LOCUS       sequence-177989-        6015 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  A bacterial expression plasmid encoding ciliary localization 
            effected RP2 (RP2-C86Y,P95L) with N-terminus GST-tag..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6015)
  AUTHORS   Madugula V, Lu L
  TITLE     A ternary complex comprising transportin1, Rab8 and the ciliary 
            targeting signal directs proteins to ciliary membranes.
  JOURNAL   J Cell Sci. 2016 Oct 15;129(20):3922-3934. Epub 2016 Sep 15.
  PUBMED    27633000
REFERENCE   2  (bases 1 to 6015)
  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"; journalName: "J Cell 
            Sci."; date: "2016-10-15"; volume: "129(20)"; pages: "3922-3934. 
            Epub 2016 Sep 15"
COMMENT     Sequence Label: GST-RP2-C86Y,P95L
FEATURES             Location/Qualifiers
     source          1..6015
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      121..709
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     610..629
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     863..880
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        953..1030
                     /gene="lacI"
                     /label=lacI promoter
     CDS             1031..2113
                     /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(1050..1069)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(2162..2167,2168..2185,2186..2192)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2200..2216
                     /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     2224..2240
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2224..2240
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             2236..2763
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(2256..2273)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2256..2272)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2265..2287)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     promoter        join(2865..2870,2871..2886,2887..2893)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    2901..2917
                     /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)."
     CDS             2940..3593
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     3551..3573
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             3600..3617
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     primer_bind     complement(4747..4769)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4914..4932)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5000..5104
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5105..5173,5174..5965)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(5323..5342)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc
       61 ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
      121 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
      181 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      241 cagcagagcg cagataccaa atactgtcct tctagtgtag ccgtagttag gccaccactt
      301 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      361 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      421 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      481 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      541 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      601 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      661 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      721 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      781 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
      841 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcctgat
      901 gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcataaatt ccgacaccat
      961 cgaatggcgc aaaacctttc gcggtatggc atgatagcgc ccggaagaga gtcaattcag
     1021 ggtggtgaat gtgaaaccag taacgttata cgatgtcgca gagtatgccg gtgtctctta
     1081 tcagaccgtt tcccgcgtgg tgaaccaggc cagccacgtt tctgcgaaaa cgcgggaaaa
     1141 agtggaagcg gcgatggcgg agctgaatta cattcccaac cgcgtggcac aacaactggc
     1201 gggcaaacag tcgttgctga ttggcgttgc cacctccagt ctggccctgc acgcgccgtc
     1261 gcaaattgtc gcggcgatta aatctcgcgc cgatcaactg ggtgccagcg tggtggtgtc
     1321 gatggtagaa cgaagcggcg tcgaagcctg taaagcggcg gtgcacaatc ttctcgcgca
     1381 acgcgtcagt gggctgatca ttaactatcc gctggatgac caggatgcca ttgctgtgga
     1441 agctgcctgc actaatgttc cggcgttatt tcttgatgtc tctgaccaga cacccatcaa
     1501 cagtattatt ttctcccatg aagacggtac gcgactgggc gtggagcatc tggtcgcatt
     1561 gggtcaccag caaatcgcgc tgttagcggg cccattaagt tctgtctcgg cgcgtctgcg
     1621 tctggctggc tggcataaat atctcactcg caatcaaatt cagccgatag cggaacggga
     1681 aggcgactgg agtgccatgt ccggttttca acaaaccatg caaatgctga atgagggcat
     1741 cgttcccact gcgatgctgg ttgccaacga tcagatggcg ctgggcgcaa tgcgcgccat
     1801 taccgagtcc gggctgcgcg ttggtgcgga tatctcggta gtgggatacg acgataccga
     1861 agacagctca tgttatatcc cgccgttaac caccatcaaa caggattttc gcctgctggg
