LOCUS       sequence-462699-        5138 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  A middle entry gateway clone containing V5 tagged human FOXF2 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 5138)
  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 5138)
  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 1060 (pME_V5_FOXF2_P2A_H2A_mCherry)
FEATURES             Location/Qualifiers
     source          1..5138
                     /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             1536..1556
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=7xHis
                     /translation="HHHHHHH"
     CDS             2082..2138
                     /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             2535..3245
                     /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(2680..2698)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2948..2967
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     3142..3165
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     protein_bind    complement(3263..3362)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(3379..3398)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3380..3398)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3403..3419)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3403..3419)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3521..3540)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             3532..4341
                     /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(3599..3618)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      4488..5076
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4977..4996
                     /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 atgggaaaac ctataccaaa tcccttgctg ggcctggact
      721 caacaggtgg tgctggtggt atgacaacag aaggtggacc cccacccgct cctctgagaa
      781 gagcttgtag tccagtgcca ggggccttgc aagcagctct catgtctcct cctccagcgg
      841 cagctgcagc agcggctgca gcacccgaaa ctactagctc ttcaagtagc agctcaagcg
      901 catcatgtgc tagtagctcc agtagcagca acagtgcttc tgctccgtcc gcggcatgta
      961 aatcagctgg cggaggaggt gcaggagcag gatctggtgg ggctaagaag gcctcttccg
     1021 gcctcagaag accagaaaag cccccttata gctatatagc actgatagta atggcgattc
     1081 aatccagccc atccaaacgt ctcaccctga gtgaaattta tcaatttctg caagctcggt
     1141 ttcctttctt tcgaggtgct tatcaaggtt ggaagaattc agtcagacat aacctgagtc
     1201 ttaatgaatg ttttattaaa cttccaaaag ggctgggccg tcctgggaaa ggtcattatt
     1261 ggacaattga tcccgcatcc gaatttatgt ttgaagaagg atcatttcgg cggcgcccca
     1321 ggggatttcg gagaaaatgt caagctctga aacctatgta tcatcgggtt gtctctgggc
     1381 tgggatttgg cgcaagcttg ctccctcaag ggtttgattt tcaagcccca ccttctgctc
     1441 ccctggggtg tcattctcaa ggtggatatg gcggtctgga tatgatgcca gctggttatg
     1501 atgcaggggc aggtgcgcca tcccatgctc atccccatca tcatcatcat catcatgtgc
     1561 cacatatgtc ccctaatcca ggaagcacgt atatggcaag ttgtcccgtc cctgctggcc
     1621 caggcggagt tggggcagct ggtggtggcg gggggggtga ttatggacca gattctagct
     1681 cttcccctgt gccttcctct ccagcaatgg cttctgcgat agagtgtcat agtccttata
     1741 caagtccagc tgctcattgg tcctcacctg gggcgtcccc gtatcttaaa caacccccag
     1801 ctctcacacc tagttccaat cctgctgcat ccgcgggatt gcatagttcc atgagttctt
     1861 attcactcga acaatcatat cttcatcaaa atgcacggga agatctgtcc gttgggcttc
     1921 ctagatatca acaccatagt acacctgttt gcgatcggaa ggactttgtt ctgaatttta
     1981 acggaatctc ctcatttcac ccgagcgcat ccggttccta ctaccatcac catcatcaat
     2041 cagtgtgcca agacatcaaa ccttgtgtga tgggtaccgg agccactaac ttctccctgt
     2101 tgaaacaagc aggggatgtc gaagagaatc ccgggccacc taggatggca ggtggaaaag
