LOCUS       sequence-446893-        4597 bp    DNA     circular SYN 15-APR-2025
DEFINITION  A Gateway compatible 3' entry clone containing an IRES H2A mCherry 
            followed by a polyA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4597)
  AUTHORS   Gillespie W, Zhang Y, Ruiz OE, Cerda III J, Ortiz-Guzman J, Turner 
            WD, Largoza G, Sherman M, Mosser LE, Fujimoto E, Chien C-B, Kwan KM,
            Arenkiel BR, Devine WP, Wythe JD
  TITLE     Multisite Assembly of Gateway Induced Clones (MAGIC): a flexible 
            cloning toolbox with diverse applications in vertebrate model 
            systems
  JOURNAL   bioRxiv 2024.07.13.603267
REFERENCE   2  (bases 1 to 4597)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "bioRxiv 
            2024.07.13.603267"
COMMENT     Sequence Label: p3E-IRES-H2A-mCherry-SV40-pA (JDW 1256)
FEATURES             Location/Qualifiers
     source          1..4597
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(103..122)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             114..923
                     /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(181..200)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      1101..1689
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1590..1609
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1843..1860
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    1977..1998
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(2013..2018,2019..2036,2037..2043)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2051..2067
                     /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)."
     primer_bind     2056..2078
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2075..2091
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2075..2091
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     2105..2122
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2106..2122
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(2131..2254)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     misc_feature    join(2263..2837,2838..2840,2841..2849)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(2430..2447)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     2657..2676
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             3246..3956
                     /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(3391..3409)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     3659..3678
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     3853..3876
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     polyA_signal    4085..4219
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4109..4128)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4163..4182
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     protein_bind    complement(4314..4409)
                     /gene="mutant version of attL"
                     /label=attL3
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(4428..4447)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4547..4569
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 tgcagctctg gcccgtgtct caaaatctct gatgttacat tgcacaagat aaaaatatat
       61 catcatgaac aataaaactg tctgcttaca taaacagtaa tacaaggggt gttatgagcc
      121 atattcaacg ggaaacgtcg aggccgcgat taaattccaa catggatgct gatttatatg
      181 ggtataaatg ggctcgcgat aatgtcgggc aatcaggtgc gacaatctat cgcttgtatg
      241 ggaagcccga tgcgccagag ttgtttctga aacatggcaa aggtagcgtt gccaatgatg
      301 ttacagatga gatggtcaga ctaaactggc tgacggaatt tatgcctctt ccgaccatca
      361 agcattttat ccgtactcct gatgatgcat ggttactcac cactgcgatc cccggaaaaa
      421 cagcattcca ggtattagaa gaatatcctg attcaggtga aaatattgtt gatgcgctgg
      481 cagtgttcct gcgccggttg cattcgattc ctgtttgtaa ttgtcctttt aacagcgatc
      541 gcgtatttcg tctcgctcag gcgcaatcac gaatgaataa cggtttggtt gatgcgagtg
      601 attttgatga cgagcgtaat ggctggcctg ttgaacaagt ctggaaagaa atgcataaac
      661 ttttgccatt ctcaccggat tcagtcgtca ctcatggtga tttctcactt gataacctta
      721 tttttgacga ggggaaatta ataggttgta ttgatgttgg acgagtcgga atcgcagacc
      781 gataccagga tcttgccatc ctatggaact gcctcggtga gttttctcct tcattacaga
      841 aacggctttt tcaaaaatat ggtattgata atcctgatat gaataaattg cagtttcatt
      901 tgatgctcga tgagtttttc taatcagaat tggttaattg gttgtaacat tattcagatt
      961 gggcttgatt taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat
