LOCUS       sequence-353467-        7273 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid containing piggyBac ITRs flanking CAG and mCherry (also see 
            doi: 10.7554/eLife.63907 ).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7273)
  AUTHORS   Yamagata M, Sanes JR
  TITLE     CRISPR-mediated Labeling of Cells in Chick Embryos Based on 
            Selectively Expressed Genes.
  JOURNAL   Bio Protoc. 2021 Aug 5;11(15):e4105. doi: 10.21769/BioProtoc.4105. 
            eCollection 2021 Aug 5.
  PUBMED    34458399
REFERENCE   2  (bases 1 to 7273)
  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: "Bio 
            Protoc."; date: "2021-08-5"; volume: "11(15):e4105. doi"; pages: " 
            10.21769/BioProtoc.4105. eCollection 2021 Aug 5"
COMMENT     Sequence Label: pXL-BacII-CAG-mCherry
FEATURES             Location/Qualifiers
     source          1..7273
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..304
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        306..584
                     /label=chicken beta-actin promoter
     intron          585..1601
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1525..1547
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1609..1628
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     protein_bind    1668..1692
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(1753..1786)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     regulatory      1912..1921
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1918..2628
                     /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(2063..2081)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2331..2350
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     protein_bind    complement(2671..2695)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(2786..2805)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    2851..2906
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(2905..2924)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     repeat_region   3529..3563
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(4054..4070)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4054..4070)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4067..4089)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4078..4094
                     /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(4102..4108,4109..4126,4127..4132))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4147..4168
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4285..4302)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4456..5044)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4536..4555)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5215..6006,6007..6075))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5838..5857
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6076..6180)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(6206..6661)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6343..6364)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6555..6574
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6788..6810
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     6802..6819
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6803..6819
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     repeat_region   6949..7011
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
ORIGIN
        1 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
       61 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      121 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      181 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      241 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      301 catggtcgag gtgagcccca cgttctgctt cactctcccc atctcccccc cctccccacc
      361 cccaattttg tatttattta ttttttaatt attttgtgca gcgatggggg cggggggggg
      421 gggggggggc gcgcccgggc ggggcggggc ggggcgaggg gcggggcggg gcgaggcgga
      481 gaggtgcggc ggcagccaat cagagcggcg cgctccgaaa gtttcctttt atggcgaggc
      541 ggcggcggcg gcggccctat aaaaagcgaa gcgcgcggcg ggcgggagtc gctgcgcgct
