LOCUS       sequence-176786-        7887 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Empty backbone, includes IRES-mCherry.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7887)
  AUTHORS   Golden RJ, Chen B, Li T, Braun J, Manjunath H, Chen X, Wu J, Schmid 
            V, Chang TC, Kopp F, Ramirez-Martinez A, Tagliabracci VS, Chen ZJ, 
            Xie Y, Mendell JT
  TITLE     An Argonaute phosphorylation cycle promotes microRNA-mediated 
            silencing.
  JOURNAL   Nature. 2017 Feb 9;542(7640):197-202. doi: 10.1038/nature21025. Epub
            2017 Jan 23.
  PUBMED    28114302
REFERENCE   2  (bases 1 to 7887)
  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: 
            "10.1038/nature21025"; journalName: "Nature"; date: "2017-02-9- 9"; 
            volume: "542"; issue: "7640"; pages: "197-202"
COMMENT     Sequence Label: pCAGImC_Empty
FEATURES             Location/Qualifiers
     source          1..7887
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        4..383
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        386..661
                     /label=chicken beta-actin promoter
     intron          662..1670
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1594..1616
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1678..1697
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             1736..1777
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     protein_bind    1787..1911
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(1936..1941,1942..1959,1960..1966)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             2020..2679
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(2086..2105)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             3000..3305
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     3184..3203
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(3346..3470)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     misc_feature    join(3547..4121,4122..4124,4125..4133)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3714..3731)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3941..3960
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             4134..4844
                     /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(4279..4297)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     4547..4566
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     complement(4911..4930)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    4976..5031
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(5030..5049)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(5390..5406)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5390..5406)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5403..5425)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5414..5430
                     /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(5438..5444,5445..5462,5463..5468))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5483..5504
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        5563..5758
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      5609..5744
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    5764..5898
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5801..5820)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5855..5874
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5966..5983)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6137..6725)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6217..6236)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6896..7687,7688..7756))
                     /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     7519..7538
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7757..7861)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgacattg attattgact agttattaat agtaatcaat tacggggtca ttagttcata
       61 gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc
      121 ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag
      181 ggactttcca ttgacgtcaa tgggtggact atttacggta aactgcccac ttggcagtac
      241 atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg
      301 cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg
      361 tattagtcat cgctattacc atgggtcgag gtgagcccca cgttctgctt cactctcccc
      421 atctcccccc cctccccacc cccaattttg tatttattta ttttttaatt attttgtgca
      481 gcgatggggg cggggggggg gggggcgcgc gccaggcggg gcggggcggg gcgaggggcg