     1021 aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta
     1081 gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa
     1141 acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt
     1201 tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag
     1261 ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta
     1321 atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca
     1381 agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag
     1441 cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa
     1501 agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga
     1561 acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc
     1621 gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc
     1681 ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt
     1741 gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt
     1801 gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag
     1861 gaagcggaag agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa
     1921 tgcagctggc acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat
     1981 gtgagttagc tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg
     2041 ttgtgtggaa ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac
     2101 accatgtaaa acgacggcca gtcttaagcc catagtgact gcatatgttg tgttttacag
     2161 tattatgtag tctgtttttt atgcaaaatc taatttaata tattgatatt tatatcattt
     2221 tacgtttctc gttcaacttt cttgtacaaa gtggcatcga tacccctctc cctccccccc
     2281 ccctaacgtt actggccgaa gccgcttgga ataaggccgg tgtgcgtttg tctatatgtt
     2341 attttccacc atattgccgt cttttggcaa tgtgagggcc cggaaacctg gccctgtctt
     2401 cttgacgagc attcctaggg gtctttcccc tctcgccaaa ggaatgcaag gtctgttgaa
     2461 tgtcgtgaag gaagcagttc ctctggaagc ttcttgaaga caaacaacgt ctgtagcgac
     2521 cctttgcagg cagcggaacc ccccacctgg cgacaggtgc ctctgcggcc aaaagccacg
     2581 tgtataagat acacctgcaa aggcggcaca accccagtgc cacgttgtga gttggatagt
     2641 tgtggaaaga gtcaaatggc tctcctcaag cgtattcaac aaggggctga aggatgccca
     2701 gaaggtaccc cattgtatgg gatctgatct ggggcctcgg tacacatgct ttacatgtgt
     2761 ttagtcgagg ttaaaaaaac gtctaggccc cccgaaccac ggggacgtgg ttttcctttg
     2821 aaaaacacga tgataatatg gccacaacca tggcaggtgg aaaagcaggt aaagacagtg
     2881 gaaaagccaa gacgaaagcc gtctccagat ctcagagagc tggtctgcag ttccctgtag
     2941 gtcgtatcca caggcacctg aagacccgaa ccaccagcca tggccgtgtc ggagctacag
     3001 ctgcggttta cagtgctgct atcctggagt atctcaccgc tgaggtgctg gagttggctg
     3061 gaaacgcttc taaggatctg aaggtgaagc gaatcactcc cagacattta cagctggcca
     3121 tccgcggaga cgaggagctg gactcgctta tcaaggccac cattgctgga ggaggagtca
     3181 ttcctcacat ccacaagtct ctgattggta agaagggcca gcagaaaacc gcaggatccg
     3241 gggggatggt gagcaagggc gaggaggaca acatggccat catcaaggag ttcatgcgct
     3301 tcaaggtgca catggagggc tccgtgaacg gccacgagtt cgagatcgag ggcgagggcg
     3361 agggccgccc ctacgagggc acccagaccg ccaagctgaa ggtgaccaag ggcggccccc
     3421 tgcccttcgc ctgggacatc ctgtcccctc agttcatgta cggctccaag gcctacgtga
     3481 agcaccccgc cgacatcccc gactacttga agctgtcctt ccccgagggc ttcaagtggg
     3541 agcgcgtgat gaacttcgag gacggcggcg tggtgaccgt gacccaggac tcctccctgc
     3601 aggacggcga gttcatctac aaggtgaagc tgcgcggcac caacttcccc tccgacggcc
     3661 ccgtaatgca gaagaagacc atgggctggg aggcctcctc cgagcggatg taccccgagg
     3721 acggcgccct gaagggcgag atcaagcaga ggctgaagct gaaggacggc ggccactacg
     3781 acgccgaggt caagaccacc tacaaggcca agaagcccgt gcagctgccc ggcgcctaca
     3841 acgtcaacat caagctggac atcacctccc acaacgagga ctacaccatc gtggaacagt
     3901 acgagcgcgc cgagggccgc cactccaccg gcggcatgga cgagctgtac aagtaataac
     3961 tcgagggagg agggtcgaca agatctacta gtaagctgaa ccctcctgat gagagtggcc
     4021 ccggctgcat gagctgcaag tgtgtgctct cctgaacccg ggctatagtg agtcgtatta
     4081 cgtagatcca gacatgataa gatacattga tgagtttgga caaaccacaa ctagaatgca
     4141 gtgaaaaaaa tgctttattt gtgaaatttg tgatgctatt gctttatttg taaccattat
     4201 aagctgcaat aaacaagtta acaacaacaa ttgcattcat tttatgtttc aggttcaggg
     4261 ggaggtgtgg gaggtttttt ctgctctaga agcgctcgaa ttcggcacgc gttcaacttt
     4321 attatacaaa gttggcatta taaaaaagca ttgcttatca atttgttgca acgaacaggt
     4381 cactatcagt caaaataaaa tcattattta atcaggtgca ctctcagtac aatctgctct
     4441 gatgccgcat agttaagcca gccccgacac ccgccaacac ccgctgacgc gccctgacgg
     4501 gcttgtctgc tcccggcatc cgcttacaga caagctgtga ccgtctccgg gagctgcatg
     4561 tgtcagaggt tttcaccgtc atcaccgaaa cgcgcga
//
