LOCUS       sequence-488753-        5611 bp    DNA     circular SYN 08-OCT-2025
DEFINITION  Wild-type zebrafish Her3 (ortholog to human HES3) in a -2A-sfGFP 
            backbone for visualization.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5611)
  AUTHORS   Kent MR, Calderon D, Silvius KM, Kucinski JP, LaVigne CA, Cannon MV,
            Kendall GC
  TITLE     Zebrafish her3 knockout impacts developmental and cancer-related 
            gene signatures.
  JOURNAL   Dev Biol. 2023 Apr;496:1-14. doi: 10.1016/j.ydbio.2023.01.003. Epub 
            2023 Jan 23.
  PUBMED    36696714
REFERENCE   2  (bases 1 to 5611)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Oct 8, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Dev Biol.";
            date: "2023-04"; volume: "496:1-14. doi"; pages: " 
            10.1016/j.ydbio.2023.01.003. Epub 2023 Jan 23"
COMMENT     Sequence Label: pCS2+her3WT-P2A-sfGFP
FEATURES             Location/Qualifiers
     source          1..5611
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        35..53
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     35..52
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     promoter        129..147
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     CDS             854..910
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             join(917..919,920..1633)
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /note="mammalian codon-optimized"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     CDS             917..1630
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Nager et al., 2011)"
                     /label=superfolder GFP
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(947..975)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     1570..1590
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     polyA_signal    1663..1797
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1687..1706)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1741..1760
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        complement(1910..1928)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1949..1965)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1949..1965)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1962..1984)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1973..1989
                     /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(1997..2003,2004..2021,2022..2027))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2042..2063
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2180..2197)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2351..2939)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2431..2450)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3110..3901,3902..3970))
                     /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     3733..3752
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3971..4075)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(4101..4556)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4238..4259)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4450..4469
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4627..5611
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
ORIGIN
        1 cgccattctg cctggggacg tcggagcaag cttgatttag gtgacactat agaatacaag
       61 ctacttgttc tttttgcagg atcccatcga ttcgaattca aggcctctcg agcctctaga
      121 ttaattaaaa ttaaccctca ctaaagggat ggctgcagca tccaacagtg cggcgacggc
      181 aaaaccccaa aacgttaaaa aggtttcaaa accactgatg gagaaaaaaa gaagggcgcg
      241 tataaataaa tgtttgaatc aattaaagag ccttctggaa agcgcctgtt ctaacaatat
      301 ccgcaagcga aaactggaaa aggctgacat tctggagctc actgtgaaac atctgaggca
      361 tctgcaaaat acgaaaagag gactgtccaa agcttgtgac tctgctgaat atcacgctgg
      421 ttacagaagt tgtctcaaca ctgtaagtca ttatctacga gcgtcggaca cagacagaga
      481 ttcccgctcc atcatgctga caaatcttac gagcggtctt aaccacaata gagttcctga
      541 tttcagtacc gtggagagcg atcccgcatt gatctttaca ctaccaagta cacttcgacg
      601 accacacaaa gttcccatta gaacagacgt ctcttattcc agctttcaac aaactgcaga
      661 gagaaaagtt tgcttaatgc ccaagaggac tgagattggt gatagcgaca gaatgtcatt
      721 ggacgcagct cttaggagcc aggaatccaa aaaggcagaa accacacact tcagacctaa
      781 agatttaaag gtcattgagt gctgcatttt caaacagaat tattggcgac cttggggcgc
      841 gccaggaagc ggagctacta acttcagcct gctgaagcag gctggagacg tggaggagaa
      901 ccctggacct ggatccatga gcaaaggaga agaacttttc actggagttg tcccaattct
      961 tgttgaatta gatggtgatg ttaatgggca caaattttct gtccgtggag agggtgaagg
     1021 tgatgctaca aacggaaaac tcacccttaa atttatttgc actactggaa aactacctgt
     1081 tccgtggcca acacttgtca ctactctgac ctatggtgtt caatgctttt cccgttatcc
     1141 ggatcacatg aaacggcatg actttttcaa gagtgccatg cccgaaggtt atgtacagga
     1201 acgcactata tctttcaaag atgacgggac ctacaagacg cgtgctgaag tcaagtttga
     1261 aggtgatacc cttgttaatc gtatcgagtt aaagggtatt gattttaaag aagatggaaa
     1321 cattcttgga cacaaactcg agtacaactt taactcacac aatgtataca tcacggcaga
