LOCUS       sequence-180692-        7589 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Human NRasQ61K fused to mCherry downstream of LexOP, placed within a
            non-autonomous maize dissociation (Ds) element for integration in 
            zebrafish. Contains pCrysB-ECFP for screening..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7589)
  AUTHORS   Kenyon A, Gavriouchkina D, Zorman J, Chong-Morrison V, Napolitani G,
            Cerundolo V, Sauka-Spengler T
  TITLE     Generation of a double binary transgenic zebrafish model to study 
            myeloid gene regulation in response to oncogene activation in 
            melanocytes.
  JOURNAL   Dis Model Mech. 2018 Apr 6;11(4). pii: dmm.030056. doi: 
            10.1242/dmm.030056.
  PUBMED    29666124
REFERENCE   2  (bases 1 to 7589)
  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: "Dis Model 
            Mech."; date: "2018-04-6"; volume: "11(4). pii: dmm.030056. doi"; 
            pages: " 10.1242/dmm.030056"
COMMENT     Sequence Label: pCrys-beta:ECFP-LexOP:mCherry-NRasQ61K
FEATURES             Location/Qualifiers
     source          1..7589
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(680..699)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(681..699)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(709..726)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(709..725)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(718..740)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      867..1322
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(954..973)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1164..1185
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1348..1452
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1453..1521,1522..2313)
                     /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     complement(1671..1690)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2484..3072
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2973..2992
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3226..3243
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    3360..3381
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(3396..3401,3402..3419,3420..3426)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3434..3450
                     /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)."
     primer_bind     3439..3461
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     3458..3474
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3458..3474
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     3493..3513
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        3495..3513
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3921..3940)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     polyA_signal    3950..4031
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     3975..3994
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(4748..5455)
                     /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(5023..5042)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     5292..5310
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     regulatory      5452..5461
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(5528..5574)
                     /label=minimal CaMV 35S promoter
                     /note="minimal 35S promoter from cauliflower mosaic virus"
     polyA_signal    6185..6409
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     6398..6415
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             complement(join(6438..7151,7152..7154,7155..7157))
                     /codon_start=1
                     /product="enhanced CFP"
                     /label=ECFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(6483..6504)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     6832..6851
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     7091..7112
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      7154..7163
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    7316..7349
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
ORIGIN
        1 gctctatgct ataggctgta tacctatggc taagagaggt ctgtgttcag gtcagaggtc
       61 acgtgatgat aatccgagca gaggagtcgc tcgctgatac actccagagc agagtagatt
      121 tcatactgaa gttaataatg tagcatgaca tatattagat ggtgtgtgtg tgtgtgtgta
      181 tatacatata tatatatata tggtagcgta tttagatgga atttattgat gtattattta
