ttcagagctg
      901 ccgtcactaa ctttcaggtt aatatgggtt tccggtttca cttccggctt gacttccggc
      961 tttgcttctt ggttgacttc cgaatcttgc aggctaccgt ggtgatggtg atggtgatga
     1021 tggtgatgaa tcttcgccat catatgtata tctccttctt aaagttaaac aaaattattt
     1081 ctagagggga attgttatcc gctcacaatt cccctatagt gagtcgtatt aatttcgcgg
     1141 gatcgagatc gatctcgatc ctctacgccg gacgcatcgt ggccggcatc accggcgcca
     1201 caggtgcggt tgctggcgcc tatatcgccg acatcaccga tggggaagat cgggctcgcc
     1261 acttcgggct catgagcgct tgtttcggcg tgggtatggt ggcaggcccc gtggccgggg
     1321 gactgttggg cgccatctcc ttgcatgcac cattccttgc ggcggcggtg ctcaacggcc
     1381 tcaacctact actgggctgc ttcctaatgc aggagtcgca taagggagag cgtcgagatc
     1441 ccggacacca tcgaatggcg caaaaccttt cgcggtatgg catgatagcg cccggaagag
     1501 agtcaattca gggtggtgaa tgtgaaacca gtaacgttat acgatgtcgc agagtatgcc
     1561 ggtgtctctt atcagaccgt ttcccgcgtg gtgaaccagg ccagccacgt ttctgcgaaa
     1621 acgcgggaaa aagtggaagc ggcgatggcg gagctgaatt acattcccaa ccgcgtggca
     1681 caacaactgg cgggcaaaca gtcgttgctg attggcgttg ccacctccag tctggccctg
     1741 cacgcgccgt cgcaaattgt cgcggcgatt aaatctcgcg ccgatcaact gggtgccagc
     1801 gtggtggtgt cgatggtaga acgaagcggc gtcgaagcct gtaaagcggc ggtgcacaat
     1861 cttctcgcgc aacgcgtcag tgggctgatc attaactatc cgctggatga ccaggatgcc
     1921 attgctgtgg aagctgcctg cactaatgtt ccggcgttat ttcttgatgt ctctgaccag
     1981 acacccatca acagtattat tttctcccat gaagacggta cgcgactggg cgtggagcat
     2041 ctggtcgcat tgggtcacca gcaaatcgcg ctgttagcgg gcccattaag ttctgtctcg
     2101 gcgcgtctgc gtctggctgg ctggcataaa tatctcactc gcaatcaaat tcagccgata
     2161 gcggaacggg aaggcgactg gagtgccatg tccggttttc aacaaaccat gcaaatgctg
     2221 aatgagggca tcgttcccac tgcgatgctg gttgccaacg atcagatggc gctgggcgca
     2281 atgcgcgcca ttaccgagtc cgggctgcgc gttggtgcgg acatctcggt agtgggatac
     2341 gacgataccg aagacagctc atgttatatc ccgccgttaa ccaccatcaa acaggatttt
     2401 cgcctgctgg ggcaaaccag cgtggaccgc ttgctgcaac tctctcaggg ccaggcggtg
     2461 aagggcaatc agctgttgcc cgtctcactg gtgaaaagaa aaaccaccct ggcgcccaat
     2521 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt
     2581 tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtaagttagc tcactcatta
     2641 ggcaccggga tctcgaccga tgcccttgag agccttcaac ccagtcagct ccttccggtg
     2701 ggcgcggggc atgactatcg tcgccgcact tatgactgtc ttctttatca tgcaactcgt
     2761 aggacaggtg ccggcagcgc tctgggtcat tttcggcgag gaccgctttc gctggagcgc
     2821 gacgatgatc ggcctgtcgc ttgcggtatt cggaatcttg cacgccctcg ctcaagcctt
     2881 cgtcactggt cccgccacca aacgtttcgg cgagaagcag gccattatcg ccggcatggc
     2941 ggccccacgg gtgcgcatga tcgtgctcct gtcgttgagg acccggctag gctggcgggg
     3001 ttgccttact ggttagcaga atgaatcacc gatacgcgag cgaacgtgaa gcgactgctg
     3061 ctgcaaaacg tctgcgacct gagcaacaac atgaatggtc ttcggtttcc gtgtttcgta
     3121 aagtctggaa acgcggaagt cagcgccctg caccattatg ttccggatct gcatcgcagg
     3181 atgctgctgg ctaccctgtg gaacacctac atctgtatta acgaagcgct ggcattgacc
     3241 ctgagtgatt tttctctggt cccgccgcat ccataccgcc agttgtttac cctcacaacg
     3301 ttccagtaac cgggcatgtt catcatcagt aacccgtatc gtgagcatcc tctctcgttt
