LOCUS       sequence-462698-        5135 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  A middle entry gateway clone containing V5 tagged human GATA4 and an
            H2A tagged mCherry, separated by a P2A peptide cleavage sequence..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5135)
  AUTHORS   Cantu Gutierrez ME, Hill MC, Largoza G, Martin JF, Wythe JD
  TITLE     Defining the Transcriptional and Epigenetic Basis of Organotypic 
            Endothelial Diversity in the Developing and Adult Mouse
  JOURNAL   bioRxiv 2021.11.15.468651
REFERENCE   2  (bases 1 to 5135)
  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: "bioRxiv 
            2021.11.15.468651"
COMMENT     Sequence Label: JDW 1059 (pME_V5_GATA4_P2A_H2A_mCherry)
FEATURES             Location/Qualifiers
     source          1..5135
                     /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    569..668
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     regulatory      675..684
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             684..725
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             2079..2135
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             2532..3242
                     /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(2677..2695)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2945..2964
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     3139..3162
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     protein_bind    complement(3260..3359)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(3376..3395)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3377..3395)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3400..3416)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3400..3416)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3518..3537)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             3529..4338
                     /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(3596..3615)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      4485..5073
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4974..4993
                     /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 tcgggcccca aataatgatt ttattttgac tgatagtgac
      601 ctgttcgttg caacaaattg atgagcaatg cttttttata atgccaactt tgtacaaaaa
      661 agcaggctgg atccgccacc atgggtaaac caatcccaaa tcccttgctg ggccttgact
      721 caacaggtgg tgctggtggt atgtaccaat ccctcgcgat ggcagctaat catggaccac
      781 cacctggagc gtatgaagcc ggcggacctg gtgcatttat gcatggtgcc ggagctgcta
      841 gttcccctgt ttatgtacca actccacgcg taccatcaag cgttcttggg ctttcttatt
      901 tgcaaggtgg cggggccggg agtgcaagcg gtggagctag tggcggatca tctggcggcg
      961 ctgctagcgg ggctggtcct ggaacacaac aagggtcccc aggctggagt caagctgggg
     1021 ctgatggtgc agcctatacg ccaccaccag tctcccctcg gtttagcttt cccggtacta
     1081 ctggtagcct cgccgcagct gctgctgccg cggctgctag agaggccgcc gcatattcat
     1141 caggtggtgg ggccgccggg gctggtctcg ctgggcgtga acaatatgga agagcggggt
     1201 ttgcagggtc ttatagttca ccttatcccg cctatatggc agatgtcgga gcctcatggg
     1261 ctgccgctgc agcagctagc gctggtccat ttgattctcc cgtgttgcat tccctgccag
     1321 ggagagcgaa tcccgcagcg aggcatccaa acctggacat gttcgatgat tttagcgagg
     1381 ggcgcgaatg cgtgaattgc ggagcgatgt caactccctt gtggcgtcgc gacggcacag
     1441 gacattactt gtgtaatgct tgtggactgt atcataaaat gaatggaata aatcgtccac
     1501 tgataaaacc ccaacgtcgc ttgtcagcga gtcggcgggt aggtctgagt tgcgcaaatt
     1561 gtcaaacaac tacaacaacc ttgtggagaa gaaacgccga aggggaacca gtctgtaacg
     1621 cttgtggact gtatatgaaa ctgcatggag ttcctcggcc gctggccatg cgtaaggaag
     1681 gcattcagac tcgcaagagg aaaccgaaga atctcaacaa gagcaaaact cctgccgcac
     1741 cgtccgggtc cgaaagtctg ccaccggcat caggagcatc aagtaattca tcaaatgcta
     1801 cgacatcttc ttctgaagaa atgcgaccta taaagactga accgggactt agttcccatt
     1861 atggacattc ttcatctgta agccaaacct ttagtgtgag cgccatgagt ggtcacggac
