LOCUS       sequence-446898-        3682 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  A Gateway compatible 3' entry clone containing an V5 mScarlet fusion
            followed by SV40 late polyA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3682)
  AUTHORS   Gillespie WB 3rd, Zhang Y, Ruiz OE, Cerda J 3rd, Ortiz-Guzman J, 
            Sherman M, Turner WD, Largoza G, Mosser LE, Fujimoto E, Chien CB, 
            Kwan KM, Arenkiel BR, Devine WP, Wythe JD
  TITLE     MultiSite Assembly of Gateway Induced Clones (MAGIC): a flexible 
            cloning toolbox for use in vertebrate model systems.
  JOURNAL   Development. 2025 Nov 15;152(22):dev204308. doi: 10.1242/dev.204308.
            Epub 2025 Nov 13.
  PUBMED    41230856
REFERENCE   2  (bases 1 to 3682)
  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: 
            "Development."; date: "2025-11-15"; volume: "152(22):dev204308. 
            doi"; pages: " 10.1242/dev.204308. Epub 2025 Nov 13"
COMMENT     Sequence Label: p3E-V5-mScarlet-I_SV40pA (JDW 968)
FEATURES             Location/Qualifiers
     source          1..3682
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     92..109
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(268..295)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(387..473)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     522..544
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     536..553
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     537..553
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(624..747)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     regulatory      765..774
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             774..815
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             831..1526
                     /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(964..982)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1232..1251
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    1594..1728
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1618..1637)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1672..1691
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     protein_bind    complement(1791..1886)
                     /gene="mutant version of attL"
                     /label=attL3
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        complement(1924..1942)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1947..1963)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1947..1963)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2065..2084)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             2076..2885
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(2143..2162)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      3032..3620
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3521..3540
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga
       61 taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga
      121 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
      181 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaata cgcgtaccgc
      241 tagccaggaa gagtttgtag aaacgcaaaa aggccatccg tcaggatggc cttctgctta
      301 gtttgatgcc tggcagttta tggcgggcgt cctgcccgcc accctccggg ccgttgcttc
      361 acaacgttca aatccgctcc cggcggattt gtcctactca ggagagcgtt caccgacaaa
      421 caacagataa aacgaaaggc ccagtcttcc gactgagcct ttcgttttat ttgatgcctg
      481 gcagttccct actctcgcgt taacgctagc atggatgttt tcccagtcac gacgttgtaa
      541 aacgacggcc agtcttaagc tcgggccctg cagctctaga gctcgaattc tacaggtcac
      601 taataccatc taagtagttg gttcatagtg actgcatatg ttgtgtttta cagtattatg
      661 tagtctgttt tttatgcaaa atctaattta atatattgat atttatatca ttttacgttt
      721 ctcgttcaac tttcttgtac aaagtgggat ccactagtgc tagcgccacc atgggtaagc
      781 ctatccctaa ccctctcctc ggtctcgatt ctacgggtgg tgctggtggt atggtgagca
      841 agggcgaggc agtgatcaag gagttcatgc ggttcaaggt gcacatggag ggctccatga
      901 acggccacga gttcgagatc gagggcgagg gcgagggccg cccctacgag ggcacccaga
      961 ccgccaagct gaaggtgacc aagggtggcc ccctgccctt ctcctgggac atcctgtccc
     1021 ctcagttcat gtacggctcc agggccttca tcaagcaccc cgccgacatc cccgactact
     1081 ataagcagtc cttccccgag ggcttcaagt gggagcgcgt gatgaacttc gaggacggcg
     1141 gcgccgtgac cgtgacccag gacacctccc tggaggacgg caccctgatc tacaaggtga
     1201 agctccgcgg caccaacttc cctcctgacg gccccgtaat gcagaagaag acaatgggct
     1261 gggaagcatc caccgagcgg ttgtaccccg aggacggcgt gctgaagggc gacattaaga
     1321 tggccctgcg cctgaaggac ggcggccgct acctggcgga cttcaagacc acctacaagg
     1381 ccaagaagcc cgtgcagatg cccggcgcct acaacgtcga ccgcaagttg gacatcacct
     1441 cccacaacga ggactacacc gtggtggaac agtacgaacg ctccgagggc cgccactcca
     1501 ccggcggcat ggacgagctg tacaagtaag gcgcgccctc gagctcaagc ttcgaattct
     1561 gcagtctaga actatagtga gtcgtattac gtagatccag acatgataag atacattgat
     1621 gagtttggac aaaccacaac tagaatgcag tgaaaaaaat gctttatttg tgaaatttgt
     1681 gatgctattg ctttatttgt aaccattata agctgcaata aacaagttaa caacaacaat
     1741 tgcattcatt ttatgtttca ggttcagggg gaggtgtggg aggtttttta caactttatt
     1801 atacaaagtt ggcattataa aaaagcattg cttatcaatt tgttgcaacg aacaggtcac
     1861 tatcagtcaa aataaaatca ttatttggag ctccatggta gcgttaacgc ggccgcgata
     1921 tcccctatag tgagtcgtat tacatggtca tagctgtttc ctggcagctc tggcccgtgt
     1981 ctcaaaatct ctgatgttac attgcacaag ataaaaatat atcatcatga acaataaaac
     2041 tgtctgctta cataaacagt aatacaaggg gtgttatgag ccatattcaa cgggaaacgt
     2101 cgaggccgcg attaaattcc aacatggatg ctgatttata tgggtataaa tgggctcgcg
     2161 ataatgtcgg gcaatcaggt gcgacaatct atcgcttgta tgggaagccc gatgcgccag
     2221 agttgtttct gaaacatggc aaaggtagcg ttgccaatga tgttacagat gagatggtca
     2281 gactaaactg gctgacggaa tttatgcctc ttccgaccat caagcatttt atccgtactc
     2341 ctgatgatgc atggttactc accactgcga tccccggaaa aacagcattc caggtattag
     2401 aagaatatcc tgattcaggt gaaaatattg ttgatgcgct ggcagtgttc ctgcgccggt
     2461 tgcattcgat tcctgtttgt aattgtcctt ttaacagcga tcgcgtattt cgtctcgctc
     2521 aggcgcaatc acgaatgaat aacggtttgg ttgatgcgag tgattttgat gacgagcgta
     2581 atggctggcc tgttgaacaa gtctggaaag aaatgcataa acttttgcca ttctcaccgg
     2641 attcagtcgt cactcatggt gatttctcac ttgataacct tatttttgac gaggggaaat
     2701 taataggttg tattgatgtt ggacgagtcg gaatcgcaga ccgataccag gatcttgcca
     2761 tcctatggaa ctgcctcggt gagttttctc cttcattaca gaaacggctt tttcaaaaat
     2821 atggtattga taatcctgat atgaataaat tgcagtttca tttgatgctc gatgagtttt
     2881 tctaatcaga attggttaat tggttgtaac actggcagag cattacgctg acttgacggg
     2941 acggcgcaag ctcatgacca aaatccctta acgtgagtta cgcgtcgttc cactgagcgt
     3001 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     3061 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     3121 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc
     3181 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     3241 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     3301 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     3361 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     3421 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     3481 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     3541 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     3601 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     3661 gctggccttt tgctcacatg tt
//
