LOCUS       sequence-217028-        3663 bp    DNA     circular SYN 14-APR-2025
DEFINITION  General donor vector with mCherry coding sequence flanking by 
            restriction sites for generating knock-in donor.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3663)
  AUTHORS   Gu B, Posfai E, Rossant J
  TITLE     Efficient generation of targeted large insertions by microinjection 
            into two-cell-stage mouse embryos.
  JOURNAL   Nat Biotechnol. 2018 Jun 11. pii: nbt.4166. doi: 10.1038/nbt.4166.
  PUBMED    29889212
REFERENCE   2  (bases 1 to 3663)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Biotechnol. 2018 Jun 11. pii: nbt.4166. doi: 10.1038/nbt.4166."
COMMENT     Sequence Label: EasyFusion mCherry
FEATURES             Location/Qualifiers
     source          1..3663
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      146..734
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     635..654
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     888..905
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    1022..1043
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(1058..1063,1064..1081,1082..1088)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1096..1112
                     /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     1101..1123
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     1120..1136
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1120..1136
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     1155..1175
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        1157..1175
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     1204..1220
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     1205..1221
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             1242..1949
                     /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(1387..1405)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1655..1674
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     complement(1957..1973)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1957..1973)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(2005..2024)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2006..2024)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2034..2051)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2034..2050)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2043..2065)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      complement(2191..2646)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2328..2349)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2540..2559
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2673..2777
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2778..2846,2847..3638)
                     /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(2996..3015)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt
       61 ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc
      121 ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct
      181 tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa
      241 ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag
      301 tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc
      361 tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg
      421 actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca
      481 cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat
      541 gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg
      601 tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc
      661 ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc
      721 ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc
      781 cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg
      841 cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga
      901 gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc
      961 attaatgcag ctggcacgac aggtttcccg actggaaagc gggcagtgag cgcaacgcaa
     1021 ttaatgtgag ttagctcact cattaggcac cccaggcttt acactttatg cttccggctc
     1081 gtatgttgtg tggaattgtg agcggataac aatttcacac aggaaacagc tatgaccatg
     1141 attacgccaa gcgcgcaatt aaccctcact aaagggaaca aaagctgggt accgggcccc
     1201 ccctcgaggt cgacggtatc gataagcttg atatcgaatt catggtgagc aagggcgagg
     1261 aggataacat ggccatcatc aaggagttca tgcgcttcaa ggtgcacatg gagggctccg
     1321 tgaacggcca cgagttcgag atcgagggcg agggcgaggg ccgcccctac gagggcaccc
     1381 agaccgccaa gctgaaggtg accaagggtg gccccctgcc cttcgcctgg gacatcctgt
     1441 cccctcagtt catgtacggc tccaaggcct acgtgaagca ccccgccgac atccccgact
     1501 acttgaagct gtccttcccc gagggcttca agtgggagcg cgtgatgaac ttcgaggacg
     1561 gcggcgtggt gaccgtgacc caggactcct ccctgcagga cggcgagttc atctacaagg
     1621 tgaagctgcg cggcaccaac ttcccctccg acggccccgt aatgcagaag aagaccatgg
     1681 gctgggaggc ctcctccgag cggatgtacc ccgaggacgg cgccctgaag ggcgagatca
     1741 agcagaggct gaagctgaag gacggcggcc actacgacgc tgaggtcaag accacctaca
     1801 aggccaagaa gcccgtgcag ctgcccggcg cctacaacgt caacatcaag ttggacatca
     1861 cctcccacaa cgaggactac accatcgtgg aacagtacga acgcgccgag ggccgccact
     1921 ccaccggcgg catggacgag ctgtacaagc ccgggggatc cactagttct agagcggccg
     1981 ccaccgcggt ggagctccaa ttcgccctat agtgagtcgt attacgcgcg ctcactggcc
     2041 gtcgttttac aacgtcgtga ctgggaaaac cctggcgtta cccaacttaa tcgccttgca
     2101 gcacatcccc ctttcgccag ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc
     2161 caacagttgc gcagcctgaa tggcgaatgg aaattgtaag cgttaatatt ttgttaaaat
     2221 tcgcgttaaa tttttgttaa atcagctcat tttttaacca ataggccgaa atcggcaaaa
     2281 tcccttataa atcaaaagaa tagaccgaga tagggttgag tgttgttcca gtttggaaca
     2341 agagtccact attaaagaac gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg
     2401 gcgatggccc actacgtgaa ccatcaccct aatcaagttt tttggggtcg aggtgccgta
     2461 aagcactaaa tcggaaccct aaagggagcc cccgatttag agcttgacgg ggaaagccgg
     2521 cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa
     2581 gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc gcttaatgcg ccgctacagg
     2641 gcgcgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct
     2701 aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat
     2761 attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt cccttttttg
     2821 cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg
     2881 aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc
     2941 ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa gttctgctat
     3001 gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc cgcatacact
     3061 attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt acggatggca
     3121 tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact
     3181 tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg
     3241 atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg
     3301 agcgtgacac cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg
     3361 aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg gataaagttg
     3421 caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag
     3481 ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc
     3541 gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga
     3601 tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa gtttactcat
     3661 ata
//
