LOCUS       sequence-204495-        4884 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  a cmlc2::mCherry (red heart) reporter gene was inserted at the 3'end
            of pKHR4 mcs, that resides within donor fragments produced by I-SceI
            digestion. Genbank accession number is KU144824..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4884)
  AUTHORS   Hoshijima K, Jurynec MJ, Grunwald DJ
  TITLE     Precise Editing of the Zebrafish Genome Made Simple and Efficient.
  JOURNAL   Dev Cell. 2016 Mar 21;36(6):654-67. doi: 
            10.1016/j.devcel.2016.02.015.
  PUBMED    27003937
REFERENCE   2  (bases 1 to 4884)
  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"; doi: 
            "10.1016/j.devcel.2016.02"; journalName: "Dev Cell"; date: 
            "2016-03-21- 21"; volume: "36"; issue: "6"; pages: "654-67"
COMMENT     Sequence Label: pKHR7
FEATURES             Location/Qualifiers
     source          1..4884
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             893..1603
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEDDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(1038..1056)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1306..1325
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     1500..1523
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     polyA_signal    1641..1775
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1665..1684)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1719..1738
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        complement(1905..1923)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1944..1960)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1944..1960)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1957..1979)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1968..1984
                     /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(1992..1998,1999..2016,2017..2022))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2037..2058
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2175..2192)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2346..2934)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2426..2445)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3105..3896,3897..3965))
                     /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     3728..3747
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3966..4070)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(4096..4551)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4233..4254)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4445..4464
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4678..4700
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4692..4709
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4693..4709
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        4716..4734
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     4816..4832
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4816..4832
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(4866..4882)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4867..4883)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
ORIGIN
        1 aaatcagttg tgttaaataa gagacattca aaataaatgt aaatgagctc tccaaatcag
       61 cagacttaac attctttaaa atgattgatt caatagtgat aaaaatcagg catagccagt
      121 tgtaacttta gataaattac agaaaatgtc aaatacagag aaccgattct tttttatgat
      181 acatccaagc acacatttaa cacaatccag gcaaaccccg aatttcacag tcacaagcac
      241 tgtttgtaca agagctttgc ctaaggacac acagtctcta taagtccagg tcgttggttt
      301 cactcttatt ttaaacatgt gacatttttc ctgccatcct gtcttaggct gctgtttgct
      361 tcattccatg tcacattaaa ttcctcagta gcacctttta cacacacagc caatcttttc
      421 cagaaaattc aattgctttg aagagataat gtgtgaacaa atccatttag aaaaggaaaa
      481 ttaagaattt gtaaaatcat ctgtaaattg ttggcattct tctgtatatg aacatcacat
      541 catttacagg taaaggtctg gtcattaatt atatgacaat ttactggtat tattttgtga
      601 aaggggctat tttcaatgcg ttcatccatc cttttcatcc ctcaaatctc tcattcacgt
      661 ccccctcccc atctgcacac tttatctcat tttccaccct gctggaatct gagcacttgt
      721 gcagttatca gggctcctgt atttaggagg ctctgggtgt ccatgtaggg gacgaacaga
      781 aacactgcag acctttatag aagaacaatt gataagagtc ctcatacata aagactccat
      841 tagtaagcca gtgacccagg agcccagacc aacagcaaag cagacagtga ccatggtgag
      901 caagggcgag gacgacaaca tggccatcat caaggagttc atgcgcttca aggtgcacat
      961 ggagggctcc gtgaacggcc acgagttcga gatcgagggc gagggcgagg gccgccccta
     1021 cgagggcacc cagaccgcca agctgaaggt gaccaagggc ggccccctgc ccttcgcctg
     1081 ggacatcctg tcccctcagt tcatgtacgg ctccaaggcc tacgtgaagc accccgccga
     1141 catccccgac tacttgaagc tgtccttccc cgagggcttc aagtgggagc gcgtgatgaa
     1201 cttcgaggac ggcggcgtgg tgaccgtgac ccaggactcc tccctgcagg acggcgagtt
     1261 catctacaag gtgaagctgc gcggcaccaa cttcccctcc gacggccccg taatgcagaa
     1321 gaagaccatg ggctgggagg cctcctccga gcggatgtac cccgaggacg gcgccctgaa
     1381 gggcgagatc aagcagaggc tgaagctgaa ggacggcggc cactacgacg ccgaggtcaa
     1441 gaccacctac aaggccaaga agcccgtgca gctgcccggc gcctacaacg tcaacatcaa
     1501 gctggacatc acctcccaca acgaggacta caccatcgtg gaacagtacg agcgcgccga
     1561 gggccgccac tccaccggcg gcatggacga gctgtacaag taaggcgcgc ctctagaact
     1621 atagtgagtc gtattacgta gatccagaca tgataagata cattgatgag tttggacaaa
     1681 ccacaactag aatgcagtga aaaaaatgct ttatttgtga aatttgtgat gctattgctt
     1741 tatttgtaac cattataagc tgcaataaac aagttaacaa caacaattgc attcatttta
     1801 tgtttcaggt tcagggggag gtgtgggagg tttttttcga gccgccatta ccctgttatc
     1861 cctacccgaa tacccgacta aaggaccggt accagctttt gttcccttta gtgagggtta
     1921 atttcgagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc
     1981 acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga
     2041 gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg
     2101 tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg
     2161 cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg
     2221 gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga
     2281 aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
     2341 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
     2401 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
     2461 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
     2521 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
     2581 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
     2641 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
     2701 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
     2761 tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca
     2821 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     2881 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     2941 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     3001 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     3061 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
     3121 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     3181 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     3241 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     3301 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     3361 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     3421 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     3481 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     3541 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     3601 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     3661 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     3721 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     3781 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     3841 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     3901 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     3961 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     4021 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc
     4081 cgaaaagtgc cacctaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt
     4141 gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa
     4201 aagaatagac cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa
     4261 agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac
     4321 gtgaaccatc accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga
     4381 accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa
     4441 aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc
     4501 tgcgcgtaac caccacaccc gccgcgctta atgcgccgct acagggcgcg tcccattcgc
     4561 cattcaggct gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc
     4621 agctggcgaa agggggatgt gctgcaaggc gattaagttg ggtaacgcca gggttttccc
     4681 agtcacgacg ttgtaaaacg acggccagtg aattgtaata cgactcacta tagggcgaat
     4741 tggagctcca ccgcggtggc ggccaggtcc tttagtcggg tattcgggta gggataacag
     4801 ggtaatggcg gccgctctag aactagtgga tcccccgggc tgcaggaatt cgatatcaag
     4861 cttatcgata ccgtcgacct cgag
//
