LOCUS       sequence-181326-        7272 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses V5-CSNK1A1, includes IRES-mCherry .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7272)
  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 7272)
  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_V5-CSNK1A1
FEATURES             Location/Qualifiers
     source          1..7272
                     /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..1811
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(2829..2853)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_feature    join(2930..3504,3505..3507,3508..3516)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3097..3114)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3324..3343
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             3517..4227
                     /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(3662..3680)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     3930..3949
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     complement(4294..4313)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    4359..4414
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(4413..4432)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(4775..4791)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4775..4791)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4788..4810)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4799..4815
                     /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(4823..4829,4830..4847,4848..4853))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4868..4889
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        4948..5143
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      4994..5129
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    5149..5283
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5186..5205)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5240..5259
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5351..5368)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5522..6110)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5602..5621)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6281..7072,7073..7141))
                     /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     6904..6923
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7142..7246)
                     /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 aaaagcaggc accatggcga gtagcagcgg ctccaaggct gaattcattg tcggagggaa
     1861 atataaactg gtacggaaga tcgggtctgg ctccttcggg gacatctatt tggcgatcaa
     1921 catcaccaac ggcgaggaag tggcagtgaa gctagaatct cagaaggcca ggcatcccca
     1981 gttgctgtac gagagcaagc tctataagat tcttcaaggt ggggttggca tcccccacat
     2041 acggtggtat ggtcaggaaa aagactacaa tgtactagtc atggatcttc tgggacctag
     2101 cctcgaagac ctcttcaatt tctgttcaag aaggttcaca atgaaaactg tacttatgtt
     2161 agctgaccag atgatcagta gaattgaata tgtgcataca aagaatttta tacacagaga
     2221 cattaaacca gataacttcc taatgggtat tgggcgtcac tgtaataagt tattccttat
     2281 tgattttggt ttggccaaaa agtacagaga caacaggaca aggcaacaca taccatacag
     2341 agaagataaa aacctcactg gcactgcccg atatgctagc atcaatgcac atcttggtat
     2401 tgagcagagt cgccgagatg acatggaatc attaggatat gttttgatgt attttaatag
     2461 aaccagcctg ccatggcaag ggctaaaggc tgcaacaaag aaacaaaaat atgaaaagat
     2521 tagtgaaaag aagatgtcca cgcctgttga agttttatgt aaggggtttc ctgcagaatt
     2581 tgcgatgtac ttaaactatt gtcgtgggct acgctttgag gaagccccag attacatgta
     2641 tctgaggcag ctattccgca ttcttttcag gaccctgaac catcaatatg actacacatt
     2701 tgattggaca atgttaaagc agaaagcagc acagcaggca gcctcttcca gtgggcaggg
     2761 tcagcaggcc caaaccccca caggcaagca aactgacaaa accaagagta acatgaaagg
     2821 tttctagaac ccagctttct tgtacaaagt ggttgatctc gaggattggc gcgccaagat
     2881 atcttaatta agttaacagg cggccgctac gtaaattccg cccccccccc ccctctccct
     2941 cccccccccc taacgttact ggccgaagcc gcttggaata aggccggtgt gcgtttgtct
     3001 atatgttatt ttccaccata ttgccgtctt ttggcaatgt gagggcccgg aaacctggcc
     3061 ctgtcttctt gacgagcatt cctaggggtc tttcccctct cgccaaagga atgcaaggtc
     3121 tgttgaatgt cgtgaaggaa gcagttcctc tggaagcttc ttgaagacaa acaacgtctg
     3181 tagcgaccct ttgcaggcag cggaaccccc cacctggcga caggtgcctc tgcggccaaa
     3241 agccacgtgt ataagataca cctgcaaagg cggcacaacc ccagtgccac gttgtgagtt
     3301 ggatagttgt ggaaagagtc aaatggctct cctcaagcgt attcaacaag gggctgaagg
     3361 atgcccagaa ggtaccccat tgtatgggat ctgatctggg gcctcggtgc acatgcttta
     3421 catgtgttta gtcgaggtta aaaaaacgtc taggcccccc gaaccacggg gacgtggttt
     3481 tcctttgaaa aacacgatga taatatggcc acaaccatgg tgagcaaggg cgaggaggat
     3541 aacatggcca tcatcaagga gttcatgcgc ttcaaggtgc acatggaggg ctccgtgaac
     3601 ggccacgagt tcgagatcga gggcgagggc gagggccgcc cctacgaggg cacccagacc
     3661 gccaagctga aggtgaccaa gggtggcccc ctgcccttcg cctgggacat cctgtcccct
     3721 cagttcatgt acggctccaa ggcctacgtg aagcaccccg ccgacatccc cgactacttg
     3781 aagctgtcct tccccgaggg cttcaagtgg gagcgcgtga tgaacttcga ggacggcggc
     3841 gtggtgaccg tgacccagga ctcctccctg caggacggcg agttcatcta caaggtgaag
     3901 ctgcgcggca ccaacttccc ctccgacggc cccgtaatgc agaagaagac catgggctgg
     3961 gaggcctcct ccgagcggat gtaccccgag gacggcgccc tgaagggcga gatcaagcag
     4021 aggctgaagc tgaaggacgg cggccactac gacgctgagg tcaagaccac ctacaaggcc
     4081 aagaagcccg tgcagctgcc cggcgcctac aacgtcaaca tcaagttgga catcacctcc
     4141 cacaacgagg actacaccat cgtggaacag tacgaacgcg ccgagggccg ccactccacc
     4201 ggcggcatgg acgagctgta caagtgagtt tggggccggc cgccagcaca gtggtcgatc
     4261 gaccaatgcc ctggctcaca aataccactg agatcttttt ccctctgcca aaaattatgg
     4321 ggacatcatg aagccccttg agcatctgac ttctggctaa taaaggaaat ttattttcat
     4381 tgcaatagtg tgttggaatt ttttgtgtct ctcactcgga aggacatatg ggagggcaaa
     4441 tcatttaaaa catcagaatg agtatttggt ttagagtttg gcaacatatg cccatatgct
     4501 ggctgccatg aacaaaggtt ggctataaag aggtcatcag tatatgaaac agccccctgc
     4561 tgtccattcc ttattccata gaaaagcctt gacttgaggt tagatttttt ttatattttg
     4621 ttttgtgtta tttttttctt taacatccct aaaattttcc ttacatgttt tactagccag
     4681 atttttcctc ctctcctgac tactcccagt catagctgtc cctcttctct tatggagatc
     4741 cctcgacctg cagcccaagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt
     4801 gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg
     4861 gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt
     4921 cgggaaacct gtcgtgccag cggatccgca tctcaattag tcagcaacca tagtcccgcc
     4981 cctaactccg cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg
     5041 ctgactaatt ttttttattt atgcagaggc cgaggccgcc tcggcctctg agctattcca
     5101 gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaaaagctaa cttgtttatt
     5161 gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt
     5221 ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgg
     5281 atccgctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct
     5341 cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat
     5401 cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga
     5461 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt
     5521 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt
     5581 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc
     5641 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
     5701 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct
     5761 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
     5821 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg
     5881 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc
     5941 ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta
     6001 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg
     6061 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
     6121 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg
     6181 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta
     6241 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
     6301 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     6361 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
     6421 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
     6481 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     6541 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
     6601 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     6661 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     6721 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     6781 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     6841 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
     6901 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     6961 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     7021 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     7081 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     7141 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
     7201 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
     7261 aagtgccacc tg
//
