LOCUS       sequence-254135-        6653 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses mCherry in mammalian cells. ~200 nt of Cyrano with miR-7 
            site replaced by miR-16 site inserted in mCherry 3'UTR..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6653)
  AUTHORS   Kleaveland B, Shi CY, Stefano J, Bartel DP
  TITLE     A Network of Noncoding Regulatory RNAs Acts in the Mammalian Brain.
  JOURNAL   Cell. 2018 Jul 12;174(2):350-362.e17. doi: 
            10.1016/j.cell.2018.05.022. Epub 2018 Jun 7.
  PUBMED    29887379
REFERENCE   2  (bases 1 to 6653)
  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.cell.2018.05.022"; journalName: "Cell"; date: 
            "2018-07-12- 12"; volume: "174"; issue: "2"; pages: "350-362.e17"
COMMENT     Sequence Label: pcDNA3.1-mCherry-CyrCR1.miR16
FEATURES             Location/Qualifiers
     source          1..6653
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     629..648
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        629..647
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      724..733
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             730..1440
                     /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(875..893)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1143..1162
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     protein_bind    1738..1756
                     /label=boxB
                     /bound_moiety="lambda-N peptide"
                     /note="lambda N peptide binding site consisting of an RNA 
                     hairpin within the N-utliization (nut) site of the lambda 
                     bacteriophage transcript (Tan and Frankel, 1995)"
     protein_bind    1782..1800
                     /label=boxB
                     /bound_moiety="lambda-N peptide"
                     /note="lambda N peptide binding site consisting of an RNA 
                     hairpin within the N-utliization (nut) site of the lambda 
                     bacteriophage transcript (Tan and Frankel, 1995)"
     protein_bind    1806..1824
                     /label=boxB
                     /bound_moiety="lambda-N peptide"
                     /note="lambda N peptide binding site consisting of an RNA 
                     hairpin within the N-utliization (nut) site of the lambda 
                     bacteriophage transcript (Tan and Frankel, 1995)"
     protein_bind    1865..1883
                     /label=boxB
                     /bound_moiety="lambda-N peptide"
                     /note="lambda N peptide binding site consisting of an RNA 
                     hairpin within the N-utliization (nut) site of the lambda 
                     bacteriophage transcript (Tan and Frankel, 1995)"
     protein_bind    1889..1907
                     /label=boxB
                     /bound_moiety="lambda-N peptide"
                     /note="lambda N peptide binding site consisting of an RNA 
                     hairpin within the N-utliization (nut) site of the lambda 
                     bacteriophage transcript (Tan and Frankel, 1995)"
     primer_bind     complement(2010..2027)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2016..2240
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2286..2714
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2373..2392)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2583..2604
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2723..2743)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2728..3057
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2908..3043
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2970..2989
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3124..3918
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3178..3197)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3788..3807
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4092..4213
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4129..4148)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4183..4202
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4262..4278)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4262..4278)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4275..4297)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4286..4302
                     /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(4310..4316,4317..4334,4335..4340))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4355..4376
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4493..4510)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4664..5252)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4744..4763)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5423..6214,6215..6283))
                     /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     6046..6065
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6284..6388)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6463..6482)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctctgg ctaactagag
      601 aacccactgc ttactggctt atcgaaatta atacgactca ctatagggag acccaagctg
      661 gctagcgttt aaacttaagc ttggtaccga gctcggatcc actagtccag tgtggtggaa
      721 ttcgccacca tggtgagcaa gggcgaggag gataacatgg ccatcatcaa ggagttcatg
      781 cgcttcaagg tgcacatgga gggctccgtg aacggccacg agttcgagat cgagggcgag
      841 ggcgagggcc gcccctacga gggcacccag accgccaagc tgaaggtgac caagggtggc
      901 cccctgccct tcgcctggga catcctgtcc cctcagttca tgtacggctc caaggcctac
      961 gtgaagcacc ccgccgacat ccccgactac ttgaagctgt ccttccccga gggcttcaag
     1021 tgggagcgcg tgatgaactt cgaggacggc ggcgtggtga ccgtgaccca ggactcctcc
     1081 ctgcaggacg gcgagttcat ctacaaggtg aagctgcgcg gcaccaactt cccctccgac
     1141 ggccccgtaa tgcagaagaa gaccatgggc tgggaggcct cctccgagcg gatgtacccc
     1201 gaggacggcg ccctgaaggg cgagatcaag cagaggctga agctgaagga cggcggccac
     1261 tacgacgctg aggtcaagac cacctacaag gccaagaagc ccgtgcagct gcccggcgcc
     1321 tacaacgtca acatcaagtt ggacatcacc tcccacaacg aggactacac catcgtggaa
     1381 cagtacgaac gcgccgaggg ccgccactcc accggcggca tggacgagct gtacaagtga
