LOCUS       sequence-239422-        6424 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Donor cassette containing myr-tagged tdTomato and SV40 polyA tail, 
            followed by FRT site-flanked Kanamycin selection gene.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6424)
  AUTHORS   Weinberger M, Simoes FC, Patient R, Sauka-Spengler T, Riley PR
  TITLE     Functional Heterogeneity within the Developing Zebrafish Epicardium.
  JOURNAL   Dev Cell. 2020 Mar 9;52(5):574-590.e6. doi: 
            10.1016/j.devcel.2020.01.023. Epub 2020 Feb 20.
  PUBMED    32084358
REFERENCE   2  (bases 1 to 6424)
  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"; doi: 
            "10.1016/j.devcel.2020.01.023"; journalName: "Dev Cell"; date: 
            "2020-03-9- 9"; volume: "52"; issue: "5"; pages: "574-590.e6"
COMMENT     Sequence Label: pGEM-myr-tdTomato-SV40pA-FKF
FEATURES             Location/Qualifiers
     source          1..6424
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(28..456)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(138..159)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     350..369
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     582..604
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     596..613
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     597..613
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     620..639
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        620..638
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      669..678
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      673..682
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             697..723
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YDVPDYASL"
     CDS             730..756
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YDVPDYASL"
     CDS             763..789
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YDVPDYASL"
     regulatory      853..862
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             931..2361
                     /codon_start=1
                     /product="tandem dimeric (pseudo-monomeric) derivative of 
                     DsRed (Shaner et al., 2004)"
                     /label=tdTomato
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVMNFEDGG
                     LVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVLKGEIH
                     QALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERSEGRH
                     HLFLGHGTGSTGSGSSGTASSEDNNMAVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPY
                     EGTQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVM
                     NFEDGGLVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGV
                     LKGEIHQALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYE
                     RSEGRHHLFLYGMDELYK"
     polyA_signal    2413..2534
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2424..2443)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2478..2497
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     protein_bind    complement(2658..2705)
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     promoter        2799..2903
                     /gene="bla"
                     /label=AmpR promoter
     enhancer        2905..3095
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2928..2948)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     CDS             3135..3929
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             3135..3929
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3189..3208)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3799..3818
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3936..3984
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     protein_bind    complement(4009..4056)
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     promoter        complement(4185..4203)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(4186..4203)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(4221..4237)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4221..4237)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4234..4256)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4245..4261
                     /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)."
     promoter        complement(join(4269..4275,4276..4293,4294..4299))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4314..4335
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4452..4469)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4623..5211)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4703..4722)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5382..6173,6174..6242))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGKRGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6005..6024
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6243..6347)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat
       61 tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga
      121 tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca
      181 acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccct
      241 aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc
      301 cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag
      361 cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca
      421 cacccgccgc gcttaatgcg ccgctacagg gcgcgtccat tcgccattca ggctgcgcaa
      481 ctgttgggaa gggcgatcgg tgcgggcctc ttcgctatta cgccagctgg cgaaaggggg
      541 atgtgctgca aggcgattaa gttgggtaac gccagggttt tcccagtcac gacgttgtaa
      601 aacgacggcc agtgaattgt aatacgactc actatagggc gaattgggcc cgacgtcgca
      661 tgctcccggc cgccatggcg gccgcggatg gccacctatg acgtcccaga ttacgcaagt
      721 ttgcccggat atgatgttcc tgattatgct agcctcccgg gttacgatgt gcccgactat
      781 gcctcacttc caggcagatc tcctatgggc tgcatcaaga gcaagcgcaa ggacaacctg
      841 aacgacgacg aggccgccat gggctgcatc aagagcaagc gcaaggacaa cctgaacgac
      901 gacgagggtg gaagcggatc tggaggtggg atggtgtcga agggcgagga ggtcatcaaa
      961 gagttcatgc gcttcaaagt tcgcatggag ggctccatga acggccacga gttcgagatc
     1021 gagggcgagg gcgagggccg cccctacgag ggcacccaga ccgccaagct gaaggtgacc
     1081 aagggcggcc ccctgccctt cgcctgggac atcctgtccc cccaattcat gtacggctcc
     1141 aaggcgtacg tgaagcaccc cgccgacatc cccgattaca agaagctgtc cttccccgag
