LOCUS       sequence-382134-        5443 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses mRuby Rab8a in mammalian cells and zebrafish embryos.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5443)
  AUTHORS   Aljiboury AA, Ingram E, Krishnan N, Ononiwu F, Pal D, Manikas J, 
            Taveras C, Hall NA, Da Silva J, Freshour J, Hehnly H
  TITLE     Rab8, Rab11, and Rab35 coordinate lumen and cilia formation during 
            zebrafish left-right organizer development.
  JOURNAL   PLoS Genet. 2023 May 15;19(5):e1010765. doi: 
            10.1371/journal.pgen.1010765. eCollection 2023 May.
  PUBMED    37186603
REFERENCE   2  (bases 1 to 5443)
  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: "PLoS 
            Genet."; date: "2023-05-15"; volume: "19(5):e1010765. doi"; pages: "
            10.1371/journal.pgen.1010765. eCollection 2023 May"
COMMENT     Sequence Label: pCS2-mRuby-Rab8a
FEATURES             Location/Qualifiers
     source          1..5443
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        35..53
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     35..52
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     regulatory      88..97
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             94..804
                     /codon_start=1
                     /product="monomeric red fluorescent protein derived from 
                     mRuby, with improved photophysical properties (Lam et al., 
                     2012)"
                     /label=mRuby2
                     /translation="MVSKGEELIKENMRMKVVMEGSVNGHQFKCTGEGEGNPYMGTQTM
                     RIKVIEGGPLPFAFDILATSFMYGSRTFIKYPKGIPDFFKQSFPEGFTWERVTRYEDGG
                     VVTVMQDTSLEDGCLVYHVQVRGVNFPSNGPVMQKKTKGWEPNTEMMYPADGGLRGYTH
                     MALKVDGGGHLSCSFVTTYRSKKTVGNIKMPGIHAVDHRLERLEESDNEMFVVQREHAV
                     AKFAGLGGGMDELYK"
     polyA_signal    1495..1629
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1519..1538)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1573..1592
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(1742..1762)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(1742..1760)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1781..1797)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1781..1797)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1794..1816)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1805..1821
                     /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(1829..1835,1836..1853,1854..1859))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1874..1895
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2012..2029)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2183..2771)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2263..2282)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2942..3733,3734..3802))
                     /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     3565..3584
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3803..3907)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(3933..4388)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4070..4091)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4282..4301
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4459..5443
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
ORIGIN
        1 cgccattctg cctggggacg tcggagcaag cttgatttag gtgacactat agaatacaag
       61 ctacttgttc tttttgcagc cgctagagcc accatggtgt ctaagggcga agagctgatc
      121 aaggaaaata tgcgtatgaa ggtggtcatg gaaggttcgg tcaacggcca ccaattcaaa
      181 tgcacaggtg aaggagaagg caatccgtac atgggaactc aaaccatgag gatcaaagtc
      241 atcgagggag gacccctgcc atttgccttt gacattcttg ccacgtcgtt catgtatggc
      301 agccgtactt ttatcaagta cccgaaaggc attcctgatt tctttaaaca gtcctttcct
      361 gagggtttta cttgggaaag agttacgaga tacgaagatg gtggagtcgt caccgtcatg
      421 caggacacca gccttgagga tggctgtctc gtttaccacg tccaagtcag aggggtaaac
      481 tttccctcca atggtcccgt gatgcagaag aagaccaagg gttgggagcc taatacagag
      541 atgatgtatc cagcagatgg tggtctgagg ggatacactc atatggcact gaaagttgat
      601 ggtggtggcc atctgtcttg ctctttcgta acaacttaca ggtcaaaaaa gaccgtcggg
      661 aacatcaaga tgcccggtat ccatgccgtt gatcaccgcc tggaaaggtt agaggaaagt
      721 gacaatgaaa tgttcgtagt acaacgcgaa cacgcagttg ccaagttcgc cgggcttggt
      781 ggtgggatgg acgagctgta caagcgagct caagcttcga attctgcgaa gacctacgat
      841 tacctgttca agctgctgct gatcggggac tcgggggtgg ggaagacctg tgtcctgttc
      901 cgcttctccg aggacgcctt caactcaact ttcatctcca ccataggaat tgactttaaa
      961 attaggacca tagagctcga tggcaagaga attaagctac agatatggga cacagctggt
     1021 caagaacggt ttcggacgat cacaacagcc tattacaggg gcgcaatggg catcatgctg
     1081 gtctatgaca tcaccaacga gaaatccttt gacaatatcc ggaactggat tcggaacatt
     1141 gaggagcatg cttctgcaga tgtcgaaaag atgatactcg gaaacaagtg tgatgtgaac
     1201 gacaaaagac aagtttccaa ggaacgggga gaaaagctgg ccctggacta tggaatcaag