     3361 catcggtatc attaccccca tgaacagaaa tcccccttac acggaggcat cagtgaccaa
     3421 acaggaaaaa accgccctta acatggcccg ctttatcaga agccagacat taacgcttct
     3481 ggagaaactc aacgagctgg acgcggatga acaggcagac atctgtgaat cgcttcacga
     3541 ccacgctgat gagctttacc gcagctgcct cgcgcgtttc ggtgatgacg gtgaaaacct
     3601 ctgacacatg cagctcccgg agacggtcac agcttgtctg taagcggatg ccgggagcag
     3661 acaagcccgt cagggcgcgt cagcgggtgt tggcgggtgt cggggcgcag ccatgaccca
     3721 gtcacgtagc gatagcggag tgtatactgg cttaactatg cggcatcaga gcagattgta
     3781 ctgagagtgc accattgcgg tgtgaaatac cgcacagatg cgtaaggaga aaataccgca
     3841 tcaggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     3901 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     3961 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     4021 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     4081 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     4141 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     4201 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     4261 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     4321 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     4381 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     4441 agtggtggcc taactacggc tacactagaa ggacagtatt tggtatctgc gctctgctga
     4501 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     4561 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     4621 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     4681 ggattttggt catgaacaat aaaactgtct gcttacataa acagtaatac aaggggtgtt
     4741 atgagccata ttcaacggga aacgtcttgc tctaggccgc gattaaattc caacatggat
     4801 gctgatttat atgggtataa atgggctcgc gataatgtcg ggcaatcagg tgcgacaatc
     4861 tatcgattgt atgggaagcc cgatgcgcca gagttgtttc tgaaacatgg caaaggtagc
     4921 gttgccaatg atgttacaga tgagatggtc agactaaact ggctgacgga atttatgcct
     4981 cttccgacca tcaagcattt tatccgtact cctgatgatg catggttact caccactgcg
     5041 atccccggga aaacagcatt ccaggtatta gaagaatatc ctgattcagg tgaaaatatt
     5101 gttgatgcgc tggcagtgtt cctgcgccgg ttgcattcga ttcctgtttg taattgtcct
     5161 tttaacagcg atcgcgtatt tcgtctcgct caggcgcaat cacgaatgaa taacggtttg
     5221 gttgatgcga gtgattttga tgacgagcgt aatggctggc ctgttgaaca agtctggaaa
     5281 gaaatgcata aacttttgcc attctcaccg gattcagtcg tcactcatgg tgatttctca
     5341 cttgataacc ttatttttga cgaggggaaa ttaataggtt gtattgatgt tggacgagtc
     5401 ggaatcgcag accgatacca ggatcttgcc atcctatgga actgcctcgg tgagttttct
     5461 ccttcattac agaaacggct ttttcaaaaa tatggtattg ataatcctga tatgaataaa
     5521 ttgcagtttc atttgatgct cgatgagttt ttctaagaat taattcatga gcggatacat
     5581 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt
     5641 gccacctaaa ttgtaagcgt taatattttg ttaaaattcg cgttaaattt ttgttaaatc
     5701 agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag
     5761 accgagatag ggttgagtgt tgttccagtt tggaacaaga gtccactatt aaagaacgtg
     5821 gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg atggcccact acgtgaacca
     5881 tcaccctaat caagtttttt ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa
     5941 gggagccccc gatttagagc ttgacgggga aagccggcga acgtggcgag aaaggaagg
//
