LOCUS       sequence-333547-        5367 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Rab27A 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 5367)
  TITLE     Generation of mRuby3-fused Rab GTPases constructs
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5367)
  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            "Unpublished"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-Rab27A
FEATURES             Location/Qualifiers
     source          1..5367
                     /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(2733..2757)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(2801..2820)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    2845..2893
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     promoter        complement(3109..3127)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3148..3164)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3148..3164)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3161..3183)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3172..3188
                     /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(3196..3202,3203..3220,3221..3226))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3241..3262
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3379..3396)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3550..4138)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3630..3649)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4309..5100,5101..5169))
                     /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     4932..4951
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5170..5274)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5342..5360
                     /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 gcaccatgtc tgatggagat tatgattacc tcatcaagtt
     2101 tttagctttg ggagactctg gtgtagggaa gaccagtgta ctttaccaat atacagatgg
     2161 taaatttaac tccaaattta tcacaacagt gggcattgat ttcagggaaa aaagagtggt
     2221 gtacagagcc agtgggccgg atggagccac tggcagaggc cagagaatcc acctgcagtt
     2281 atgggacaca gcagggcagg agaggtttcg tagcttaacg acagcgttct tcagagatgc
     2341 tatgggtttt cttctacttt ttgatctgac aaatgagcaa agtttcctca atgtcagaaa
     2401 ctggataagc cagctacaga tgcatgcata ttgtgaaaac ccagatatag tgctgtgtgg
     2461 aaacaagagt gatctggagg accagagagt agtgaaagag gaggaagcca tagcactcgc
     2521 agagaaatat ggaatcccct actttgaaac tagtgctgcc aatgggacaa acataagcca
     2581 agcaattgag atgcttctgg acctgataat gaagcgaatg gaacggtgtg tggacaagtc
     2641 ctggattcct gaaggagtgg tgcgatcaaa tggtcatgcc tctacggatc agttaagtga
     2701 agaaaaggag aaaggggcat gtggctgtta gaacccagct ttcttgtaca aagtggtgat
     2761 atcgtaggtt gacctgcagg gggagatggg ggaggctaac tgaaacacgg aaggagacaa
     2821 taccggaagg aacccgcgct atgacggcaa taaaaagaca gaataaaacg cacgggtgtt
     2881 gggtcgtttg ttcataaacg cggggttcgg tcccagggct ggcactctgt cgatacccca
     2941 ccgagacccc attggggcca atacgcccgc gtttcttcct tttccccacc ccacccccca
     3001 agttcgggtg aaggcccagg gctcgcagcc aacgtcgggg cggcaagccc tgccatagcc
     3061 acgggccccg tgggttaggg acggggtccc ccatgcaagc ttttgttccc tttagtgagg
     3121 gttaatttcg agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc
     3181 gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta
     3241 atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa
     3301 cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat
     3361 tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg
     3421 agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc
     3481 aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt
     3541 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag
     3601 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc
     3661 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc
     3721 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt
     3781 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt
     3841 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc
     3901 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa
     3961 gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa
     4021 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg
     4081 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga
     4141 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg
     4201 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg
     4261 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt
     4321 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact
     4381 ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat
     4441 gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg
     4501 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg
     4561 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat
     4621 tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc
     4681 ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt
     4741 cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc
     4801 agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga
     4861 gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc
     4921 gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa
     4981 acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta
     5041 acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg
     5101 agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg
     5161 aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat
     5221 gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt
     5281 tccccgaaaa gtgccacctg acgtctaaga aaccattatt atcatgacat taacctataa
     5341 aaataggcgt atcacgaggc cctttcg
//
