LOCUS       sequence-170767-        4844 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Middle entry vector for generating MBS tagged RNAs with a 
            fluorescent protein reporter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4844)
  AUTHORS   Campbell PD, Chao JA, Singer RH, Marlow FL
  TITLE     Dynamic visualization of transcription and RNA subcellular 
            localization in zebrafish.
  JOURNAL   Development. 2015 Apr 1;142(7):1368-74. doi: 10.1242/dev.118968. 
            Epub 2015 Mar 10.
  PUBMED    25758462
REFERENCE   2  (bases 1 to 4844)
  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"; doi: 
            "10.1242/dev.118968"; journalName: "Development"; date: "2015-04-1- 
            1"; volume: "142"; issue: "7"; pages: "1368-74"
COMMENT     Sequence Label: pME-mCherry-24xMBS
FEATURES             Location/Qualifiers
     source          1..4844
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      85..673
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     574..593
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     827..844
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(1003..1030)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(1122..1208)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     1257..1279
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1271..1288
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1272..1288
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    1304..1403
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             1428..2138
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEDDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(1573..1591)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1841..1860
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     2035..2058
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     protein_bind    complement(3458..3557)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(3574..3593)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3575..3593)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3598..3614)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3598..3614)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             4045..4836
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
ORIGIN
        1 ccacccatga ccaaaatccc ttaacgtgag ttacgcgtcg ttccactgag cgtcagaccc
       61 cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt
      121 gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac
      181 tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt
      241 gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct
      301 gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga
      361 ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac
      421 acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg
      481 agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt
      541 cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc
      601 tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg
      661 gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc
      721 ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc
      781 ctttgagtga gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag
      841 cgaggaagcg gaagagcgcc caatacgcaa accgcctctc cccgcgcgtt ggccgattca
      901 ttaatgcagc tggcacgaca ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat
      961 taatacgcgt accgctagcc aggaagagtt tgtagaaacg caaaaaggcc atccgtcagg
     1021 atggccttct gcttagtttg atgcctggca gtttatggcg ggcgtcctgc ccgccaccct
     1081 ccgggccgtt gcttcacaac gttcaaatcc gctcccggcg gatttgtcct actcaggaga
     1141 gcgttcaccg acaaacaaca gataaaacga aaggcccagt cttccgactg agcctttcgt
     1201 tttatttgat gcctggcagt tccctactct cgcgttaacg ctagcatgga tgttttccca
     1261 gtcacgacgt tgtaaaacga cggccagtct taagctcggg ccccaaataa tgattttatt
     1321 ttgactgata gtgacctgtt cgttgcaaca aattgatgag caatgctttt ttataatgcc
     1381 aactttgtac aaaaaagcag gctccgaatt cgcccttcgc gggatccatg gtgagcaagg
     1441 gcgaggacga caacatggcc atcatcaagg agttcatgcg cttcaaggtg cacatggagg
     1501 gctccgtgaa cggccacgag ttcgagatcg agggcgaggg cgagggccgc ccctacgagg
