LOCUS       sequence-268878-        4503 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of membrane-mCherry-LOV mRNA for microinjection for use 
            of optogenetics in S. purpuratus.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4503)
  AUTHORS   Uchida A, Yajima M
  TITLE     An optogenetic approach to control protein localization during 
            embryogenesis of the sea urchin.
  JOURNAL   Dev Biol. 2018 Sep 1;441(1):19-30. doi: 10.1016/j.ydbio.2018.06.015.
            Epub 2018 Jun 26.
  PUBMED    29958898
REFERENCE   2  (bases 1 to 4503)
  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: 
            "10.1016/j.ydbio.2018.06.015"; journalName: "Dev Biol"; date: 
            "2018-09-1- 1"; volume: "441"; issue: "1"; pages: "19-30"
COMMENT     Sequence Label: pSP64-membrane-mCherry-LOV
FEATURES             Location/Qualifiers
     source          1..4503
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     5'UTR           1..43
                     /label=Xenopus globin 5'-UTR
                     /note="translational enhancer from the 5'-UTR of the major 
                     beta-globin gene of Xenopus laevis"
     CDS             136..843
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(281..299)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     549..568
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     complement(1560..1576)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1560..1576)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1573..1595)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1584..1600
                     /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(1608..1614,1615..1632,1633..1638))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1653..1674
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1791..1808)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1962..2550)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2042..2061)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2721..3512,3513..3581))
                     /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     3344..3363
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3582..3686)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3754..3772
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(3810..3832)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     3932..3951
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4165..4184
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        4474..4492
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     4474..4491
                     /label=SP6
                     /note="SP6 promoter, forward primer"
ORIGIN
        1 cttgttcttt ttgcagaagc tcagaataaa cgctcaactt tggcagatct gatcatgcca
       61 tgggggccca tgctgtgctg tatgagaaga accaaacagg ttgaaaagaa tgatgacgac
      121 caaaagattg gcggcatggt gagcaagggc gaggaggata acatggccat catcaaggag
      181 ttcatgcgct tcaaggtgca catggagggc tccgtgaacg gccacgagtt cgagatcgag
      241 ggcgagggcg agggccgccc ctacgagggc acccagaccg ccaagctgaa ggtgaccaag
      301 ggtggccccc tgcccttcgc ctgggacatc ctgtcccctc agttcatgta cggctccaag
      361 gcctacgtga agcaccccgc cgacatcccc gactacttga agctgtcctt ccccgagggc
      421 ttcaagtggg agcgcgtgat gaacttcgag gacggcggcg tggtgaccgt gacccaggac
      481 tcctccctgc aggacggcga gttcatctac aaggtgaagc tgcgcggcac caacttcccc
      541 tccgacggcc ccgtaatgca gaagaagacc atgggctggg aggcctcctc cgagcggatg
      601 taccccgagg acggcgccct gaagggcgag atcaagcaga ggctgaagct gaaggacggc
      661 ggccactacg acgctgaggt caagaccacc tacaaggcca agaagcccgt gcagctgccc
      721 ggcgcctaca acgtcaacat caagttggac atcacctccc acaacgagga ctacaccatc
      781 gtggaacagt acgaacgcgc cgagggccgc cactccaccg gcggcatgga cgagctgtac
      841 aagtccggag gggatcggtg gagctccacc ggtggcggcc gcggtggatc tggaggttca
      901 ggtggaagtc ctaggttggc tactacactt gaacgtattg agaagaactt tgtcattact
      961 gacccaagat tgccagataa tcccattata ttcgcgtccg atagtttctt gcagttgaca
     1021 gaatatagcc gtgaagaaat tttgggaaga aactgcaggt ttctacaagg tcctgaaact
     1081 gatcgcgcga cagtgagaaa aattagagat gccatagata accaaacaga ggtcactgtt
     1141 cagctgatta attatacaaa gagtggtaaa aagttctgga acctctttca cttgcagcct
     1201 atgcgagatc agaagggaga tgtccagtac tttattgggg ttcagttgga tggaactgag
     1261 catgtccgag atgctgccga gagagaggca gtcatgctga ttaagaaaac tgcagaagag
     1321 attgataaag cggtggatac ctgggtgtaa actagtgact gactaggatc tggttaccac
     1381 taaaccagcc tcaagaacac ccgaatggag tctctaagct acataatacc aacttacact
     1441 ttacaaaatg ttgtccccca aaatgtagcc attcgtatct gctcctaata aaaagaaagt
     1501 cccccccccc tgcaggtcga ctctagagga tccccgggcg agctcgaatt cgtaatcatg
     1561 tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc
     1621 ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg
     1681 ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc
     1741 ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact
     1801 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta
     1861 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag
     1921 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
     1981 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
     2041 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg
     2101 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc
     2161 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac
     2221 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
     2281 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
     2341 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
     2401 agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
     2461 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag
     2521 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct
     2581 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg
     2641 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat
     2701 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc
     2761 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg
     2821 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct
     2881 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca
     2941 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg
     3001 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg
     3061 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc
     3121 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag
     3181 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg
     3241 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag
     3301 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat
     3361 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg
     3421 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca
     3481 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca
     3541 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat
     3601 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag
     3661 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgtctaa
     3721 gaaaccatta ttatcatgac attaacctat aaaaataggc gtatcacgag gccctttcgt
     3781 ctcgcgcgtt tcggtgatga cggtgaaaac ctctgacaca tgcagctccc ggagacggtc
     3841 acagcttgtc tgtaagcgga tgccgggagc agacaagccc gtcagggcgc gtcagcgggt
     3901 gttggcgggt gtcggggctg gcttaactat gcggcatcag agcagattgt actgagagtg
     3961 caccattcga cgctctccct tatgcgactc ctgcattagg aagcagccca gtagtaggtt
     4021 gaggccgttg agcaccgccg ccgcaaggaa tggtgcatgc aaggagatgg cgcccaacag
     4081 tcccccggcc acggggcctg ccaccatacc cacgccgaaa caagcgctca tgagcccgaa
     4141 gtggcgagcc cgatcttccc catcggtgat gtcggcgata taggcgccag caaccgcacc
     4201 tgtggcgccg gtgatgccgg ccacgatgcg tccggcgtag aggatctggc tagcgatgac
     4261 cctgctgatt ggttcgctga ccatttccgg gtgcgggacg gcgttaccag aaactcagaa
     4321 ggttcgtcca accaaaccga ctctgacggc agtttacgag agagatgata gggtctgctt
     4381 cagtaagcca gatgctacac aattaggctt gtacatattg tcgttagaac gcggctacaa
     4441 ttaatacata accttatgta tcatacacat acgatttagg tgacactata gaatacaagc
     4501 ttg
//