      541 gggcggggcg aggcggagag gtgcggcggc agccaatcag agcggcgcgc tccgaaagtt
      601 tccttttatg gcgaggcggc ggcggcggcg gccctataaa aagcgaagcg cgcggcgggc
      661 gggagtcgct gcgttgcctt cgccccgtgc cccgctccgc gccgcctcgc gccgcccgcc
      721 ccggctctga ctgaccgcgt tactcccaca ggtgagcggg cgggacggcc cttctcctcc
      781 gggctgtaat tagcgcttgg tttaatgacg gctcgtttct tttctgtggc tgcgtgaaag
      841 ccttaaaggg ctccgggagg gccctttgtg cgggggggag cggctcgggg ggtgcgtgcg
      901 tgtgtgtgtg cgtggggagc gccgcgtgcg gcccgcgctg cccggcggct gtgagcgctg
      961 cgggcgcggc gcggggcttt gtgcgctccg cgtgtgcgcg aggggagcgc ggccgggggc
     1021 ggtgccccgc ggtgcggggg ggctgcgagg ggaacaaagg ctgcgtgcgg ggtgtgtgcg
     1081 tgggggggtg agcagggggt gtgggcgcgg cggtcgggct gtaacccccc cctgcacccc
     1141 cctccccgag ttgctgagca cggcccggct tcgggtgcgg ggctccgtgc ggggcgtggc
     1201 gcggggctcg ccgtgccggg cggggggtgg cggcaggtgg gggtgccggg cggggcgggg
     1261 ccgcctcggg ccggggaggg ctcgggggag gggcgcggcg gccccggagc gccggcggct
     1321 gtcgaggcgc ggcgagccgc agccattgcc ttttatggta atcgtgcgag agggcgcagg
     1381 gacttccttt gtcccaaatc tggcggagcc gaaatctggg aggcgccgcc gcaccccctc
     1441 tagcgggcgc gggcgaagcg gtgcggcgcc ggcaggaagg aaatgggcgg ggagggcctt
     1501 cgtgcgtcgc cgcgccgccg tccccttctc catctccagc ctcggggctg ccgcaggggg
     1561 acggctgcct tcggggggga cggggcaggg cggggttcgg cttctggcgt gtgaccggcg
     1621 gctctagagc ctctgctaac catgttcatg ccttcttctt tttcctacag ctcctgggca
     1681 acgtgctggt tattgtgctg tctcatcatt ttggcaaaga attaccacca tggcaggcaa
     1741 gccaatccct aaccctctgc tgggcctgga cagcaccgaa ttgatcacaa gtttgtacaa
     1801 aaaagctgaa cgagaaacgt aaaatgatat aaatatcaat atattaaatt agattttgca
     1861 taaaaaacag actacataat actgtaaaac acaacatatc cagtcactat ggcggccgca
     1921 ttaggcaccc caggctttac actttatgct tccggctcgt ataatgtgtg gattttgagt
     1981 taggatccgg cgagattttc aggagctaag gaagctaaaa tggagaaaaa aatcactgga
     2041 tataccaccg ttgatatatc ccaatggcat cgtaaagaac attttgaggc atttcagtca
     2101 gttgctcaat gtacctataa ccagaccgtt cagctggata ttacggcctt tttaaagacc
     2161 gtaaagaaaa ataagcacaa gttttatccg gcctttattc acattcttgc ccgcctgatg
     2221 aatgctcatc cggaattccg tatggcaatg aaagacggtg agctggtgat atgggatagt
     2281 gttcaccctt gttacaccgt tttccatgag caaactgaaa cgttttcatc gctctggagt
     2341 gaataccacg acgatttccg gcagtttcta cacatatatt cgcaagatgt ggcgtgttac
     2401 ggtgaaaacc tggcctattt ccctaaaggg tttattgaga atatgttttt cgtctcagcc
     2461 aatccctggg tgagtttcac cagttttgat ttaaacgtgg ccaatatgga caacttcttc
     2521 gcccccgttt tcaccatggg caaatattat acgcaaggcg acaaggtgct gatgccgctg
     2581 gcgattcagg ttcatcatgc cgtctgtgat ggcttccatg tcggcagaat gcttaatgaa
     2641 ttacaacagt actgcgatga gtggcagggc ggggcgtaaa cgcgtggatc cggcttacta
     2701 aaagccagat aacagtatgc gtatttgcgc gcaccggtgc tagcgtatac ccgaagtatg
     2761 tcaaaaagag gtgtgctatg aagcagcgta ttacagtgac agttgacagc gacagctatc
     2821 agttgctcaa ggcatatatg atgtcaatat ctccggtctg gtaagcacaa ccatgcagaa
     2881 tgaagcccgt cgtctgcgtg ccgaacgctg gaaagcggaa aatcaggaag ggatggctga
     2941 ggtcgcccgg tttattgaaa tgaacggctc ttttgctgac gagaacaggg actggtgaaa
     3001 tgcagtttaa ggtttacacc tataaaagag agagccgtta tcgtctgttt gtggatgtac
     3061 agagtgatat tattgacacg cccgggcgac ggatggtgat ccccctggcc agtgcacgtc
     3121 tgctgtcaga taaagtctcc cgtgaacttt acccggtggt gcatatcggg gatgaaagct
     3181 ggcgcatgat gaccaccgat atggccagtg tgccggtctc cgttatcggg gaagaagtgg
     3241 ctgatctcag ccaccgcgaa aatgacatca aaaacgccat taacctgatg ttctggggaa
     3301 tataaatgtc aggctccgtt atacacagcc agtctgcagg tcgaccatag tgactggata
     3361 tgttgtgttt tacagtatta tgtagtctgt tttttatgca aaatctaatt taatatattg
     3421 atatttatat cattttacgt ttctcgttca gctttcttgt acaaagtggt tgatctcgag
     3481 gattggcgcg ccaagatatc ttaattaagt taacaggcgg ccgctacgta aattccgccc
     3541 cccccccccc tctccctccc ccccccctaa cgttactggc cgaagccgct tggaataagg
     3601 ccggtgtgcg tttgtctata tgttattttc caccatattg ccgtcttttg gcaatgtgag
     3661 ggcccggaaa cctggccctg tcttcttgac gagcattcct aggggtcttt cccctctcgc
     3721 caaaggaatg caaggtctgt tgaatgtcgt gaaggaagca gttcctctgg aagcttcttg
     3781 aagacaaaca acgtctgtag cgaccctttg caggcagcgg aaccccccac ctggcgacag
     3841 gtgcctctgc ggccaaaagc cacgtgtata agatacacct gcaaaggcgg cacaacccca
     3901 gtgccacgtt gtgagttgga tagttgtgga aagagtcaaa tggctctcct caagcgtatt
     3961 caacaagggg ctgaaggatg cccagaaggt accccattgt atgggatctg atctggggcc
     4021 tcggtgcaca tgctttacat gtgtttagtc gaggttaaaa aaacgtctag gccccccgaa