     1201 ggcttcaagt gggagcgcgt gatgaacttc gaggacggcg gtctggtgac cgtgacccag
     1261 gactcctccc tgcaagacgg cacgctgatc tacaaggtga agatgcgcgg caccaacttc
     1321 ccccccgacg gccccgtaat gcagaagaag accatgggct gggaggcctc caccgagcgc
     1381 ctgtaccccc gcgacggcgt gctgaagggc gagatccacc aggccctgaa gctgaaggac
     1441 ggcggccact acctggtgga gttcaagacc atctacatgg ccaagaagcc cgtgcaactg
     1501 cccggctact actacgtgga caccaagctg gacatcacct cccacaacga ggactacacc
     1561 atcgtggaac agtacgagcg ctccgagggc cgccaccacc tgttcctggg gcatggcacc
     1621 ggcagcaccg gcagcggcag ctccggcacc gcctcctccg aggacaacaa catggccgtc
     1681 atcaaagagt tcatgcgctt caaagttcgc atggagggct ccatgaacgg ccacgagttc
     1741 gagatcgagg gcgagggcga gggccgcccc tacgagggca cccagaccgc caagctgaag
     1801 gtgaccaagg gcggccccct gcccttcgcc tgggacatcc tgtcccccca attcatgtac
     1861 ggctccaagg cgtacgtgaa gcaccccgcc gacatccccg attacaagaa gctgtccttc
     1921 cccgagggct tcaagtggga gcgcgtgatg aacttcgagg acggcggtct ggtgaccgtg
     1981 acccaggact cctccctgca agacggcacg ctgatctaca aggtgaagat gcgcggcacc
     2041 aacttccccc ccgacggccc cgtaatgcag aagaagacca tgggctggga ggcctccacc
     2101 gagcgcctgt acccccgcga cggcgtgctg aagggcgaga tccaccaggc cctgaagctg
     2161 aaggacggcg gccactacct ggtggagttc aagaccatct acatggccaa gaagcccgtg
     2221 caactgcccg gctactacta cgtggacacc aagctggaca tcacctccca caacgaggac
     2281 tacaccatcg tggaacagta cgagcgctcc gagggccgcc accacctgtt cctgtacggc
     2341 atggacgagc tgtacaagta aagcggccgc gactctagag tcggggcggc cggccgcttc
     2401 gagcagacat gataagatac attgatgagt ttggacaaac cacaactaga atgcagtgaa
     2461 aaaaatgctt tatttgtgaa atttgtgatg ctattgcttt atttgtaacc attataagct
     2521 gcaataaaca agttaacaac aacaattgca ttcattttat gtttcgggtt cagggggagg
     2581 tgtgggaggt tttttaaagc aagtaaaacc tctacaaatg tggtaaaatc gataaggatc
     2641 cgtcgaccga cctgcttgaa gttcctatac tttctagaga ataggaactt cggaatagga
     2701 acttcaagat ccccctggcg aaagggggat gtgctgcaag gcgattaagt tgggtaacgc
     2761 cagggttttc gtcaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat
     2821 ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc
     2881 aataatattg aaaaaggaag agtcctgagg cggaaagaac cagctgtgga atgtgtgtca
     2941 gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     3001 caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     3061 aagcatgcat ctcaattagt cagtaaccat agtccgcaaa gatcgatcaa gagacaggat
     3121 gaggatcgtt tcgcatgatt gaacaagatg gattgcacgc aggttctccg gccgcttggg
     3181 tggagaggct attcggctat gactgggcac aacagacaat cggctgctct gatgccgccg
     3241 tgttccggct gtcagcgcag gggcgcccgg ttctttttgt caagaccgac ctgtccggtg
     3301 ccctgaatga actgcaagac gaggcagcgc ggctatcgtg gctggccacg acgggcgttc
     3361 cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag ggactggctg ctattgggcg
     3421 aagtgccggg gcaggatctc ctgtcatctc accttgctcc tgccgagaaa gtatccatca
     3481 tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc tacctgccca ttcgaccacc
     3541 aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga agccggtctt gtcgatcagg
     3601 atgatctgga cgaagagcat caggggctcg cgccagccga actgttcgcc aggctcaagg
     3661 cgagcatgcc cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc ttgccgaata
     3721 tcatggtgga aaatggccgc ttttctggat tcatcgactg tggccggctg ggtgtggcgg
     3781 accgctatca ggacatagcg ttggctaccc gtgatattgc tgaagagctt ggcggcgaat
     3841 gggctgaccg cttcctcgtg ctttacggta tcgccgctcc cgattcgcag cgcatcgcct
     3901 tctatcgcct tcttgacgag ttcttctgag gggatcggca ataaaaagac agaataaaac
     3961 gcacgggtgt tgggtcgttt gttcggatcc gagcttcaaa agcgctctga agttcctata
     4021 ctttctagag aataggaact tcggaatagt aacttctcca tggtagcctc caaaaaagcc
     4081 tcctcactac ttctggaata gctcaatcac tagtgaattc gcggccgcct gcaggtcgac
     4141 catatgggag agctcccaac gcgttggatg catagcttga gtattctata gtgtcaccta
     4201 aatagcttgg cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca
     4261 attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg
     4321 agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg
     4381 tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc
     4441 tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta
     4501 tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag
     4561 aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg
     4621 tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg
     4681 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg
     4741 cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga
     4801 agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc
     4861 tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt
     4921 aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact
     4981 ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg
     5041 cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt
     5101 accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt
     5161 ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct
     5221 ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg
     5281 gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt
     5341 aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt
     5401 gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc
     5461 gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg
     5521 cgagacccac gcttaccggc tccagattta tcagcaataa accagccagc cggaagggcc
     5581 gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg
     5641 gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca
     5701 ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga
     5761 tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct
     5821 ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg
     5881 cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca
     5941 accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata
     6001 cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct
     6061 tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact
     6121 cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa
     6181 acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc
     6241 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga
     6301 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga
     6361 aaagtgccac ctgatgcggt gtgaaatacc gcacagatgc gtaaggagaa aataccgcat
     6421 cagg
//
