LOCUS       sequence-366082-        7172 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Used to generate the Tg(QUAS:mApple; he1.1:CFP)c788 transgenic line.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7172)
  AUTHORS   Spikol ED, Cheng J, Macurak M, Subedi A, Halpern ME
  TITLE     Genetically defined nucleus incertus neurons differ in connectivity 
            and function.
  JOURNAL   Elife. 2024 May 31;12:RP89516. doi: 10.7554/eLife.89516.
  PUBMED    38819436
REFERENCE   2  (bases 1 to 7172)
  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: "Elife."; 
            date: "2024-05-31"; volume: "12:RP89516. doi"; pages: " 
            10.7554/eLife.89516"
COMMENT     Sequence Label: pDestTol2-QUAS:mApple-he1.1:CFP
FEATURES             Location/Qualifiers
     source          1..7172
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(144..165)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     misc_feature    join(514..525,526..688)
                     /label=Tol2 right end
                     /note="right end of the Tol2 transposon from the medaka 
                     fish Oryzias latipes (Kawakami, 2007)"
     primer_bind     1061..1077
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    1097..1117
                     /gene="mutant version of attB"
                     /label=attB4
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
     protein_bind    1308..1332
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             1338..2048
                     /codon_start=1
                     /product="photostable monomeric derivative of DsRed (Shaner
                     et al., 2008)"
                     /label=mApple
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEENNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEA
                     FQTAKLKVTKGGPLPFAWDILSPQFMYGSKVYIKHPADIPDYFKLSFPEGFRWERVMNF
                     EDGGIIHVNQDSSLQDGVFIYKVKLRGTNFPSDGPVMQKKTMGWEASEERMYPEDGALK
                     SEIKKRLKLKDGGHYAAEVKTTYKAKKPVQLPGAYIVDIKLDIVSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     1751..1770
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     protein_bind    complement(2050..2074)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     polyA_signal    2074..2208
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     regulatory      2749..2758
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2755..2757,2758..2760,2761..3474)
                     /codon_start=1
                     /product="enhanced CFP"
                     /label=ECFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2800..2821)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3061..3080)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3408..3429
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(3837..3854)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3843..4068
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    complement(4069..4089)
                     /gene="mutant version of attB"
                     /label=attB3
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
     primer_bind     complement(4097..4113)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     polyA_signal    4129..4263
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     misc_feature    join(4702..4889,4890..4901)
                     /label=Tol2 left end
                     /note="left end of the Tol2 transposon from the medaka fish
                     Oryzias latipes (Kawakami, 2007)"
     primer_bind     complement(4935..4951)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4935..4951)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(4988..5008)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(4988..5006)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5027..5043)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5040..5062)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5051..5067
                     /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(5075..5081,5082..5099,5100..5105))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5120..5141
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5258..5275)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5429..6017)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5509..5528)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6188..6979,6980..7048))
                     /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     6811..6830
