LOCUS       sequence-333537-        5325 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Rab8A fused to mRuby3 fluorescent protein (ex558/em592) tag in N-ter
            under control of a weak L30 promoter..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5325)
  TITLE     Generation of mRuby3-fused Rab GTPases constructs
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5325)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pBS L30 mRuby3-Rab8A
FEATURES             Location/Qualifiers
     source          1..5325
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(31..53)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     153..172
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      239..694
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(326..345)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     536..557
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     820..842
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     834..851
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     835..851
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     858..877
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        858..876
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      1289..1298
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1295..2005
                     /codon_start=1
                     /product="monomeric red fluorescent protein derived from 
                     mRuby2, with improved brightness and photostability (Bajar 
                     et al., 2016)"
                     /label=mRuby3
                     /translation="MVSKGEELIKENMRMKVVMEGSVNGHQFKCTGEGEGRPYEGVQTM
                     RIKVIEGGPLPFAFDILATSFMYGSRTFIKYPADIPDFFKQSFPEGFTWERVTRYEDGG
                     VVTVTQDTSLEDGELVYNVKVRGVNFPSNGPVMQKKTKGWEPNTEMMYPADGGLRGYTD
                     IALKVDGGGHLHCNFVTTYRSKKTVGNIKMPGVHAVDHRLERIEESDNETYVVQREVAV
                     AKYSNLGGGMDELYK"
     protein_bind    2039..2063
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(2691..2715)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(2759..2778)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    2803..2851
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     promoter        complement(3067..3085)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3106..3122)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3106..3122)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3119..3141)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3130..3146
                     /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)."
     promoter        complement(join(3154..3160,3161..3178,3179..3184))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3199..3220
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3337..3354)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3508..4096)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3588..3607)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4267..5058,5059..5127))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     4890..4909
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5128..5232)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5300..5318
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tctcgcgcgt ttcggtgatg acggtgaaaa cctctgacac atgcagctcc cggagacggt
       61 cacagcttgt ctgtaagcgg atgccgggag cagacaagcc cgtcagggcg cgtcagcggg
      121 tgttggcggg tgtcggggct ggcttaacta tgcggcatca gagcagattg tactgagagt
      181 gcaccatatg cggtgtgaaa taccgcacag atgcgtaagg agaaaatacc gcatcaggac
      241 gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct
      301 acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg
      361 ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt
      421 gctttacggc acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca
      481 tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga
      541 ctcttgttcc aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa
      601 gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac
      661 gcgaatttta acaaaatatt aacgcttaca atttccattc gccattcagg ctgcgcaact
      721 gttgggaagg gcgatcggtg cgggcctctt cgctattacg ccagctggcg aaagggggat
      781 gtgctgcaag gcgattaagt tgggtaacgc cagggttttc ccagtcacga cgttgtaaaa
      841 cgacggccag tgaattgtaa tacgactcac tatagggcga attcgagctc ggtacccggg
      901 gatcgcagaa caaacgccca gaaccgaagg ccggagccca gacgccgcag cagcccagcc
      961 tgcgttgtaa cccggtcccc atagcaacca actaccgcag actactctcg caagcgctgt
     1021 tctgcttgtg acccggaccg gcttctcttc cgcttccggt cccacaatcc tctgtcggcc
     1081 tagaagagct ttgcattgtg ggagctcctt cctttctcgc tccccggcca tcttggcggc
     1141 tggtgttggt gagtgagctc tgcggggtaa acgattaggc ggctcgggga gctccgctag
     1201 ctggtgtttg acgctctgga ttataatcct ctcttctgtc ctctgtgtat gctaggttgg