      241 ttatatgtat atacagatga agtcaaaatt tttcgctccc tgtatatttt ccccttaaat
      301 cctgtttaag agaaagaaac atatttctaa tcataatagt tttaataact catttctaat
      361 aactgatttc ttctctcttt gtcatgatga tagcaccaat tgctcgaggg atctgttgca
      421 gtcatcccga attagaaaat acggtaacga aacgggatca tcccgattaa aaacgggatc
      481 ccggtgaaac ggtcgggaaa ctagctctac cgtttccgtt tccgtttacc gttttgtata
      541 tcccgtttcc gttccgtttt cgttttttac ctcgggttcg aaatcgatcg ggataaaact
      601 aacaaaatcg gttatacgat aacggtcggt acgggatttt cccatcctac tttcatccct
      661 gcgtcgggta cccaattcgc cctatagtga gtcgtattac gcgcgctcac tggccgtcgt
      721 tttacaacgt cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca
      781 tccccctttc gccagctggc gtaatagcga agaggcccgc accgatcgcc cttcccaaca
      841 gttgcgcagc ctgaatggcg aatgggacgc gccctgtagc ggcgcattaa gcgcggcggg
      901 tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt
      961 cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg
     1021 ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga
     1081 ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac
     1141 gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc
     1201 tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct attggttaaa
     1261 aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa cgcttacaat
     1321 ttaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata
     1381 cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga
     1441 aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca
     1501 ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat
     1561 cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag
     1621 agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc
     1681 gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct
     1741 cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca
     1801 gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt
     1861 ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat
     1921 gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt
     1981 gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta
     2041 cttactctag cttcccggca acaattaata gactggatgg aggcggataa agttgcagga
     2101 ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt
     2161 gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc
     2221 gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct
     2281 gagataggtg cctcactgat taagcattgg taactgtcag accaagttta ctcatatata
     2341 ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt
     2401 gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc
     2461 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg
     2521 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact
     2581 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg
     2641 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg
     2701 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac
     2761 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca
     2821 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga
     2881 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc
     2941 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct
     3001 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg
     3061 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct
     3121 tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc
     3181 tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc
     3241 gaggaagcgg aagagcgccc aatacgcaaa ccgcctctcc ccgcgcgttg gccgattcat
     3301 taatgcagct ggcacgacag gtttcccgac tggaaagcgg gcagtgagcg caacgcaatt
     3361 aatgtgagtt agctcactca ttaggcaccc caggctttac actttatgct tccggctcgt
     3421 atgttgtgtg gaattgtgag cggataacaa tttcacacag gaaacagcta tgaccatgat
     3481 tacgccaagc gcgcaattaa ccctcactaa agggaacaaa agctggagct ccccgactag
     3541 ggatgaaaac ggtcggtaac ggtcggtaaa atacctctac cgttttcatt ttcatattta
     3601 acttgcggga cggaaacgaa aacgggatat accggtaacg aaaacgaacg ggataaatac
     3661 ggtaatcgaa aaccgatacg atccggtcgg gttaaagtcg aaatcggacg ggaaccggta
     3721 tttttgttcg gtaaaatcac acatgaaaac atatattcaa aacttaaaaa caaatataaa
     3781 aaattgtaaa cacaagtctt aattaaacat agataaaatc catataaatc tggagcacac
     3841 atagtttaat gtagcacata agtgataagt cttgggctct tggctaacat aagaagccat
     3901 ataagtctac tctacgtatt gatgagtttg gacaaaccac aactagaatg cagtgaaaaa
     3961 aatgctttat ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt ataagctgca
     4021 ataaacaagt taacaacaac aattgcattc attttatgtt tcaggttcag ggggaggtgt