     4081 ccacggggac gtggttttcc tttgaaaaac acgatgataa tatggccaca accatggtga
     4141 gcaagggcga ggaggataac atggccatca tcaaggagtt catgcgcttc aaggtgcaca
     4201 tggagggctc cgtgaacggc cacgagttcg agatcgaggg cgagggcgag ggccgcccct
     4261 acgagggcac ccagaccgcc aagctgaagg tgaccaaggg tggccccctg cccttcgcct
     4321 gggacatcct gtcccctcag ttcatgtacg gctccaaggc ctacgtgaag caccccgccg
     4381 acatccccga ctacttgaag ctgtccttcc ccgagggctt caagtgggag cgcgtgatga
     4441 acttcgagga cggcggcgtg gtgaccgtga cccaggactc ctccctgcag gacggcgagt
     4501 tcatctacaa ggtgaagctg cgcggcacca acttcccctc cgacggcccc gtaatgcaga
     4561 agaagaccat gggctgggag gcctcctccg agcggatgta ccccgaggac ggcgccctga
     4621 agggcgagat caagcagagg ctgaagctga aggacggcgg ccactacgac gctgaggtca
     4681 agaccaccta caaggccaag aagcccgtgc agctgcccgg cgcctacaac gtcaacatca
     4741 agttggacat cacctcccac aacgaggact acaccatcgt ggaacagtac gaacgcgccg
     4801 agggccgcca ctccaccggc ggcatggacg agctgtacaa gtgagtttgg ggccggccgc
     4861 cagcacagtg gtcgatcgac caatgccctg gctcacaaat accactgaga tctttttccc
     4921 tctgccaaaa attatgggga catcatgaag ccccttgagc atctgacttc tggctaataa
     4981 aggaaattta ttttcattgc aatagtgtgt tggaattttt tgtgtctctc actcggaagg
     5041 acatatggga gggcaaatca tttaaaacat cagaatgagt atttggttta gagtttggca
     5101 acatatgcca tatgctggct gccatgaaca aaggtggcta taaagaggtc atcagtatat
     5161 gaaacagccc cctgctgtcc attccttatt ccatagaaaa gccttgactt gaggttagat
     5221 tttttttata ttttgttttg tgttattttt ttctttaaca tccctaaaat tttccttaca
     5281 tgttttacta gccagatttt tcctcctctc ctgactactc ccagtcatag ctgtccctct
     5341 tctcttatga agatccctcg acctgcagcc caagcttggc gtaatcatgg tcatagctgt
     5401 ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa
     5461 agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac
     5521 tgcccgcttt ccagtcggga aacctgtcgt gccagcggat ccgcatctca attagtcagc
     5581 aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca
     5641 ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg ccgcctcggc
     5701 ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa
     5761 gctaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc
     5821 acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta
     5881 tcttatcatg tctggatccg ctgcattaat gaatcggcca acgcgcgggg agaggcggtt
     5941 tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
     6001 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
     6061 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
     6121 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
     6181 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
     6241 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
     6301 ttctcccttc gggaagcgtg gcgctttctc aatgctcacg ctgtaggtat ctcagttcgg
     6361 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
     6421 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
     6481 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
     6541 tcttgaagtg gtggcctaac tacggctaca ctagaaggac agtatttggt atctgcgctc
     6601 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
     6661 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
     6721 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
     6781 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt
     6841 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc
     6901 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
     6961 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg
     7021 ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc
     7081 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta
     7141 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg
     7201 ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct
     7261 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta
     7321 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg
     7381 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga
     7441 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt
     7501 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca
     7561 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt
     7621 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt
     7681 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga
     7741 aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat cagggttatt
     7801 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
     7861 gcacatttcc ccgaaaagtg ccacctg
//