     1921 ctagcataca tccagttctt tccgcactca aactgagtcc ccagggatac gcctcaccgg
     1981 tgagtcaaag tccccaaact agttcaaaac aagatagttg gaatagtctt gtgctcgcag
     2041 attctcatgg tgacataatc actgcgggtg gtaccggagc cactaacttc tccctgttga
     2101 aacaagcagg ggatgtcgaa gagaatcccg ggccacctag gatggcaggt ggaaaagcag
     2161 gtaaagacag tggaaaagcc aagacgaaag ccgtctccag atctcagaga gctggtctgc
     2221 agttccctgt aggtcgtatc cacaggcacc tgaagacacg aaccaccagc catggccgtg
     2281 tcggagctac agctgcggtt tacagtgctg ctatcctgga gtatctcacc gctgaggtgc
     2341 tggagttggc tggaaacgct tctaaggatc tgaaggtgaa gcgaatcact cccagacatt
     2401 tacagctggc catccgcgga gacgaggagc tggactcgct tatcaaggcc accattgctg
     2461 gaggaggagt cattcctcac atccacaagt ctctgattgg taagaagggc cagcagaaaa
     2521 ccgcaggggg gatggtgagc aagggcgagg aggacaacat ggccatcatc aaggagttca
     2581 tgcgcttcaa ggtgcacatg gagggctccg tgaacggcca cgagttcgag atcgagggcg
     2641 agggcgaggg ccgcccctac gagggcaccc agaccgccaa gctgaaggtg accaagggcg
     2701 gccccctgcc cttcgcctgg gacatcctgt cccctcagtt catgtacggc tccaaggcct
     2761 acgtgaagca ccccgccgac atccccgact acttgaagct gtccttcccc gagggcttca
     2821 agtgggagcg cgtgatgaac ttcgaggacg gcggcgtggt gaccgtgacc caggactcct
     2881 ccctgcagga cggcgagttc atctacaagg tgaagctgcg cggcaccaac ttcccctccg
     2941 acggccccgt aatgcagaag aagaccatgg gctgggaggc ctcctccgag cggatgtacc
     3001 ccgaggacgg cgccctgaag ggcgagatca agcagaggct gaagctgaag gacggcggcc
     3061 actacgacgc cgaggtcaag accacctaca aggccaagaa gcccgtgcag ctgcccggcg
     3121 cctacaacgt caacatcaag ctggacatca cctcccacaa cgaggactac accatcgtgg
     3181 aacagtacga gcgcgccgag ggccgccact ccaccggcgg catggacgag ctgtacaagt
     3241 aatgatagct cgagagatct cccagctttc ttgtacaaag ttggcattat aagaaagcat
     3301 tgcttatcaa tttgttgcaa cgaacaggtc actatcagtc aaaataaaat cattatttgc
     3361 catccagctg atatccccta tagtgagtcg tattacatgg tcatagctgt ttcctggcag
     3421 ctctggcccg tgtctcaaaa tctctgatgt tacattgcac aagataaaaa tatatcatca
     3481 tgaacaataa aactgtctgc ttacataaac agtaatacaa ggggtgttat gagccatatt
     3541 caacgggaaa cgtcgaggcc gcgattaaat tccaacatgg atgctgattt atatgggtat
     3601 aaatgggctc gcgataatgt cgggcaatca ggtgcgacaa tctatcgctt gtatgggaag
     3661 cccgatgcgc cagagttgtt tctgaaacat ggcaaaggta gcgttgccaa tgatgttaca
     3721 gatgagatgg tcagactaaa ctggctgacg gaatttatgc ctcttccgac catcaagcat
     3781 tttatccgta ctcctgatga tgcatggtta ctcaccactg cgatccccgg aaaaacagca
     3841 ttccaggtat tagaagaata tcctgattca ggtgaaaata ttgttgatgc gctggcagtg
     3901 ttcctgcgcc ggttgcattc gattcctgtt tgtaattgtc cttttaacag cgatcgcgta
     3961 tttcgtctcg ctcaggcgca atcacgaatg aataacggtt tggttgatgc gagtgatttt
     4021 gatgacgagc gtaatggctg gcctgttgaa caagtctgga aagaaatgca taaacttttg
     4081 ccattctcac cggattcagt cgtcactcat ggtgatttct cacttgataa ccttattttt
     4141 gacgagggga aattaatagg ttgtattgat gttggacgag tcggaatcgc agaccgatac
     4201 caggatcttg ccatcctatg gaactgcctc ggtgagtttt ctccttcatt acagaaacgg
     4261 ctttttcaaa aatatggtat tgataatcct gatatgaata aattgcagtt tcatttgatg
     4321 ctcgatgagt ttttctaatc agaattggtt aattggttgt aacactggca gagcattacg
     4381 ctgacttgac gggacggcgc aagctcatga ccaaaatccc ttaacgtgag ttacgcgtcg
     4441 ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt
     4501 ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg
     4561 ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata
     4621 ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca
     4681 ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag
     4741 tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc
     4801 tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga
     4861 tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg
     4921 tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac
     4981 gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg
     5041 tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg
     5101 ttcctggcct tttgctggcc ttttgctcac atgtt
//
