LOCUS       sequence-373587-        5536 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Destination vector for the expression of a fluorescent fusion of a 
            protein of interest with C-ter mScarlet-I into embryos, by mRNA 
            microinjection.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5536)
  TITLE     Lemaire Lab Plasmids 2022
REFERENCE   2  (bases 1 to 5536)
  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"
COMMENT     Sequence Label: pDEST-Spe4-RfA-mScarlet-I-stop
FEATURES             Location/Qualifiers
     source          1..5536
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(87..106)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     297..318
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     581..603
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     595..612
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     596..612
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     619..638
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        619..637
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     gap             682..699
                     /estimated_length=18
     3'UTR           complement(700..825)
                     /label=Xenopus globin 3'-UTR
                     /note="3'-UTR of the major beta-globin gene of Xenopus 
                     laevis"
     CDS             complement(835..1530)
                     /codon_start=1
                     /product="bright monomeric red fluorescent protein evolved 
                     from a synthetic template (Bindels et al., 2016)"
                     /label=mScarlet-I
                     /note="matures faster than mScarlet"
                     /translation="MVSKGEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFSWDILSPQFMYGSRAFIKHPADIPDYYKQSFPEGFKWERVMNFEDGG
                     AVTVTQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDGVLKGDIK
                     MALRLKDGGRYLADFKTTYKAKKPVQMPGAYNVDRKLDITSHNEDYTVVEQYERSEGRH
                     STGGMDELYK"
     primer_bind     complement(1110..1129)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     1379..1397
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     protein_bind    1544..1668
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             complement(1709..2014)
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     complement(1811..1830)
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             complement(2356..3015)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     2930..2949
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(join(3069..3075,3076..3093,3094..3099))
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    complement(3124..3248)
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     5'UTR           complement(3277..3319)
                     /label=Xenopus globin 5'-UTR
                     /note="translational enhancer from the 5'-UTR of the major 
                     beta-globin gene of Xenopus laevis"
     promoter        complement(3344..3362)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3383..3399)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3383..3399)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3396..3418)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3407..3423
                     /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(3431..3437,3438..3455,3456..3461))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3476..3497
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3614..3631)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3785..4373)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3865..3884)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4544..5335,5336..5404))
                     /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     5167..5186
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5405..5509)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      join(5536,1..455)
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
ORIGIN
        1 cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc
       61 tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac
      121 gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag
      181 tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc
      241 atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg
      301 actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata
      361 agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa
      421 cgcgaatttt aacaaaatat taacgcttac aatttccatt cgccattcag gctgcgcaac
      481 tgttgggaag ggcgatcggt gcgggcctct tcgctattac gccagctggc gaaaggggga
      541 tgtgctgcaa ggcgattaag ttgggtaacg ccagggtttt cccagtcacg acgttgtaaa
      601 acgacggcca gtgaattgta atacgactca ctatagggcg aatggcgcgc cggccgactt
      661 ggccctgcag ggggggggkk rnnnnnnnnn nnnnnnnnng tgaagaaact ttctttttat
      721 taggagcaga tacgaatggc tacattttgg gggacaacat tttgtaaagt gtaagttggt
      781 attatgtagc ttagagactc cattcgggtg ttcttgaggc tggttcctag gttacttgta
      841 cagctcgtcc atgccgccgg tggagtggcg gccctcggag cgttcgtact gttccaccac
      901 ggtgtagtcc tcgttgtggg aggtgatgtc caacttgcgg tcgacgttgt aggcgccggg
      961 catctgcacg ggcttcttgg ccttgtaggt ggtcttgaag tccgccaggt agcggccgcc
     1021 gtccttcagg cgcagggcca tcttaatgtc gcccttcagc acgccgtcct cggggtacaa