     1261 ttcatggaga ccagtgcgaa ggccaacatc aatgtggaga acgcattttt cactctcgcc
     1321 agagacatca aagcaaagat ggacaaaaaa ttggaaggca acagtcccca agggagcaac
     1381 cagggagtca aaatcacacc agaccagcag aagaggagca gctttttccg atgtgttctt
     1441 ctgtgatcgc cagcacagtc tgcagtctag aactatagtg agtcgtatta cgtagatcca
     1501 gacatgataa gatacattga tgagtttgga caaaccacaa ctagaatgca gtgaaaaaaa
     1561 tgctttattt gtgaaatttg tgatgctatt gctttatttg taaccattat aagctgcaat
     1621 aaacaagtta acaacaacaa ttgcattcat tttatgtttc aggttcaggg ggaggtgtgg
     1681 gaggtttttt aattcgcggc cgcggcgcca atgcattggg cccggtaccc agcttttgtt
     1741 ccctttagtg agggttaatt gcgcgcttgg cgtaatcatg gtcatagctg tttcctgtgt
     1801 gaaattgtta tccgctcaca attccacaca acatacgagc cgggagcata aagtgtaaag
     1861 cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt
     1921 tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag
     1981 gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg
     2041 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat
     2101 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     2161 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     2221 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     2281 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     2341 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     2401 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     2461 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     2521 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     2581 cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct
     2641 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     2701 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     2761 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
     2821 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
     2881 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca
     2941 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca
     3001 tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc
     3061 ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa
     3121 accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc
     3181 agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca
     3241 acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat
     3301 tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag
     3361 cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac
     3421 tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt
     3481 ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt
     3541 gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc
     3601 tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat
     3661 ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca
     3721 gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga
     3781 cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg
     3841 gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg
     3901 ttccgcgcac atttccccga aaagtgccac ctaaattgta agcgttaata ttttgttaaa
     3961 attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa
     4021 aatcccttat aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa
     4081 caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca
     4141 gggcgatggc ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg
     4201 taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc
     4261 ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc
     4321 aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca
     4381 gggcgcgtcc cattcgccat tcaggctgcg caactgttgg gaagggcgat cggtgcgggc
     4441 ctcttcgcta ttacgccagt cgaccgccaa ttcaatatgg cgtatatgga ctcatgccaa
     4501 ttcaatatgg tggatctgga cctgtgccaa ttcaatatgg cgtatatgga ctcgtgccaa
     4561 ttcaatatgg tggatctgga ccccagccaa ttcaatatgg cggacttggc accatgccaa
     4621 ttcaatatgg cggacctggc actgtgccaa ctggggaggg gtctacttgg cacggtgcca
     4681 agtttgagga ggggtcttgg ccctgtgcca agtccgccat attgaattgg catggtgcca
     4741 ataatggcgg ccatattggc tatatgccag gatcaatata taggcaatat ccaatatggc
     4801 cctatgccaa tatggctatt ggccaggttc aatactatgt attggcccta tgccatatag
     4861 tattccatat atgggttttc ctattgacgt agatagcccc tcccaatggg cggtcccata
     4921 taccatatat ggggcttcct aataccgccc atagccactc ccccattgac gtcaatggtc
     4981 tctatatatg gtctttccta ttgacgtcat atgggcggtc ctattgacgt atatggcgcc
     5041 tcccccattg acgtcaatta cggtaaatgg cccgcctggc tcaatgccca ttgacgtcaa
     5101 taggaccacc caccattgac gtcaatggga tggctcattg cccattcata tccgttctca
     5161 cgccccctat tgacgtcaat gacggtaaat ggcccacttg gcagtacatc aatatctatt
     5221 aatagtaact tggcaagtac attactattg gaagtacgcc agggtacatt ggcagtactc
     5281 ccattgacgt caatggcggt aaatggcccg cgatggctgc caagtacatc cccattgacg
     5341 tcaatgggga ggggcaatga cgcaaatggg cgttccattg acgtaaatgg gcggtaggcg
     5401 tgcctaatgg gaggtctata taagcaatgc tcgtttaggg aac
//