     4081 gggaggtttt ttaaagcaag taaaacctct acaaatgtgg tatggctgat tatgatctag
     4141 agtcgcggcc gctttacatc accacacatg gcaatcccat acaaccctga gtcccatcat
     4201 cactgctgtt gagttttttc attcggtact ggcgtatttc tcttaccagt gtgtaaaaag
     4261 catcttcaac accctgtctg gtcttggctg aggtttcaat gaatggaatc ccgtaactct
     4321 tggccagttc gtgggcttgt tttgtatcaa ctgtccttgt tggcaaatca cacttgtttc
     4381 ccactagcac cataggtaca tcatccgagt cttttactcg cttaatctgc tccctgtaga
     4441 ggttaatatc cgcaaatgac ttgctattat tgatggcaaa tacacagagg aagccttcgc
     4501 ctgtcctcat gtattggtct ctcatggcac tgtactcttc ttttccagct gtatccagta
     4561 tgtccaacaa acaggtttca ccatctataa ccacttgttt tctgtaagaa tcctctatgg
     4621 tgggatcata ttcatctaca aagtggttct ggattagctg gattgtcagt gcgcttttcc
     4681 caacaccacc tgctccaacc accaccagtt tgtactcagt cataccagaa ccacctcgcg
     4741 agtcgacctt gtacagctcg tccatgccgc cggtggagtg gcggccctcg gcgcgttcgt
     4801 actgttccac gatggtgtag tcctcgttgt gggaggtgat gtccaacttg atgttgacgt
     4861 tgtaggcgcc gggcagctgc acgggcttct tggccttgta ggtggtcttg acctcagcgt
     4921 cgtagtggcc gccgtccttc agcttcagcc tctgcttgat ctcgcccttc agggcgccgt
     4981 cctcggggta catccgctcg gaggaggcct cccagcccat ggtcttcttc tgcattacgg
     5041 ggccgtcgga ggggaagttg gtgccgcgca gcttcacctt gtagatgaac tcgccgtcct
     5101 gcagggagga gtcctgggtc acggtcacca cgccgccgtc ctcgaagttc atcacgcgct
     5161 cccacttgaa gccctcgggg aaggacagct tcaagtagtc ggggatgtcg gcggggtgct
     5221 tcacgtaggc cttggagccg tacatgaact gaggggacag gatgtcccag gcgaagggca
     5281 gggggccacc cttggtcacc ttcagcttgg cggtctgggt gccctcgtag gggcggccct
     5341 cgccctcgcc ctcgatctcg aactcgtggc cgttcacgga gccctccatg tgcaccttga
     5401 agcgcatgaa ctccttgatg atggccatgt tatcctcctc gcccttgctc accatggtgg
     5461 cgggatcccg ggcccgcggt accgtcgact gcagaattcg aggctagagt cgactagctt
     5521 cagcgtgtcc tctccaaatg aaatgaactt ccttatatag aggaagggtc ttgcgaagat
     5581 cctagagtcg agcatattac atggcattac caccatatac atatccatat acatatccat
     5641 atctaatctt acctcgactg ctgtatataa aaccagtggt tatatgtaca gtactgctgt
     5701 atataaaacc agtggttata tgtacagtac gtcgactgct gtatataaaa ccagtggtta
     5761 tatgtacagt actgctgtat ataaaaccag tggttatatg tacagtacgt cgaggggatg
     5821 ataatgcgat tagtttttta gcctcgacct gcagcccaag cggcatgcaa gctgtcgagg
     5881 ggggatcaat tcccgatcta gtaacataga tgacaccgcg cgcgataatt tatcctagtt
     5941 tgcgcgctat attttgtttt ctatcgcgta ttaaatgtat aattgcggga ctctaatcat
     6001 aaaaacccat ctcataaata acgtcatgca ttacatgtta attattacat gcttaacgta
     6061 attcaacaga aattatatga taatcatcgc aagaccggca acaggattca atcttaattt
     6121 aaattataac tagtcgacga tcggcggcct aggcgccacg tgctggttct ttccgcctca
     6181 gaagccatag agcccaccgc atccccagca tgcctgctat tgtcttccca atcctccccc
     6241 ttgctgtcct gccccacccc accccccaga atagaatgac acctactcag acaatgcgat
     6301 gcaatttcct cattttatta ggaaaggaca gtgggagtgg caccttccag ggtcaaggaa
     6361 ggcacggggg aggggcaaac aacagatggc tggcaactag aaggcacagt cgaggctgat
     6421 cagcgggttt ggccgcttta cttgtacagc tcgtccatgc cgagagtgat cccggcggcg
     6481 gtcacgaact ccagcaggac catgtgatcg cgcttctcgt tggggtcttt gctcagggcg
     6541 gactgggtgc tcaggtagtg gttgtcgggc agcagcacgg ggccgtcgcc gatgggggtg
     6601 ttctgctggt agtggtcggc gagctgcacg ctgccgtcct cgatgttgtg gcggatcttg
     6661 aagttggcct tgatgccgtt cttctgcttg tcggcggtga tatagacgtt gtggctgatg
     6721 tagttgtact ccagcttgtg ccccaggatg ttgccgtcct ccttgaagtc gatgcccttc
     6781 agctcgatgc ggttcaccag ggtgtcgccc tcgaacttca cctcggcgcg ggtcttgtag
     6841 ttgccgtcgt ccttgaagaa gatggtgcgc tcctggacgt agccttcggg catggcggac
     6901 ttgaagaagt cgtgctgctt catgtggtcg gggtagcggc tgaagcactg cacgccccag
     6961 gtcagggtgg tcacgagggt gggccagggc acgggcagct tgccggtggt gcagatgaac
     7021 ttcagggtca gcttgccgta ggtggcatcg ccctcgccct cgccggacac gctgaacttg
     7081 tggccgttta cgtcgccgtc cagctcgacc aggatgggca ccaccccggt gaacagctcc
     7141 tcgcccttgc tcaccatggt ggcgggatcc cctagcaaaa atacagatac tgcagtttct
     7201 gataggaagg ttgagcagaa tagcagggga aggaaacagg aggaagacac ttacctgtta
     7261 attaacacgg ggaatgcgcg ttccaatgca ccgttcccgg caacctagca tgcagataac
     7321 ttcgtataaa gtatcctata cgaagttatg gcaacgcgtt taaactacag ctgtgctggg
     7381 aaggctggct cgctctggcc ttcctcacca ctgccgcagc ttttatacgc tggcaagggc
     7441 agcacagcgg cgcgcggcgg acaatagaaa cacgccgtca tcagagttca gccgtgccga
     7501 agcgcagggt catcgcatcg cccagaaggc cgtaagcatt ttactgtcaa tggaaagccc
     7561 tgacggcggc actttccctc tcagcccga
//