     1921 gcaaaccagc gtggaccgct tgctgcaact ctctcagggc caggcggtga agggcaatca
     1981 gctgttgccc gtctcactgg tgaaaagaaa aaccaccctg gcgcccaata cgcaaaccgc
     2041 ctctccccgc gcgttggccg attcattaat gcagctggca cgacaggttt cccgactgga
     2101 aagcgggcag tgagcgcaac gcaattaatg taagttagct cactcattag gcaccccagg
     2161 ctttacactt tatgcttccg gctcgtatgt tgtgtggaat tgtgagcgga taacaatttc
     2221 acacaggaaa cagctatgac catgattacg gattcactgg ccgtcgtttt acaacgtcgt
     2281 gactgggaaa accctggcgt tacccaactt aatcgccttg cagcacatcc ccctttcgcc
     2341 agctggcgta atagcgaaga ggcccgcacc gatcgccctt cccaacagtt gcgcagcctg
     2401 aatggcgaat ggcgctttgc ctggtttccg gcaccagaag cggtgccgga aagctggctg
     2461 gagtgcgatc ttcctgaggc cgatactgtc gtcgtcccct caaactggca gatgcacggt
     2521 tacgatgcgc ccatctacac caacgtgacc tatcccatta cggtcaatcc gccgtttgtt
     2581 cccacggaga atccgacggg ttgttactcg ctcacattta atgttgatga aagctggcta
     2641 caggaaggcc agacgcgaat tatttttgat ggcgttggaa ttagcttatc gactgcacgg
     2701 tgcaccaatg cttctggcgt caggcagcca tcggaagctg tggtatggct gtgcaggtcg
     2761 taaatcactg cataattcgt gtcgctcaag gcgcactccc gttctggata atgttttttg
     2821 cgccgacatc ataacggttc tggcaaatat tctgaaatga gctgttgaca attaatcatc
     2881 ggctcgtata atgtgtggaa ttgtgagcgg ataacaattt cacacaggaa acagtattca
     2941 tgtcccctat actaggttat tggaaaatta agggccttgt gcaacccact cgacttcttt
     3001 tggaatatct tgaagaaaaa tatgaagagc atttgtatga gcgcgatgaa ggtgataaat
     3061 ggcgaaacaa aaagtttgaa ttgggtttgg agtttcccaa tcttccttat tatattgatg
     3121 gtgatgttaa attaacacag tctatggcca tcatacgtta tatagctgac aagcacaaca
     3181 tgttgggtgg ttgtccaaaa gagcgtgcag agatttcaat gcttgaagga gcggttttgg
     3241 atattagata cggtgtttcg agaattgcat atagtaaaga ctttgaaact ctcaaagttg
     3301 attttcttag caagctacct gaaatgctga aaatgttcga agatcgttta tgtcataaaa
     3361 catatttaaa tggtgatcat gtaacccatc ctgacttcat gttgtatgac gctcttgatg
     3421 ttgttttata catggaccca atgtgcctgg atgcgttccc aaaattagtt tgttttaaaa
     3481 aacgtattga agctatccca caaattgata agtacttgaa atccagcaag tatatagcat
     3541 ggcctttgca gggctggcaa gccacgtttg gtggtggcga ccatcctcca aaatcggatc
     3601 tggttccgcg tggatccccg gaattcatgg gctgcttctt ctccaagaga cggaaggctg
     3661 acaaggagtc gcggcccgag aacgaggagg agcggccaaa gcagtacagc tgggatcagc
     3721 gcgagaaggt tgatccaaaa gactacatgt tcagtggact gaaggatgaa acagtaggtc
     3781 gcttacctgg gacggtagca ggacaacagt ttctcattca agactgtgag aactgtaaca
     3841 tctatatttt tgatcactct gctacagtta ccattgatga ctacactaac tgcataattt
     3901 ttctgggact ggtgaaaggc agcgtgtttt tccggaattg cagagattgc aagtgcacat
     3961 tagcctgcca acaatttcgt gtgcgagatt gtagaaagct ggaagtcttt ttgtgttgtg
     4021 ccactcaacc catcattgag tcttcctcaa atatcaaatt tggatgtttt caatggtact
     4081 atcctgaatt agctttccag ttcaaagatg cagggctaag tatcttcaac aatacatgga
     4141 gtaacattca tgactttaca cctgtgtcag gagaactcaa ctggagcctt cttccagaag
     4201 atgctgtggt tcaggactat gttcctatac ctactaccga agagctcaaa gctgttcgtg
     4261 tttccacaga agccaataga agcattgttc caatatcccg gggtcagaga cagaagagca
     4321 gcgatgaatc atgcttagtg gtattatttg ctggtgatta cactattgca aatgccagaa
     4381 aactaattga tgagatggtt ggtaaaggct ttttcctagt tcagacaaag gaagtgtcca
     4441 tgaaagctga ggatgctcaa agggtttttc gggaaaaagc acctgacttc cttcctcttc
     4501 tgaacaaagg tcctgttatt gccttggagt ttaatgggga tggtgctgta gaagtatgtc
     4561 aacttattgt aaacgagata ttcaatggga ccaagatgtt tgtatctgaa agcaaggaga
     4621 cggcatctgg agatgtagac agcttctaca actttgctga tatacagatg ggaatagtcg
     4681 actcgagcgg ccgcatcgtg actgactgac gatctgcctc gcgcgtttcg gtgatgacgg
     4741 tgaaaacctc tgacacatgc agctcccgga gacggtcaca gcttgtctgt aagcggatgc
     4801 cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc ggggcgcagc
     4861 catgacccag tcacgtagcg atagcggagt gtataattct tgaagacgaa agggcctcgt
     4921 gatacgccta tttttatagg ttaatgtcat gataataatg gtttcttaga cgtcaggtgg
     4981 cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa
     5041 tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa
     5101 gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct
     5161 tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg
     5221 tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg
     5281 ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt
     5341 atcccgtgtt gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga
     5401 cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga
     5461 attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac
     5521 gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg
     5581 ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac
     5641 gatgcctgca gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct
     5701 agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct
     5761 gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg
     5821 gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat
     5881 ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg
     5941 tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat
     6001 tgatttaaaa cttca
//