     1561 gcacccagac cgccaagctg aaggtgacca agggcggccc cctgcccttc gcctgggaca
     1621 tcctgtcccc tcagttcatg tacggctcca aggcctacgt gaagcacccc gccgacatcc
     1681 ccgactactt gaagctgtcc ttccccgagg gcttcaagtg ggagcgcgtg atgaacttcg
     1741 aggacggcgg cgtggtgacc gtgacccagg actcctccct gcaggacggc gagttcatct
     1801 acaaggtgaa gctgcgcggc accaacttcc cctccgacgg ccccgtaatg cagaagaaga
     1861 ccatgggctg ggaggcctcc tccgagcgga tgtaccccga ggacggcgcc ctgaagggcg
     1921 agatcaagca gaggctgaag ctgaaggacg gcggccacta cgacgccgag gtcaagacca
     1981 cctacaaggc caagaagccc gtgcagctgc ccggcgccta caacgtcaac atcaagctgg
     2041 acatcacctc ccacaacgag gactacacca tcgtggaaca gtacgagcgc gccgagggcc
     2101 gccactccac cggcggcatg gacgagctgt acaagtaagg atcctacggt acttattgcc
     2161 aagaaagcac gagcatcagc cgtgcctcca ggtcgaatct tcaaacgacg acgatcacgc
     2221 gtcgctccag tattccaggg ttcatcagat cctacggtac ttattgccaa gaaagcacga
     2281 gcatcagccg tgcctccagg tcgaatcttc aaacgacgac gatcacgcgt cgctccagta
     2341 ttccagggtt catcagatcc tacggtactt attgccaaga aagcacgagc atcagccgtg
     2401 cctccaggtc gaatcttcaa acgacgacga tcacgcgtcg ctccagtatt ccagggttca
     2461 tcagatccta cggtacttat tgccaagaaa gcacgagcat cagccgtgcc tccaggtcga
     2521 atcttcaaac gacgacgatc acgcgtcgct ccagtattcc agggttcatc agatcctacg
     2581 gtacttattg ccaagaaagc acgagcatca gccgtgcctc caggtcgaat cttcaaacga
     2641 cgacgatcac gcgtcgctcc agtattccag ggttcatcag atcctacggt acttattgcc
     2701 aagaaagcac gagcatcagc cgtgcctcca ggtcgaatct tcaaacgacg acgatcacgc
     2761 gtcgctccag tattccaggg ttcatcagat cctacggtac ttattgccaa gaaagcacga
     2821 gcatcagccg tgcctccagg tcgaatcttc aaacgacgac gatcacgcgt cgctccagta
     2881 ttccagggtt catcagatcc tacggtactt attgccaaga aagcacgagc atcagccgtg
     2941 cctccaggtc gaatcttcaa acgacgacga tcacgcgtcg ctccagtatt ccagggttca
     3001 tcagatccta cggtacttat tgccaagaaa gcacgagcat cagccgtgcc tccaggtcga
     3061 atcttcaaac gacgacgatc acgcgtcgct ccagtattcc agggttcatc agatcctacg
     3121 gtacttattg ccaagaaagc acgagcatca gccgtgcctc caggtcgaat cttcaaacga
     3181 cgacgatcac gcgtcgctcc agtattccag ggttcatcag atcctacggt acttattgcc
     3241 aagaaagcac gagcatcagc cgtgcctcca ggtcgaatct tcaaacgacg acgatcacgc
     3301 gtcgctccag tattccaggg ttcatcagat cctacggtac ttattgccaa gaaagcacga
     3361 gcatcagccg tgcctccagg tcgaatcttc aaacgacgac gatcacgcgt cgctccagta
     3421 ttccagggtt catcagatct cgcgaagggc gaattcgacc cagctttctt gtacaaagtt
     3481 ggcattataa aaaataattg ctcatcaatt tgttgcaacg aacaggtcac tatcagtcaa
     3541 aataaaatca ttatttgcca tccagctgat atcccctata gtgagtcgta ttacatggtc
     3601 atagctgttt cctggcagct ctggcccgtg tctcaaaatc tctgatgtta cattgcacaa
     3661 gataaaaata tatcatcatg cctcctctag accagccagg acagaaatgc ctcgacttcg
     3721 ctgctaccca aggttgccgg gtgacgcaca ccgtggaaac ggatgaaggc acgaacccag
     3781 tggacataag cctgttcggt tcgtaagctg taatgcaagt agcgtatgcg ctcacgcaac
     3841 tggtccagaa ccttgaccga acgcagcggt ggtaacggcg cagtggcggt tttcatggct
     3901 tgttatgact gtttttttgg ggtacagtct atgcctcggg catccaagca gcaagcgcgt
     3961 tacgccgtgg gtcgatgttt gatgttatgg agcagcaacg atgttacgca gcagggcagt
     4021 cgccctaaaa caaagttaaa cattatgagg gaagcggtga tcgccgaagt atcgactcaa
     4081 ctatcagagg tagttggcgt catcgagcgc catctcgaac cgacgttgct ggccgtacat
     4141 ttgtacggct ccgcagtgga tggcggcctg aagccacaca gtgatattga tttgctggtt
     4201 acggtgaccg taaggcttga tgaaacaacg cggcgagctt tgatcaacga ccttttggaa
     4261 acttcggctt cccctggaga gagcgagatt ctccgcgctg tagaagtcac cattgttgtg
     4321 cacgacgaca tcattccgtg gcgttatcca gctaagcgcg aactgcaatt tggagaatgg
     4381 cagcgcaatg acattcttgc aggtatcttc gagccagcca cgatcgacat tgatctggct
     4441 atcttgctga caaaagcaag agaacatagc gttgccttgg taggtccagc ggcggaggaa
     4501 ctctttgatc cggttcctga acaggatcta tttgaggcgc taaatgaaac cttaacgcta
     4561 tggaactcgc cgcccgactg ggctggcgat gagcgaaatg tagtgcttac gttgtcccgc
     4621 atttggtaca gcgcagtaac cggcaaaatc gcgccgaagg atgtcgctgc cgactgggca
     4681 atggagcgcc tgccggccca gtatcagccc gtcatacttg aagctagaca ggcttatctt
     4741 ggacaagaag aagatcgctt ggcctcgcgc gcagatcagt tggaagaatt tgtccactac
     4801 gtgaaaggcg agatcaccaa ggtagtcggc aaataaccct cgag
//