     1381 caaacaaaag aatggaatca aagctaactt caaaattcgc cacaacgttg aagatggttc
     1441 cgttcaacta gcagaccatt atcaacaaaa tactccaatt ggcgatggcc ctgtcctttt
     1501 accagacaac cattacctgt cgacacaatc tgtcctgtcg aaagatccca acgaaaagcg
     1561 tgaccacatg gtccttcttg agtttgtaac tgctgctggg attacacatg gcatggatga
     1621 gctctacaaa taatctagaa ctatagtgag tcgtattacg tagatccaga catgataaga
     1681 tacattgatg agtttggaca aaccacaact agaatgcagt gaaaaaaatg ctttatttgt
     1741 gaaatttgtg atgctattgc tttatttgta accattataa gctgcaataa acaagttaac
     1801 aacaacaatt gcattcattt tatgtttcag gttcaggggg aggtgtggga ggttttttaa
     1861 ttcgcggccg cggcgccaat gcattgggcc cggtacccag cttttgttcc ctttagtgag
     1921 ggttaattgc gcgcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc
     1981 cgctcacaat tccacacaac atacgagccg ggagcataaa gtgtaaagcc tggggtgcct
     2041 aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
     2101 acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta
     2161 ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     2221 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     2281 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     2341 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     2401 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     2461 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     2521 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     2581 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     2641 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     2701 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     2761 agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga
     2821 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     2881 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     2941 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     3001 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     3061 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     3121 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     3181 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     3241 tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     3301 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     3361 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     3421 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     3481 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     3541 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     3601 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     3661 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     3721 cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     3781 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     3841 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     3901 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     3961 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     4021 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
     4081 ttccccgaaa agtgccacct aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa
     4141 tttttgttaa atcagctcat tttttaacca ataggccgaa atcggcaaaa tcccttataa
     4201 atcaaaagaa tagaccgaga tagggttgag tgttgttcca gtttggaaca agagtccact
     4261 attaaagaac gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc
     4321 actacgtgaa ccatcaccct aatcaagttt tttggggtcg aggtgccgta aagcactaaa
     4381 tcggaaccct aaagggagcc cccgatttag agcttgacgg ggaaagccgg cgaacgtggc
     4441 gagaaaggaa gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt
     4501 cacgctgcgc gtaaccacca cacccgccgc gcttaatgcg ccgctacagg gcgcgtccca
     4561 ttcgccattc aggctgcgca actgttggga agggcgatcg gtgcgggcct cttcgctatt
     4621 acgccagtcg accgccaatt caatatggcg tatatggact catgccaatt caatatggtg
     4681 gatctggacc tgtgccaatt caatatggcg tatatggact cgtgccaatt caatatggtg
     4741 gatctggacc ccagccaatt caatatggcg gacttggcac catgccaatt caatatggcg
     4801 gacctggcac tgtgccaact ggggaggggt ctacttggca cggtgccaag tttgaggagg
     4861 ggtcttggcc ctgtgccaag tccgccatat tgaattggca tggtgccaat aatggcggcc
     4921 atattggcta tatgccagga tcaatatata ggcaatatcc aatatggccc tatgccaata
     4981 tggctattgg ccaggttcaa tactatgtat tggccctatg ccatatagta ttccatatat
     5041 gggttttcct attgacgtag atagcccctc ccaatgggcg gtcccatata ccatatatgg
     5101 ggcttcctaa taccgcccat agccactccc ccattgacgt caatggtctc tatatatggt
     5161 ctttcctatt gacgtcatat gggcggtcct attgacgtat atggcgcctc ccccattgac
     5221 gtcaattacg gtaaatggcc cgcctggctc aatgcccatt gacgtcaata ggaccaccca
     5281 ccattgacgt caatgggatg gctcattgcc cattcatatc cgttctcacg ccccctattg
     5341 acgtcaatga cggtaaatgg cccacttggc agtacatcaa tatctattaa tagtaacttg
     5401 gcaagtacat tactattgga agtacgccag ggtacattgg cagtactccc attgacgtca
     5461 atggcggtaa atggcccgcg atggctgcca agtacatccc cattgacgtc aatggggagg
     5521 ggcaatgacg caaatgggcg ttccattgac gtaaatgggc ggtaggcgtg cctaatggga
     5581 ggtctatata agcaatgctc gtttagggaa c
//