     1441 gcggccgcgt taaaaccggg atatgtgcaa tagaaatata tatatatata tgtatatttt
     1501 aatacttgtg gacaaatgtt acaagttgtt taagacgcca atatttaaca ttgctgctat
     1561 tttgagatgt gtatagtttt gtaagcatta gtgcttggta gcatattgta gtgccatgtt
     1621 aggggttagt gcatgagtct agtgtctaga gggccgggta agctcgcttt cttgctgtcc
     1681 aatttctatt aaaggttcct ttgttcccta agtccaacta ctaaactggg gattcctggg
     1741 ccctgaagaa gggcccctcg actaagtcca actactaaac tgggccctga agaagggccc
     1801 atatagggcc ctgaagaagg gccctatcga ggatattatc tcgactaagt ccaactacta
     1861 aactgggccc tgaagaaggg cccatatagg gccctgaaga agggccctat cgaggatatt
     1921 atctcgagga tattatgaag ggccttgagc atctggattc tattctatag tgtcacctgc
     1981 tagagggccc gtttaaaccc gctgatcagc ctcgactgtg ccttctagtt gccagccatc
     2041 tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct
     2101 ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg
     2161 gggtggggtg gggcaggaca gcaaggggga ggattgggaa gacaatagca ggcatgctgg
     2221 ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc agctggggct ctagggggta
     2281 tccccacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt
     2341 gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct
     2401 cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg
     2461 atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtag
     2521 tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa
     2581 tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcggtct attcttttga
     2641 tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa
     2701 atttaacgcg aattaattct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc
     2761 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga
     2821 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca
     2881 accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat
     2941 tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc cgcctctgcc
     3001 tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag
     3061 ctcccgggag cttgtatatc cattttcgga tctgatcaag agacaggatg aggatcgttt
     3121 cgcatgattg aacaagatgg attgcacgca ggttctccgg ccgcttgggt ggagaggcta
     3181 ttcggctatg actgggcaca acagacaatc ggctgctctg atgccgccgt gttccggctg
     3241 tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa
     3301 ctgcaggacg aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct
     3361 gtgctcgacg ttgtcactga agcgggaagg gactggctgc tattgggcga agtgccgggg
     3421 caggatctcc tgtcatctca ccttgctcct gccgagaaag tatccatcat ggctgatgca
     3481 atgcggcggc tgcatacgct tgatccggct acctgcccat tcgaccacca agcgaaacat
     3541 cgcatcgagc gagcacgtac tcggatggaa gccggtcttg tcgatcagga tgatctggac
     3601 gaagagcatc aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc gcgcatgccc
     3661 gacggcgagg atctcgtcgt gacccatggc gatgcctgct tgccgaatat catggtggaa
     3721 aatggccgct tttctggatt catcgactgt ggccggctgg gtgtggcgga ccgctatcag
     3781 gacatagcgt tggctacccg tgatattgct gaagagcttg gcggcgaatg ggctgaccgc
     3841 ttcctcgtgc tttacggtat cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt
     3901 cttgacgagt tcttctgagc gggactctgg ggttcgaaat gaccgaccaa gcgacgccca
     3961 acctgccatc acgagatttc gattccaccg ccgccttcta tgaaaggttg ggcttcggaa
     4021 tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg ctggagttct
     4081 tcgcccaccc caacttgttt attgcagctt ataatggtta caaataaagc aatagcatca
     4141 caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca
     4201 tcaatgtatc ttatcatgtc tgtataccgt cgacctctag ctagagcttg gcgtaatcat
     4261 ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag
     4321 ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg
     4381 cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa
     4441 tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca
     4501 ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg
     4561 taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc
     4621 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc
     4681 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
     4741 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
     4801 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
     4861 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
     4921 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
     4981 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
     5041 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
     5101 gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
     5161 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
     5221 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
     5281 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
     5341 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
     5401 atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga
     5461 tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac
     5521 gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg
     5581 ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg
     5641 caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt
     5701 cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct
     5761 cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat
     5821 cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta
     5881 agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca
     5941 tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat
     6001 agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac
     6061 atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa
     6121 ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt
     6181 cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg
     6241 caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat
     6301 attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt
     6361 agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtcg
     6421 acggatcggg agatctcccg atcccctatg gtgcactctc agtacaatct gctctgatgc
     6481 cgcatagtta agccagtatc tgctccctgc ttgtgtgttg gaggtcgctg agtagtgcgc
     6541 gagcaaaatt taagctacaa caaggcaagg cttgaccgac aattgcatga agaatctgct
     6601 tagggttagg cgttttgcgc tgcttcgcga tgtacgggcc agatatacgc gtt
//