     1081 ccgctcggtg gacgcttccc agcccattgt cttcttctgc attacggggc cgtcaggagg
     1141 gaagttggtg ccgcggagct tcaccttgta gatcagggtg ccgtcctcca gggaggtgtc
     1201 ctgggtcacg gtcacggcgc cgccgtcctc gaagttcatc acgcgctccc acttgaagcc
     1261 ctcggggaag gactgcttat agtagtcggg gatgtcggcg gggtgcttga tgaaggccct
     1321 ggagccgtac atgaactgag gggacaggat gtcccaggag aagggcaggg ggccaccctt
     1381 ggtcaccttc agcttggcgg tctgggtgcc ctcgtagggg cggccctcgc cctcgccctc
     1441 gatctcgaac tcgtggccgt tcatggagcc ctccatgtgc accttgaacc gcatgaactc
     1501 cttgatcact gcctcgccct tgctcaccat atctaagttt atcaccactt tgtacaagaa
     1561 agctgaacga gaaacgtaaa atgatataaa tatcaatata ttaaattaga ttttgcataa
     1621 aaaacagact acataatact gtaaaacaca acatatccag tcactatggt cgacctgcag
     1681 actggctgtg tataagggag cctgacattt atattcccca gaacatcagg ttaatggcgt
     1741 ttttgatgtc attttcgcgg tggctgagat cagccacttc ttccccgata acggagaccg
     1801 gcacactggc catatcggtg gtcatcatgc gccagctttc atccccgata tgcaccaccg
     1861 ggtaaagttc acgggagact ttatctgaca gcagacgtgc actggccagg gggatcacca
     1921 tccgtcgccc gggcgtgtca ataatatcac tctgtacatc cacaaacaga cgataacggc
     1981 tctctctttt ataggtgtaa accttaaact gcatttcacc agtccctgtt ctcgtcagca
     2041 aaagagccgt tcatttcaat aaaccgggcg acctcagcca tcccttcctg attttccgct
     2101 ttccagcgtt cggcacgcag acgacgggct tcattctgca tggttgtgct taccagaccg
     2161 gagatattga catcatatat gccttgagca actgatagct gtcgctgtca actgtcactg
     2221 taatacgctg cttcatagca cacctctttt tgacatactt cgggtataca tatcagtata
     2281 tattcttata ccgcaaaaat cagcgcgcaa atacgcatac tgttatctgg cttttagtaa
     2341 gccggatcca cgcgtttacg ccccgccctg ccactcatcg cagtactgtt gtaattcatt
     2401 aagcattctg ccgacatgga agccatcaca gacggcatga tgaacctgaa tcgccagcgg
     2461 catcagcacc ttgtcgcctt gcgtataata tttgcccatg gtgaaaacgg gggcgaagaa
     2521 gttgtccata ttggccacgt ttaaatcaaa actggtgaaa ctcacccagg gattggctga
     2581 gacgaaaaac atattctcaa taaacccttt agggaaatag gccaggtttt caccgtaaca
     2641 cgccacatct tgcgaatata tgtgtagaaa ctgccggaaa tcgtcgtggt attcactcca
     2701 gagcgatgaa aacgtttcag tttgctcatg gaaaacggtg taacaagggt gaacactatc
     2761 ccatatcacc agctcaccgt ctttcattgc catacggaat tccggatgag cattcatcag
     2821 gcgggcaaga atgtgaataa aggccggata aaacttgtgc ttatttttct ttacggtctt
     2881 taaaaaggcc gtaatatcca gctgaacggt ctggttatag gtacattgag caactgactg
     2941 aaatgcctca aaatgttctt tacgatgcca ttgggatata tcaacggtgg tatatccagt
     3001 gatttttttc tccattttag cttccttagc tcctgaaaat ctcgccggat cctaactcaa
     3061 aatccacaca ttatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgcggc
     3121 cgccatagtg actggatatg ttgtgtttta cagtattatg tagtctgttt tttatgcaaa
     3181 atctaattta atatattgat atttatatca ttttacgttt ctcgttcagc ttttttgtac
     3241 aaacttgtga ttacgtaagg cctgacgtca gatctgccaa agttgagcgt ttattctgag
     3301 cttctgcaaa aagaacaagc aagcttagct ccagcttttg ttccctttag tgagggttaa
     3361 tttcgagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca
     3421 caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag
     3481 tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt
     3541 cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc
     3601 gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg
     3661 tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa
     3721 agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg
     3781 cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga
     3841 ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg
     3901 tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg
     3961 gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc
     4021 gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg
     4081 gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca
     4141 ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt
     4201 ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag
     4261 ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg
     4321 gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc
     4381 ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt
     4441 tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt
     4501 ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca
     4561 gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg
     4621 tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac
     4681 cgcgagatcc acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg
     4741 ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc
     4801 gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta
     4861 caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac
     4921 gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc
     4981 ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac
     5041 tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact
     5101 caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa
     5161 tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt
     5221 cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca
     5281 ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa
     5341 aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac
     5401 tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg
     5461 gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc
     5521 gaaaagtgcc acctga
//