     2161 caggtaaaga cagtggaaaa gccaagacga aagccgtctc cagatctcag agagctggtc
     2221 tgcagttccc tgtaggtcgt atccacaggc acctgaagac acgaaccacc agccatggcc
     2281 gtgtcggagc tacagctgcg gtttacagtg ctgctatcct ggagtatctc accgctgagg
     2341 tgctggagtt ggctggaaac gcttctaagg atctgaaggt gaagcgaatc actcccagac
     2401 atttacagct ggccatccgc ggagacgagg agctggactc gcttatcaag gccaccattg
     2461 ctggaggagg agtcattcct cacatccaca agtctctgat tggtaagaag ggccagcaga
     2521 aaaccgcagg ggggatggtg agcaagggcg aggaggacaa catggccatc atcaaggagt
     2581 tcatgcgctt caaggtgcac atggagggct ccgtgaacgg ccacgagttc gagatcgagg
     2641 gcgagggcga gggccgcccc tacgagggca cccagaccgc caagctgaag gtgaccaagg
     2701 gcggccccct gcccttcgcc tgggacatcc tgtcccctca gttcatgtac ggctccaagg
     2761 cctacgtgaa gcaccccgcc gacatccccg actacttgaa gctgtccttc cccgagggct
     2821 tcaagtggga gcgcgtgatg aacttcgagg acggcggcgt ggtgaccgtg acccaggact
     2881 cctccctgca ggacggcgag ttcatctaca aggtgaagct gcgcggcacc aacttcccct
     2941 ccgacggccc cgtaatgcag aagaagacca tgggctggga ggcctcctcc gagcggatgt
     3001 accccgagga cggcgccctg aagggcgaga tcaagcagag gctgaagctg aaggacggcg
     3061 gccactacga cgccgaggtc aagaccacct acaaggccaa gaagcccgtg cagctgcccg
     3121 gcgcctacaa cgtcaacatc aagctggaca tcacctccca caacgaggac tacaccatcg
     3181 tggaacagta cgagcgcgcc gagggccgcc actccaccgg cggcatggac gagctgtaca
     3241 agtaatgata gctcgagaga tctcccagct ttcttgtaca aagttggcat tataagaaag
     3301 cattgcttat caatttgttg caacgaacag gtcactatca gtcaaaataa aatcattatt
     3361 tgccatccag ctgatatccc ctatagtgag tcgtattaca tggtcatagc tgtttcctgg
     3421 cagctctggc ccgtgtctca aaatctctga tgttacattg cacaagataa aaatatatca
     3481 tcatgaacaa taaaactgtc tgcttacata aacagtaata caaggggtgt tatgagccat
     3541 attcaacggg aaacgtcgag gccgcgatta aattccaaca tggatgctga tttatatggg
     3601 tataaatggg ctcgcgataa tgtcgggcaa tcaggtgcga caatctatcg cttgtatggg
     3661 aagcccgatg cgccagagtt gtttctgaaa catggcaaag gtagcgttgc caatgatgtt
     3721 acagatgaga tggtcagact aaactggctg acggaattta tgcctcttcc gaccatcaag
     3781 cattttatcc gtactcctga tgatgcatgg ttactcacca ctgcgatccc cggaaaaaca
     3841 gcattccagg tattagaaga atatcctgat tcaggtgaaa atattgttga tgcgctggca
     3901 gtgttcctgc gccggttgca ttcgattcct gtttgtaatt gtccttttaa cagcgatcgc
     3961 gtatttcgtc tcgctcaggc gcaatcacga atgaataacg gtttggttga tgcgagtgat
     4021 tttgatgacg agcgtaatgg ctggcctgtt gaacaagtct ggaaagaaat gcataaactt
     4081 ttgccattct caccggattc agtcgtcact catggtgatt tctcacttga taaccttatt
     4141 tttgacgagg ggaaattaat aggttgtatt gatgttggac gagtcggaat cgcagaccga
     4201 taccaggatc ttgccatcct atggaactgc ctcggtgagt tttctccttc attacagaaa
     4261 cggctttttc aaaaatatgg tattgataat cctgatatga ataaattgca gtttcatttg
     4321 atgctcgatg agtttttcta atcagaattg gttaattggt tgtaacactg gcagagcatt
     4381 acgctgactt gacgggacgg cgcaagctca tgaccaaaat cccttaacgt gagttacgcg
     4441 tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt
     4501 tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt
     4561 ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag
     4621 ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa gaactctgta
     4681 gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat
     4741 aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg
     4801 ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg
     4861 agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac
     4921 aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga
     4981 aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt
     5041 ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta
     5101 cggttcctgg ccttttgctg gccttttgct cacatgtt
//
