LOCUS       sequence-177998-        5618 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  A bacterial expression plasmid encoding Rab8a GTPase-deficient 
            mutant (Rab8a-Q67L) with N-terminus GST..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5618)
  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 5618)
  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-Rab8a-QL
FEATURES             Location/Qualifiers
     source          1..5618
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      12..600
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     501..520
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     754..771
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        844..921
                     /gene="lacI"
                     /label=lacI promoter
     CDS             922..2004
                     /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(941..960)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(2053..2058,2059..2076,2077..2083)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2091..2107
                     /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     2115..2131
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2115..2131
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             2127..2654
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(2147..2164)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2147..2163)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2156..2178)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     promoter        join(2756..2761,2762..2777,2778..2784)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    2792..2808
                     /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             2831..3484
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     3442..3464
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             3491..3508
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     primer_bind     complement(4241..4263)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4408..4426)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4494..4598
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4599..4667,4668..5459)
                     /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(4817..4836)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa
       61 ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag
      121 gtaactggct tcagcagagc gcagatacca aatactgtcc ttctagtgta gccgtagtta
      181 ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta
      241 ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag
      301 ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg
      361 gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg
      421 cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag
      481 cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc
      541 cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa
      601 aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg
      661 ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct
      721 gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa
      781 gagcgcctga tgcggtattt tctccttacg catctgtgcg gtatttcaca ccgcataaat
      841 tccgacacca tcgaatggcg caaaaccttt cgcggtatgg catgatagcg cccggaagag
      901 agtcaattca gggtggtgaa tgtgaaacca gtaacgttat acgatgtcgc agagtatgcc
      961 ggtgtctctt atcagaccgt ttcccgcgtg gtgaaccagg ccagccacgt ttctgcgaaa
     1021 acgcgggaaa aagtggaagc ggcgatggcg gagctgaatt acattcccaa ccgcgtggca
     1081 caacaactgg cgggcaaaca gtcgttgctg attggcgttg ccacctccag tctggccctg
     1141 cacgcgccgt cgcaaattgt cgcggcgatt aaatctcgcg ccgatcaact gggtgccagc
     1201 gtggtggtgt cgatggtaga acgaagcggc gtcgaagcct gtaaagcggc ggtgcacaat
     1261 cttctcgcgc aacgcgtcag tgggctgatc attaactatc cgctggatga ccaggatgcc
     1321 attgctgtgg aagctgcctg cactaatgtt ccggcgttat ttcttgatgt ctctgaccag
     1381 acacccatca acagtattat tttctcccat gaagacggta cgcgactggg cgtggagcat
     1441 ctggtcgcat tgggtcacca gcaaatcgcg ctgttagcgg gcccattaag ttctgtctcg
     1501 gcgcgtctgc gtctggctgg ctggcataaa tatctcactc gcaatcaaat tcagccgata
     1561 gcggaacggg aaggcgactg gagtgccatg tccggttttc aacaaaccat gcaaatgctg
     1621 aatgagggca tcgttcccac tgcgatgctg gttgccaacg atcagatggc gctgggcgca
     1681 atgcgcgcca ttaccgagtc cgggctgcgc gttggtgcgg atatctcggt agtgggatac
     1741 gacgataccg aagacagctc atgttatatc ccgccgttaa ccaccatcaa acaggatttt
     1801 cgcctgctgg ggcaaaccag cgtggaccgc ttgctgcaac tctctcaggg ccaggcggtg
     1861 aagggcaatc agctgttgcc cgtctcactg gtgaaaagaa aaaccaccct ggcgcccaat