      601 gccttcgccc cgtgccccgc tccgccgccg cctcgcgccg cccgccccgg ctctgactga
      661 ccgcgttact cccacaggtg agcgggcggg acggcccttc tcctccgggc tgtaattagc
      721 gcttggttta atgacggctt gtttcttttc tgtggctgcg tgaaagcctt gaggggctcc
      781 gggagggccc tttgtgcggg gggagcggct cggggggtgc gtgcgtgtgt gtgtgcgtgg
      841 ggagcgccgc gtgcggctcc gcgctgcccg gcggctgtga gcgctgcggg cgcggcgcgg
      901 ggctttgtgc gctccgcagt gtgcgcgagg ggagcgcggc cgggggcggt gccccgcggt
      961 gcgggggggg ctgcgagggg aacaaaggct gcgtgcgggg tgtgtgcgtg ggggggtgag
     1021 cagggggtgt gggcgcgtcg gtcgggctgc aaccccccct gcacccccct ccccgagttg
     1081 ctgagcacgg cccggcttcg ggtgcggggc tccgtacggg gcgtggcgcg gggctcgccg
     1141 tgccgggcgg ggggtggcgg caggtggggg tgccgggcgg ggcggggccg cctcgggccg
     1201 gggagggctc gggggagggg cgcggcggcc cccggagcgc cggcggctgt cgaggcgcgg
     1261 cgagccgcag ccattgcctt ttatggtaat cgtgcgagag ggcgcaggga cttcctttgt
     1321 cccaaatctg tgcggagccg aaatctggga ggcgccgccg caccccctct agcgggcgcg
     1381 gggcgaagcg gtgcggcgcc ggcaggaagg aaatgggcgg ggagggcctt cgtgcgtcgc
     1441 cgcgccgccg tccccttctc cctctccagc ctcggggctg tccgcggggg gacggctgcc
     1501 ttcggggggg acggggcagg gcggggttcg gcttctggcg tgtgaccggc ggctctagag
     1561 cctctgctaa ccatgttcat gccttcttct ttttcctaca gctcctgggc aacgtgctgg
     1621 ttattgtgct gtctcatcat tttggcaaag aattcctcga ggatatcaca agtttgtaca
     1681 aaaaagcagg ctccgaattc gcccttcggg ccgcgcttaa ttaagctagc gctaccggac
     1741 tcagatctcg agataacttc gtatagcata cattatacga agttataccg gcagatctcc
     1801 gccaccatgc tgtgctgcat cagaagaact aaaccggttg agaagaatga agaggccgat
     1861 caggagctgc agtcgacggt accgcgggcc cgggatccac cggtagcatc cgccaccatg
     1921 gtgagcaagg gcgaggagga taacatggcc atcatcaagg agttcatgcg cttcaaggtg
     1981 cacatggagg gctccgtgaa cggccacgag ttcgagatcg agggcgaggg cgagggccgc
     2041 ccctacgagg gcacccagac cgccaagctg aaggtgacca agggtggccc cctgcccttc
     2101 gcctgggaca tcctgtcccc tcagttcatg tacggctcca aggcctacgt gaagcacccc
     2161 gccgacatcc ccgactactt gaagctgtcc ttccccgagg gcttcaagtg ggagcgcgtg
     2221 atgaacttcg aggacggcgg cgtggtgacc gtgacccagg actcctccct gcaggacggc
     2281 gagttcatct acaaggtgaa gctgcgcggc accaacttcc cctccgacgg ccccgtaatg
     2341 cagaagaaga ccatgggctg ggaggcctcc tccgagcgga tgtaccccga ggacggcgcc
     2401 ctgaagggcg agatcaagca gaggctgaag ctgaaggacg gcggccacta cgacgctgag
     2461 gtcaagacca cctacaaggc caagaagccc gtgcagctgc ccggcgccta caacgtcaac
     2521 atcaagttgg acatcacctc ccacaacgag gactacacca tcgtggaaca gtacgaacgc
     2581 gccgagggcc gccactccac cggcggcatg gacgagctgt acaagtaaac acgtgcggcc
     2641 gcgactctag atcataaaag ggcgaattcg acccagcttt cttgtacaaa gtggtgattc
     2701 gcggccgcac tcctcaggtg caggctgcct atcagaaggt ggtggctggt gtggccaatg
     2761 ccctggctca caaataccac tgagatcttt ttccctctgc caaaaattat ggggacatca
     2821 tgaagcccct tgagcatctg acttctggct aataaaggaa atttattttc attgcaatag
     2881 tgtgttggaa ttttttgtgt ctctcactcg gaaggacata tgggagggca aatcatttaa
     2941 aacatcagaa tgagtatttg gtttagagtt tggcaacata tgcccatatg ctggctgcca
     3001 tgaacaaagg ttggctataa agaggtcatc agtatatgaa acagccccct gctgtccatt
     3061 ccttattcca tagaaaagcc ttgacttgag gttagatttt ttttatattt tgttttgtgt
     3121 tatttttttc tttaacatcc ctaaaatttt ccttacatgt tttactagcc agatttttcc
     3181 tcctctcctg actactccca gtcatagctg tccctcttct cttatggaga tccctcgacc
     3241 tgcagcccaa gcttgatatc tataacaaga aaatatatat ataataagtt atcacgtaag
     3301 tagaacatga aataacaata taattatcgt atgagttaaa tcttaaaagt cacgtaaaag
     3361 ataatcatgc gtcattttga ctcacgcggt cgttatagtt caaaatcagt gacacttacc
     3421 gcattgacaa gcacgcctca cgggagctcc aagcggcgac tgagatgtcc taaatgcaca
     3481 gcgacggatt cgcgctattt agaaagagag agcaatattt caagaatgca tgcgtcaatt
     3541 ttacgcagac tatctttcta gggttaatct agctgcatca ggatcatatc gtcgggtctt
     3601 ttttccggct cagtcatcgc ccaagctggc gctatctggg catcggggag gaagaagccc
     3661 gtgccttttc ccgcgaggtt gaagcggcat ggaaagagtt tgccgaggat gactgctgct
     3721 gcattgacgt tgagcgaaaa cgcacgttta ccatgatgat tcgggaaggt gtggccatgc
     3781 acgcctttaa cggtgaactg ttcgttcagg ccacctggga taccagttcg tcgcggcttt
