LOCUS       sequence-366108-        7162 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Used to generate the Tg(QUAS:GFP-CAAX; he1.1:YFP)c631 transgenic 
            line.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7162)
  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 7162)
  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:GFP-CAAX-he1.1:YFP
FEATURES             Location/Qualifiers
     source          1..7162
                     /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             join(1333..1335,1336..1338,1339..2049)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1333..1335,1336..1338,1339..2049)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     polyA_signal    2165..2299
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     protein_bind    complement(2361..2385)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             join(2870..2872,2873..2875,2876..3589)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3827..3844)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3833..4058
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    complement(4059..4079)
                     /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(4087..4103)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     polyA_signal    4119..4253
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     misc_feature    join(4692..4879,4880..4891)
                     /label=Tol2 left end
                     /note="left end of the Tol2 transposon from the medaka fish
                     Oryzias latipes (Kawakami, 2007)"
     primer_bind     complement(4925..4941)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4925..4941)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(4978..4998)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(4978..4996)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5017..5033)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5030..5052)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5041..5057
                     /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(5065..5071,5072..5089,5090..5095))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5110..5131
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5248..5265)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5419..6007)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5499..5518)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6178..6969,6970..7038))
                     /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     6801..6820
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7039..7143)
                     /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 ctatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg
     1381 gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc
     1441 gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg
     1501 ccctggccca ccctcgtgac caccctgacc tacggcgtgc agtgcttcag ccgctacccc
     1561 gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag
     1621 cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag
     1681 ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac
     1741 atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat catggccgac
     1801 aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga ggacggcagc
     1861 gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg
     1921 cccgacaacc actacctgag cacccagtcc gccctgagca aagaccccaa cgagaagcgc
     1981 gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag
     2041 ctgtacaagg gaggaggaag atctaagctg aaccctcctg atgagagtgg ccccggctgc
     2101 atgagctgca agtgtgtgct ctcctgaggc gcgcctctag aactatagtg agtcgtatta
     2161 cgtagatcca gacatgataa gatacattga tgagtttgga caaaccacaa ctagaatgca
     2221 gtgaaaaaaa tgctttattt gtgaaatttg tgatgctatt gctttatttg taaccattat
     2281 aagctgcaat aaacaagtta acaacaacaa ttgcattcat tttatgtttc aggttcaggg
     2341 ggaggtgtgg gaggtttttt acccagcttt cttgtacaaa gtggtcaacc actccaggca
     2401 tagctagagc cactgatgca tctatagagc tgaagtaaag atagggttga tgcaaatcga
     2461 tttagaattt tcaatgtaga aaaaacaaaa taaaaagtct attcacttga tggatagatt
     2521 agtcaaacaa acatagagta ggcacttccc ttgactctgt aaacttttgg atttaacaga
     2581 ttgtgcactc cttttagtta aacacattag tccctgaaaa ctaagaccct aaccataatc
     2641 ccattgtaga cccacctact cgggtgaata taaagctcgc acagatcatc tcacagacat
     2701 cagaccttgg caaaggagca tcctgaacat ctctacacac tgaggcaacc ctagggccgc
     2761 caccatgctg tgttgtatca gaagaacaaa gcctgtggag aagaatgagg aggctgacca
     2821 ggaactgcag agcacggtac ctagagctag agaccctcct gtggctacaa tggtgagcaa
     2881 gggcgaggag ctgttcaccg gggtggtgcc catcctggtc gagctggacg gcgacgtaaa
     2941 cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg gcaagctgac
     3001 cctgaagttc atctgcacca ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac
     3061 cttcggctac ggcctgcagt gcttcgcccg ctaccccgac cacatgaagc agcacgactt
     3121 cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc accatcttct tcaaggacga
     3181 cggcaactac aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat
     3241 cgagctgaag ggcatcgact tcaaggagga cggcaacatc ctggggcaca agctggagta
     3301 caactacaac agccacaacg tctatatcat ggccgacaag cagaagaacg gcatcaaggt
     3361 gaacttcaag atccgccaca acatcgagga cggcagcgtg cagctcgccg accactacca
     3421 gcagaacacc cccatcggcg acggccccgt gctgctgccc gacaaccact acctgagcta
     3481 ccagtccgcc ctgagcaaag accccaacga gaagcgcgat cacatggtcc tgctggagtt
     3541 cgtgaccgcc gccgggatca ctctcggcat ggacgagctg tataagtgag aattctgcag
     3601 gaattgggtg gcatccctgt gacccctccc cagtgcctct cctggccctg gaagttgcca
     3661 ctccagtgcc caccagcctt gtcctaataa aattaagttg catcattttg tctgactagg
     3721 tgtccttcta taatattatg gggtggaggg gggtggtatg gagcaagggg caagttggga
     3781 agacaacctg tagggcacga gggggggccc gaaacccgct gatcagcctc gactgtgcct
     3841 tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt
     3901 gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg
     3961 tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac
     4021 aatagcaagg catgctgggg atgcggtggg ctctatggca actttattat acatagttga
     4081 taattcactg gccgtcgttt tacggtacca tcgatgatga tccagacatg ataagataca
     4141 ttgatgagtt tggacaaacc acaactagaa tgcagtgaaa aaaatgcttt atttgtgaaa
     4201 tttgtgatgc tattgcttta tttgtaacca ttataagctg caataaacaa gttaacaaca
     4261 acaattgcat tcattttatg tttcaggttc agggggaggt gtgggaggtt ttttaaagca
     4321 agtaaaacct ctacaaatgt ggtatggctg attatgatcc tctagatcag atctgcgaag
     4381 atacggccac gggtgctctt gatcctgtgg ctgattttgg actgtgctgc tcgcagctgc
     4441 tgatgaatca catacttcct ccattttctt ccactgattg actgttataa tttccctaat
     4501 ttccaggtca aggtgctgtg cattgtggta atagatgtga catgacgtca cttccaaagg
     4561 accaatgaac atgtctgacc aatttcatat aatgtgaaaa cgattttcat aggcagaata
     4621 aataacattt aaattaaact gggcatcagc gcaattcaat tggtttggta atagcaaggg
     4681 aaaatagaat gaagtgatct ccaaaaaata agtacttttt gactgtaaat aaaattgtaa
     4741 ggagtaaaaa gtactttttt ttctaaaaaa atgtaattaa gtaaaagtaa aagtattgat
     4801 ttttaattgt actcaagtaa agtaaaaatc cccaaaaata atacttaagt acagtaatca
     4861 agtaaaatta ctcaagtact ttacacctct ggttcttgac cccctacctt cagcaagccc
     4921 agcagatcca ctagttctag agcggccgcc accgcggtgg agctccagct tttgttccct
     4981 ttagtgaggg ttaattgcgc gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa
     5041 ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg
     5101 gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca
     5161 gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg
     5221 tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg
     5281 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
     5341 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     5401 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     5461 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     5521 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     5581 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     5641 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     5701 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     5761 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     5821 gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc
     5881 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     5941 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     6001 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     6061 acgttaaggc attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     6121 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta
     6181 ccaatgctta ataagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     6241 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag
     6301 tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag caataaacca
     6361 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc
     6421 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt
     6481 tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag
     6541 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt
     6601 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat
     6661 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt
     6721 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc
     6781 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat
     6841 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
     6901 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
     6961 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
     7021 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
     7081 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
     7141 gcgcacattt ccccgaaaag tg
//