     1921 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt
     1981 tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtaagttagc tcactcatta
     2041 ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg
     2101 ataacaattt cacacaggaa acagctatga ccatgattac ggattcactg gccgtcgttt
     2161 tacaacgtcg tgactgggaa aaccctggcg ttacccaact taatcgcctt gcagcacatc
     2221 cccctttcgc cagctggcgt aatagcgaag aggcccgcac cgatcgccct tcccaacagt
     2281 tgcgcagcct gaatggcgaa tggcgctttg cctggtttcc ggcaccagaa gcggtgccgg
     2341 aaagctggct ggagtgcgat cttcctgagg ccgatactgt cgtcgtcccc tcaaactggc
     2401 agatgcacgg ttacgatgcg cccatctaca ccaacgtgac ctatcccatt acggtcaatc
     2461 cgccgtttgt tcccacggag aatccgacgg gttgttactc gctcacattt aatgttgatg
     2521 aaagctggct acaggaaggc cagacgcgaa ttatttttga tggcgttgga attagcttat
     2581 cgactgcacg gtgcaccaat gcttctggcg tcaggcagcc atcggaagct gtggtatggc
     2641 tgtgcaggtc gtaaatcact gcataattcg tgtcgctcaa ggcgcactcc cgttctggat
     2701 aatgtttttt gcgccgacat cataacggtt ctggcaaata ttctgaaatg agctgttgac
     2761 aattaatcat cggctcgtat aatgtgtgga attgtgagcg gataacaatt tcacacagga
     2821 aacagtattc atgtccccta tactaggtta ttggaaaatt aagggccttg tgcaacccac
     2881 tcgacttctt ttggaatatc ttgaagaaaa atatgaagag catttgtatg agcgcgatga
     2941 aggtgataaa tggcgaaaca aaaagtttga attgggtttg gagtttccca atcttcctta
     3001 ttatattgat ggtgatgtta aattaacaca gtctatggcc atcatacgtt atatagctga
     3061 caagcacaac atgttgggtg gttgtccaaa agagcgtgca gagatttcaa tgcttgaagg
     3121 agcggttttg gatattagat acggtgtttc gagaattgca tatagtaaag actttgaaac
     3181 tctcaaagtt gattttctta gcaagctacc tgaaatgctg aaaatgttcg aagatcgttt
     3241 atgtcataaa acatatttaa atggtgatca tgtaacccat cctgacttca tgttgtatga
     3301 cgctcttgat gttgttttat acatggaccc aatgtgcctg gatgcgttcc caaaattagt
     3361 ttgttttaaa aaacgtattg aagctatccc acaaattgat aagtacttga aatccagcaa
     3421 gtatatagca tggcctttgc agggctggca agccacgttt ggtggtggcg accatcctcc
     3481 aaaatcggat ctggttccgc gtggatcaga attcgcgaag acctacgatt acctgttcaa
     3541 gctgctgctg atcggggact cgggggtggg gaagacctgt gtcctgttcc gcttctccga
     3601 ggacgccttc aactccactt ttatctccac cataggaatt gactttaaaa ttaggaccat
     3661 agagctcgat ggcaagagaa ttaaactgca gatatgggac acagccggtc tggaacggtt
     3721 tcggacgatc acaacggcct actacagggg tgcaatgggc atcatgctgg tctacgacat
     3781 caccaacgag aagtccttcg acaacatccg gaactggatt cgcaacattg aggagcacgc
     3841 ctctgcagac gtcgaaaaga tgatactcgg gaacaagtgt gatgtgaatg acaagagaca
     3901 agtttccaag gaacggggag aaaagctggc cctcgactat ggaatcaagt tcatggagac
     3961 cagcgcgaag gccaacatca atgtggaaaa tgcatttttc actctcgcca gagatatcaa
     4021 agcaaaaatg gacaaaaaat tggaaggcaa cagcccccag gggagcaacc agggagtcaa
     4081 aatcacaccg gaccagcaga agaggagcag ctttttccga tgtgttcttc tgtagggatc
     4141 cttaacaatt ctagactcca tgggtcgact cgagctcaag cttaattcat cgtgactgac
     4201 tgacgatctg cctcgcgcgt ttcggtgatg acggtgaaaa cctctgacac atgcagctcc
     4261 cggagacggt cacagcttgt ctgtaagcgg atgccgggag cagacaagcc cgtcagggcg
     4321 cgtcagcggg tgttggcggg tgtcggggcg cagccatgac ccagtcacgt agcgatagcg
     4381 gagtgtataa ttcttgaaga cgaaagggcc tcgtgatacg cctattttta taggttaatg
     4441 tcatgataat aatggtttct tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa
     4501 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
     4561 cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg
     4621 tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc
     4681 tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg
     4741 atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga
     4801 gcacttttaa agttctgcta tgtggcgcgg tattatcccg tgttgacgcc gggcaagagc
     4861 aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag
     4921 aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga
     4981 gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg
     5041 cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga
     5101 atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgcagcaatg gcaacaacgt
     5161 tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact
     5221 ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt
     5281 ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg
     5341 ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta
     5401 tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac
     5461 tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat ttttaattta
     5521 aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt
     5581 tttcgttcca ctgagcgtca gaccccgtag aaaagatc
//