     1261 gggttgtccc gcacctacta gaggatccgc caccatggtg tctaagggcg aagagctgat
     1321 caaggaaaat atgcgtatga aggtggtcat ggaaggttcg gtcaacggcc accaattcaa
     1381 atgcacaggt gaaggagaag gcagaccgta cgagggagtg caaaccatga ggatcaaagt
     1441 catcgaggga ggacccctgc catttgcctt tgacattctt gccacgtcgt tcatgtatgg
     1501 cagccgtacc tttatcaagt acccggccga catccctgat ttctttaaac agtcctttcc
     1561 tgagggtttt acttgggaaa gagttacgag atacgaagat ggtggagtcg tcaccgtcac
     1621 gcaggacacc agccttgagg atggcgagct cgtctacaac gtcaaggtca gaggggtaaa
     1681 ctttccctcc aatggtcccg tgatgcagaa gaagaccaag ggttgggagc ctaatacaga
     1741 gatgatgtat ccagcagatg gtggtctgag aggatacact gacatcgcac tgaaagttga
     1801 tggtggtggc catctgcact gcaacttcgt gacaacttac aggtcaaaaa agaccgtcgg
     1861 gaacatcaag atgcccggtg tccatgccgt tgatcaccgc ctggaaagga tcgaggagag
     1921 tgacaatgaa acctacgtag tgcaaagaga agtggcagtt gccaaataca gcaaccttgg
     1981 tggtggcatg gacgagctgt acaagggtgg ttccggtgga tctggaggtt ctgatatcac
     2041 aagtttgtac aaaaaagcag gcaccatggc gaagacctac gattacctgt tcaagctgct
     2101 gctgatcggg gactcggggg tggggaagac ctgtgtcctg ttccgcttct ccgaggacgc
     2161 cttcaactcc acttttatct ccaccatagg aattgacttt aaaattagga ccatagagct
     2221 cgatggcaag agaattaaac tgcagatatg ggacacagcc ggtcaggaac ggtttcggac
     2281 gatcacaacg gcctactaca ggggtgcaat gggcatcatg ctggtctacg acatcaccaa
     2341 cgagaagtcc ttcgacaaca tccggaactg gattcgcaac attgaggagc acgcctctgc
     2401 agacgtcgaa aagatgatac tcgggaacaa gtgtgatgtg aatgacaaga gacaagtttc
     2461 caaggaacgg ggagaaaagc tggccctcga ctatggaatc aagttcatgg agaccagcgc
     2521 gaaggccaac atcaatgtgg aaaatgcatt tttcactctc gccagagata tcaaagcaaa
     2581 aatggacaaa aaattggaag gcaacagccc ccaggggagc aaccagggag tcaaaatcac
     2641 accggaccag cagaagagga gcagcttttt ccgatgtgtt cttctgtaga acccagcttt
     2701 cttgtacaaa gtggtgatat cgtaggttga cctgcagggg gagatggggg aggctaactg
     2761 aaacacggaa ggagacaata ccggaaggaa cccgcgctat gacggcaata aaaagacaga
     2821 ataaaacgca cgggtgttgg gtcgtttgtt cataaacgcg gggttcggtc ccagggctgg
     2881 cactctgtcg ataccccacc gagaccccat tggggccaat acgcccgcgt ttcttccttt
     2941 tccccacccc accccccaag ttcgggtgaa ggcccagggc tcgcagccaa cgtcggggcg
     3001 gcaagccctg ccatagccac gggccccgtg ggttagggac ggggtccccc atgcaagctt
     3061 ttgttccctt tagtgagggt taatttcgag cttggcgtaa tcatggtcat agctgtttcc
     3121 tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg
     3181 taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc
     3241 cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg
     3301 gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc
     3361 ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac
     3421 agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa
     3481 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca
     3541 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
     3601 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
     3661 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
     3721 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
     3781 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
     3841 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
     3901 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg
     3961 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
     4021 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     4081 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     4141 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     4201 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
     4261 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     4321 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc
     4381 tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc
     4441 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
     4501 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
     4561 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc
     4621 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa
     4681 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
     4741 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
     4801 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
     4861 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa
     4921 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
     4981 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
     5041 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
     5101 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta
     5161 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
     5221 aggggttccg cgcacatttc cccgaaaagt gccacctgac gtctaagaaa ccattattat
     5281 catgacatta acctataaaa ataggcgtat cacgaggccc tttcg
//
