LOCUS       sequence-462721-        5248 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  A middle entry gateway clone containing V5 tagged human GATA2 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 5248)
  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 5248)
  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 1051 (pME_V5_GATA2_P2A_H2A_mCherry)
FEATURES             Location/Qualifiers
     source          1..5248
                     /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             2192..2248
                     /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             2645..3355
                     /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(2790..2808)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     3058..3077
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     3252..3275
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     protein_bind    complement(3373..3472)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(3489..3508)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3490..3508)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3513..3529)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3513..3529)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3631..3650)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             3642..4451
                     /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(3709..3728)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      4598..5186
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5087..5106
                     /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 atctgccacc atgggtaagc ctatccctaa ccctctcctc ggtctcgatt
      721 ctacgggtgg tgctggtggt atggaggtgg cgccggagca gccgcgctgg atggcgcacc
      781 cggccgtgct gaatgcgcag caccccgact cacaccaccc gggcctggcg cacaactaca
      841 tggaacccgc gcagctgctg cctccagacg aggtggacgt cttcttcaat cacctcgact
      901 cgcagggcaa cccctactat gccaaccccg ctcacgcgcg ggcgcgcgtc tcctacagcc
      961 ccgcgcacgc ccgcctgacc ggaggccaga tgtgccgccc acacttgttg cacagcccgg
     1021 gtttgccctg gctggacggg ggcaaagcag ccctctctgc cgctgcggcc caccaccaca
     1081 acccctggac cgtgagcccc ttctccaaga cgccactgca cccctcagct gctggaggcc
     1141 ctggaggccc actctctgtg tacccagggg ctgggggtgg gagcggggga ggcagcggga
     1201 gctcagtggc ctccctcacc cctacagcaa cccactctgg ctcccacctt ttcggcttcc
     1261 cacccacgcc acccaaagaa gtgtctcctg accctagcac cacgggggct gcgtctccag
     1321 cctcatcttc cgcggggggt agtgcagccc gaggagagga caaggacggc gtcaagtacc
     1381 aggtgtcact gacggagagc atgaagatgg aaagtggcag tcccctgcgc ccaggcctag
     1441 ctactatggg cacccagcct gctacacacc accccatccc cacctacccc tcctatgtgc
     1501 cggcggctgc ccacgactac agcagcggac tcttccaccc cggaggcttc ctggggggac
     1561 cggcctccag cttcacccct aagcagcgca gcaaggctcg ttcctgttca gaaggccggg
     1621 agtgtgtcaa ctgtggggcc acagccaccc ctctctggcg gcgggacggc accggccact
     1681 acctgtgcaa tgcctgtggc ctctaccaca agatgaatgg gcagaaccga ccactcatca
     1741 agcccaagcg aagactgtcg gccgccagaa gagccggcac ctgttgtgca aattgtcaga
     1801 cgacaaccac caccttatgg cgccgaaacg ccaacgggga ccctgtctgc aacgcctgtg
     1861 gcctctacta caagctgcac aatgttaaca ggccactgac catgaagaag gaagggatcc
     1921 agactcggaa ccggaagatg tccaacaagt ccaagaagag caagaaaggg gcggagtgct
     1981 tcgaggagct gtcaaagtgc atgcaggaga agtcatcccc cttcagtgca gctgccctgg
     2041 ctggacacat ggcacctgtg ggccacctcc cgcccttcag ccactccgga cacatcctgc
     2101 ccactccgac gcccatccac ccctcctcca gcctctcctt cggccacccc cacccgtcca
     2161 gcatggtgac agccatgggc taggtaccgg agccactaac ttctccctgt tgaaacaagc
     2221 aggggatgtc gaagagaatc ccgggccacc taggatggca ggtggaaaag caggtaaaga