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7049..7153)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ccacctaaat tgtaagcgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca
       61 gctcattttt taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga
      121 ccgagatagg gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg
      181 actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacgc tggctaagaa
      241 ctcatcagcc tccccggtcc atctacccac gtaccaatgc accaattggc cacaatgacg
      301 gctactacat ggtgccattc cttcctcttt ataggaatgg agactacctc ctgtccaaca
      361 atgctcttgg atacgagtac gcctacctgt tggacccagg tcattgcaca acaccagaaa
      421 tgccctctga tctgcaaaag acgtgaatat ctgttcagac acccatatcc actctgttcc
      481 acacaggtca gaggtttgtc caggagttct tgacagaggt gtaaaaagta ctcaaaaatt
      541 ttactcaagt gaaagtacaa gtacttaggg aaaattttac tcaattaaaa gtaaaagtat
      601 ctggctagaa tcttacttga gtaaaagtaa aaaagtactc cattaaaatt gtacttgagt
      661 attaaggaag taaaagtaaa agcaagaaag aaaactagag attcttgttt aagcttttaa
      721 tctcaaaaaa cattaaatga aatgcataca aggttttatc ctgctttaga actgtttgta
      781 tttaattatc aaactataag acagacaatc taatgccagt acacgctact caaagttgta
      841 aaacctcaga tttaacttca gtagaagctg attctcaaaa ttgttagtgt caagcctagc
      901 tcttttgggg ctgaaaagca atcctgcagt gctgaaaagc ctctcacagg cagccgatgc
      961 gggaagaggt gtattagtct tgatagagag gctgcaaata gcaggaaacg tgagcagaga
     1021 ctccctggtg tctgaaacac aggccagatg ggccctcgag caggaaacag ctatgaccat
     1081 gattacgcca agctatcaac tttgtataga aaagttgggg taatcgctta tcctcggata
     1141 aacaattatc ctcacgggta atcgcttatc cgctcgggta atcgcttatc ctcgggtaat
     1201 cgcttatcct tgcaagggtc gactctagag ggtatataat ggatcccatc gcgtctcagc
     1261 ctcactttga gctcctccac acgaattccc tcgacctcga agacgcgtca agtttgtaca
     1321 aaaaagcagg ctggaccatg gtgagcaagg gcgaggagaa taacatggcc atcatcaagg
     1381 agttcatgcg cttcaaggtg cacatggagg gctccgtgaa cggccacgag ttcgagatcg
     1441 agggcgaggg cgagggccgc ccctacgagg cctttcagac cgctaagctg aaggtgacca
     1501 agggtggccc cctgcccttc gcctgggaca tcctgtcccc tcagttcatg tacggctcca
     1561 aggtctacat taagcaccca gccgacatcc ccgactactt caagctgtcc ttccccgagg
     1621 gcttcaggtg ggagcgcgtg atgaacttcg aggacggcgg cattattcac gttaaccagg
     1681 actcctccct gcaggacggc gtgttcatct acaaggtgaa gctgcgcggc accaacttcc
     1741 cctccgacgg ccccgtaatg cagaagaaga ccatgggctg ggaggcctcc gaggagcgga
     1801 tgtaccccga ggacggcgcc ctgaagagcg agatcaagaa gaggctgaag ctgaaggacg
     1861 gcggccacta cgccgccgag gtcaagacca cctacaaggc caagaagccc gtgcagctgc
     1921 ccggcgccta catcgtcgac atcaagttgg acatcgtgtc ccacaacgag gactacacca
     1981 tcgtggaaca gtacgaacgc gccgagggcc gccactccac cggcggcatg gacgagctgt
     2041 acaagtaaca cccagctttc ttgtacaaag tgggatccag acatgataag atacattgat
     2101 gagtttggac aaaccacaac tagaatgcag tgaaaaaaat gctttatttg tgaaatttgt
     2161 gatgctattg ctttatttgt aaccattata agctgcaata aacaagttaa caacaacaat
     2221 tgcattcatt ttatgtttca ggttcagggg gaggtgtggg aggttttttt caaccactcc
     2281 aggcatagct agagccactg atgcatctat agagctgaag taaagatagg gttgatgcaa
     2341 atcgatttag aattttcaat gtagaaaaaa caaaataaaa agtctattca cttgatggat
     2401 agattagtca aacaaacata gagtaggcac ttcccttgac tctgtaaact tttggattta
     2461 acagattgtg cactcctttt agttaaacac attagtccct gaaaactaag accctaacca
     2521 taatcccatt gtagacccac ctactcgggt gaatataaag ctcgcacaga tcatctcaca
     2581 gacatcagac cttggcaaag gagcatcctg aacatctcta cacactgagg caaccctagg
     2641 gccgccacca tgctgtgttg tatcagaaga acaaagcctg tggagaagaa tgaggaggct
     2701 gaccaggaac tgcagagcac ggtaccgcgg gcccgggatc caccggccgc caccatggtg
     2761 agcaagggcg aggagctgtt caccggggtg gtgcccatcc tggtcgagct ggacggcgac
     2821 gtaaacggcc acaagttcag cgtgtccggc gagggcgagg gcgatgccac ctacggcaag
     2881 ctgaccctga agttcatctg caccaccggc aagctgcccg tgccctggcc caccctcgtg
     2941 accaccctga cctggggcgt gcagtgcttc agccgctacc ccgaccacat gaagcagcac
     3001 gacttcttca agtccgccat gcccgaaggc tacgtccagg agcgcaccat cttcttcaag
     3061 gacgacggca actacaagac ccgcgccgag gtgaagttcg agggcgacac cctggtgaac
     3121 cgcatcgagc tgaagggcat cgacttcaag gaggacggca acatcctggg gcacaagctg
     3181 gagtacaact acatcagcca caacgtctat atcaccgccg acaagcagaa gaacggcatc
     3241 aaggccaact tcaagatccg ccacaacatc gaggacggca gcgtgcagct cgccgaccac
     3301 taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg
     3361 agcacccagt ccgccctgag caaagacccc aacgagaagc gcgatcacat ggtcctgctg
     3421 gagttcgtga ccgccgccgg gatcactctc ggcatggacg agctgtacaa gtaaagcggc
     3481 cgcgactcta gatcataatc agccatacca catttgtaga ggttttactt gctttaaaaa
     3541 acctcccaca cctccccctg aacctgaaac ataaaatgaa tgcaattgtt gttgttatcg
     3601 aattctgcag gaattgggtg gcatccctgt gacccctccc cagtgcctct cctggccctg
     3661 gaagttgcca ctccagtgcc caccagcctt gtcctaataa aattaagttg catcattttg
     3721 tctgactagg tgtccttcta taatattatg gggtggaggg gggtggtatg gagcaagggg
     3781 caagttggga agacaacctg tagggcacga gggggggccc gaaacccgct gatcagcctc
     3841 gactgtgcct tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac
     3901 cctggaaggt gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg
     3961 tctgagtagg tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga
     4021 ttgggaagac aatagcaagg catgctgggg atgcggtggg ctctatggca actttattat
     4081 acatagttga taattcactg gccgtcgttt tacggtacca tcgatgatga tccagacatg
     4141 ataagataca ttgatgagtt tggacaaacc acaactagaa tgcagtgaaa aaaatgcttt
     4201 atttgtgaaa tttgtgatgc tattgcttta tttgtaacca ttataagctg caataaacaa
     4261 gttaacaaca acaattgcat tcattttatg tttcaggttc agggggaggt gtgggaggtt
     4321 ttttaaagca agtaaaacct ctacaaatgt ggtatggctg attatgatcc tctagatcag
     4381 atctgcgaag atacggccac gggtgctctt gatcctgtgg ctgattttgg actgtgctgc
     4441 tcgcagctgc tgatgaatca catacttcct ccattttctt ccactgattg actgttataa
     4501 tttccctaat ttccaggtca aggtgctgtg cattgtggta atagatgtga catgacgtca
     4561 cttccaaagg accaatgaac atgtctgacc aatttcatat aatgtgaaaa cgattttcat
     4621 aggcagaata aataacattt aaattaaact gggcatcagc gcaattcaat tggtttggta
     4681 atagcaaggg aaaatagaat gaagtgatct ccaaaaaata agtacttttt gactgtaaat
     4741 aaaattgtaa ggagtaaaaa gtactttttt ttctaaaaaa atgtaattaa gtaaaagtaa
     4801 aagtattgat ttttaattgt actcaagtaa agtaaaaatc cccaaaaata atacttaagt
     4861 acagtaatca agtaaaatta ctcaagtact ttacacctct ggttcttgac cccctacctt
     4921 cagcaagccc agcagatcca ctagttctag agcggccgcc accgcggtgg agctccagct
     4981 tttgttccct ttagtgaggg ttaattgcgc gcttggcgta atcatggtca tagctgtttc
     5041 ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt
     5101 gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc
     5161 ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg
     5221 ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct
     5281 cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca
     5341 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga
     5401 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc
     5461 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
     5521 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
     5581 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt
     5641 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
     5701 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
     5761 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
     5821 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg
     5881 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
     5941 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
     6001 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga
     6061 acgaaaactc acgttaaggc attttggtca tgagattatc aaaaaggatc ttcacctaga
     6121 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt
     6181 ctgacagtta ccaatgctta ataagtgagg cacctatctc agcgatctgt ctatttcgtt
     6241 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
     6301 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag
     6361 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
     6421 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
     6481 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg
     6541 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca
     6601 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
     6661 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
     6721 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
     6781 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa
     6841 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt
     6901 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
     6961 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
     7021 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt
     7081 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
     7141 taggggttcc gcgcacattt ccccgaaaag tg
//