     3841 tccggacaca gttccggatg gtcagcccga agcgcatcag caacccgaac aataccggcg
     3901 acagccggaa ctgccgtgcc ggtgtgcaga ttaatgacag cggtgcggcg ctgggatatt
     3961 acgtcagcga ggacgggtat cctggctgga tgccgcagaa atggacatgg ataccccgtg
     4021 agttacccgg cgggcgcgct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg
     4081 ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg
     4141 tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc
     4201 gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt
     4261 gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct
     4321 gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga
     4381 taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc
     4441 cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg
     4501 ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg
     4561 aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt
     4621 tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt
     4681 gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg
     4741 cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact
     4801 ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt
     4861 cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct
     4921 gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac
     4981 cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc
     5041 tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg
     5101 ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta
     5161 aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca
     5221 atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc
     5281 ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc
     5341 tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc
     5401 agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat
     5461 taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt
     5521 tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc
     5581 cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag
     5641 ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt
     5701 tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac
     5761 tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg
     5821 cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat
     5881 tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc
     5941 gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc
     6001 tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa
     6061 atgttgaata ctcatactct tcctttttca atattattga agcatttatc agggttattg
     6121 tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg
     6181 cacatttccc cgaaaagtgc cacctaaatt gtaagcgtta atattttgtt aaaattcgcg
     6241 ttaaattttt gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct
     6301 tataaatcaa aagaatagac cgagataggg ttgagtgttg ttccagtttg gaacaagagt
     6361 ccactattaa agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat
     6421 ggcccactac gtgaaccatc accctaatca agttttttgg ggtcgaggtg ccgtaaagca
     6481 ctaaatcgga accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac
     6541 gtggcgagaa aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta
     6601 gcggtcacgc tgcgcgtaac caccacaccc gccgcgctta atgcgccgct acagggcgcg
     6661 tcccattcgc cattcaggct gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg
     6721 ctattacgcc agctggcgaa agggggatgt gctgcaaggc gattaagttg ggtaacgcca
     6781 gggttttccc agtcacgacg ttgtaaaacg acggccagtg agcgcgcctc gttcattcac
     6841 gtttttgaac ccgtggagga cgggcagact cgcggtgcaa atgtgtttta cagcgtgatg
     6901 gagcagatga agatgctcga cacgctgcag aacacgcagc tagattaacc ctagaaagat
     6961 aatcatattg tgacgtacgt taaagataat catgtgtaaa attgacgcat gtgttttatc
     7021 ggtctgtata tcgaggttta tttattaatt tgaatagata ttaagtttta ttatatttac
     7081 acttacatac taataataaa ttcaacaaac aatttattta tgtttattta tttattaaaa
     7141 aaaacaaaaa ctcaaaattt cttctataaa gtaacaaaac ttttatcgaa ttcctgcagc
     7201 ccgggggatc cactagttat taatagtaat caattacggg gtcattagtt catagcccat
     7261 atatggagtt ccg
//