     2281 cagtggaaaa gccaagacga aagccgtctc cagatctcag agagctggtc tgcagttccc
     2341 tgtaggtcgt atccacaggc acctgaagac acgaaccacc agccatggcc gtgtcggagc
     2401 tacagctgcg gtttacagtg ctgctatcct ggagtatctc accgctgagg tgctggagtt
     2461 ggctggaaac gcttctaagg atctgaaggt gaagcgaatc actcccagac atttacagct
     2521 ggccatccgc ggagacgagg agctggactc gcttatcaag gccaccattg ctggaggagg
     2581 agtcattcct cacatccaca agtctctgat tggtaagaag ggccagcaga aaaccgcagg
     2641 ggggatggtg agcaagggcg aggaggacaa catggccatc atcaaggagt tcatgcgctt
     2701 caaggtgcac atggagggct ccgtgaacgg ccacgagttc gagatcgagg gcgagggcga
     2761 gggccgcccc tacgagggca cccagaccgc caagctgaag gtgaccaagg gcggccccct
     2821 gcccttcgcc tgggacatcc tgtcccctca gttcatgtac ggctccaagg cctacgtgaa
     2881 gcaccccgcc gacatccccg actacttgaa gctgtccttc cccgagggct tcaagtggga
     2941 gcgcgtgatg aacttcgagg acggcggcgt ggtgaccgtg acccaggact cctccctgca
     3001 ggacggcgag ttcatctaca aggtgaagct gcgcggcacc aacttcccct ccgacggccc
     3061 cgtaatgcag aagaagacca tgggctggga ggcctcctcc gagcggatgt accccgagga
     3121 cggcgccctg aagggcgaga tcaagcagag gctgaagctg aaggacggcg gccactacga
     3181 cgccgaggtc aagaccacct acaaggccaa gaagcccgtg cagctgcccg gcgcctacaa
     3241 cgtcaacatc aagctggaca tcacctccca caacgaggac tacaccatcg tggaacagta
     3301 cgagcgcgcc gagggccgcc actccaccgg cggcatggac gagctgtaca agtaatgata
     3361 gctcgagaga tctcccagct ttcttgtaca aagttggcat tataagaaag cattgcttat
     3421 caatttgttg caacgaacag gtcactatca gtcaaaataa aatcattatt tgccatccag
     3481 ctgatatccc ctatagtgag tcgtattaca tggtcatagc tgtttcctgg cagctctggc
     3541 ccgtgtctca aaatctctga tgttacattg cacaagataa aaatatatca tcatgaacaa
     3601 taaaactgtc tgcttacata aacagtaata caaggggtgt tatgagccat attcaacggg
     3661 aaacgtcgag gccgcgatta aattccaaca tggatgctga tttatatggg tataaatggg
     3721 ctcgcgataa tgtcgggcaa tcaggtgcga caatctatcg cttgtatggg aagcccgatg
     3781 cgccagagtt gtttctgaaa catggcaaag gtagcgttgc caatgatgtt acagatgaga
     3841 tggtcagact aaactggctg acggaattta tgcctcttcc gaccatcaag cattttatcc
     3901 gtactcctga tgatgcatgg ttactcacca ctgcgatccc cggaaaaaca gcattccagg
     3961 tattagaaga atatcctgat tcaggtgaaa atattgttga tgcgctggca gtgttcctgc
     4021 gccggttgca ttcgattcct gtttgtaatt gtccttttaa cagcgatcgc gtatttcgtc
     4081 tcgctcaggc gcaatcacga atgaataacg gtttggttga tgcgagtgat tttgatgacg
     4141 agcgtaatgg ctggcctgtt gaacaagtct ggaaagaaat gcataaactt ttgccattct
     4201 caccggattc agtcgtcact catggtgatt tctcacttga taaccttatt tttgacgagg
     4261 ggaaattaat aggttgtatt gatgttggac gagtcggaat cgcagaccga taccaggatc
     4321 ttgccatcct atggaactgc ctcggtgagt tttctccttc attacagaaa cggctttttc
     4381 aaaaatatgg tattgataat cctgatatga ataaattgca gtttcatttg atgctcgatg
     4441 agtttttcta atcagaattg gttaattggt tgtaacactg gcagagcatt acgctgactt
     4501 gacgggacgg cgcaagctca tgaccaaaat cccttaacgt gagttacgcg tcgttccact
     4561 gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg
     4621 taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc
     4681 aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata
     4741 ctgttcttct agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta
     4801 catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc
     4861 ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg
     4921 ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac
     4981 agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg
     5041 taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt
     5101 atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct
     5161 cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg
     5221 ccttttgctg gccttttgct cacatgtt
//
