LOCUS       sequence-324263-        4875 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Allows for transcription of mKate2 control for injection into 
            zebrafish embryos for BiFC assays.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4875)
  AUTHORS   Don EK, Maschirow A, Radford RAW, Scherer NM, Vidal-Itriago A, Hogan
            A, Maurel C, Formella I, Stoddart JJ, Hall TE, Lee A, Shi B, Cole 
            NJ, Laird AS, Badrock AP, Chung RS, Morsch M
  TITLE     In vivo Validation of Bimolecular Fluorescence Complementation 
            (BiFC) to Investigate Aggregate Formation in Amyotrophic Lateral 
            Sclerosis (ALS).
  JOURNAL   Mol Neurobiol. 2021 May;58(5):2061-2074. doi: 
            10.1007/s12035-020-02238-0. Epub 2021 Jan 7.
  PUBMED    33415684
REFERENCE   2  (bases 1 to 4875)
  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: 
            "10.1007/s12035-020-02238-0"; journalName: "Mol Neurobiol"; date: 
            "2021-05"; volume: "58"; issue: "5"; pages: "2061-2074"
COMMENT     Sequence Label: pCS2plus-mKate2
FEATURES             Location/Qualifiers
     source          1..4875
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        35..53
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     35..52
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     regulatory      103..112
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    complement(820..919)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     polyA_signal    927..1061
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(951..970)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1005..1024
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(1174..1194)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(1174..1192)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1213..1229)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1213..1229)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1226..1248)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1237..1253
                     /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(1261..1267,1268..1285,1286..1291))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1306..1327
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1444..1461)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1615..2203)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1695..1714)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2374..3165,3166..3234))
                     /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     2997..3016
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3235..3339)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(3365..3820)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3502..3523)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3714..3733
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3891..4875
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
ORIGIN
        1 cgccattctg cctggggacg tcggagcaag cttgatttag gtgacactat agaatacaag
       61 ctacttgttc tttttgcagg atcccatcga ttcgaattcg ccgccaccat ggtgagcgag
      121 ctgatcaaag aaaacatgca catgaagctg tacatggaag gcaccgtgaa caaccaccac
      181 ttcaagtgca ccagcgaggg cgagggaaag ccttacgagg gaacccagac catgagaatc
      241 aaggctgtgg aaggcggacc tctgcctttc gccttcgaca tcctggccac cagctttatg
      301 tacggcagca agaccttcat caaccacacc cagggaatcc ctgacttctt caagcagagc
      361 ttccctgagg gattcacctg ggagagagtg accacctacg aggatggcgg cgtgctgacc
      421 gccacccagg acacaagcct gcaggacgga tgcctgatct acaacgtgaa gatcagaggc
      481 gtgaacttcc ctagcaacgg acctgtgatg cagaaaaaga ccctgggatg ggaggccagc
      541 accgagactc tgtaccctgc cgacggcgga ctggaaggca gagccgacat ggctctgaag
      601 ctggtcggag gcggacacct gatctgcaac ctgaaaacca cctacagaag caagaagccc
      661 gccaagaacc tgaagatgcc tggcgtgtac tacgtggaca gacgcctgga aagaatcaaa
      721 gaggccgaca aagagacata cgtcgagcag cacgaggtgg ccgtggccag atactgcaac
      781 ctgcctagca agctgggcca caaactgaac taaactagta cccagctttc ttgtacaaag
      841 ttggcattat aagaaagcat tgcttatcaa tttgttgcaa cgaacaggtc actatcagtc
      901 aaaataaaat cattatttga tacgtagatc cagacatgat aagatacatt gatgagtttg
      961 gacaaaccac aactagaatg cagtgaaaaa aatgctttat ttgtgaaatt tgtgatgcta
     1021 ttgctttatt tgtaaccatt ataagctgca ataaacaagt taacaacaac aattgcattc
     1081 attttatgtt tcaggttcag ggggaggtgt gggaggtttt ttaattcgcg gccgcggcgc
     1141 caatgcattg ggcccggtac ccagcttttg ttccctttag tgagggttaa ttgcgcgctt
     1201 ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca
     1261 caacatacga gccgggagca taaagtgtaa agcctggggt gcctaatgag tgagctaact
     1321 cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct
     1381 gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc
     1441 ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca
     1501 ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg
     1561 agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca
     1621 taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa
     1681 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc
     1741 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc
     1801 gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct
     1861 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg
     1921 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag
     1981 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta
     2041 cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg
     2101 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt
     2161 tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt
     2221 ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag
     2281 attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat
     2341 ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc
     2401 tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat
     2461 aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc
     2521 acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag
     2581 aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag
     2641 agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt
     2701 ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg
     2761 agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt
     2821 tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc
     2881 tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc
     2941 attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa
     3001 taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg
     3061 aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc
     3121 caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag
     3181 gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt
     3241 cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt
     3301 tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc
     3361 acctaaattg taagcgttaa tattttgtta aaattcgcgt taaatttttg ttaaatcagc
     3421 tcatttttta accaataggc cgaaatcggc aaaatccctt ataaatcaaa agaatagacc
     3481 gagatagggt tgagtgttgt tccagtttgg aacaagagtc cactattaaa gaacgtggac
     3541 tccaacgtca aagggcgaaa aaccgtctat cagggcgatg gcccactacg tgaaccatca
     3601 ccctaatcaa gttttttggg gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg
     3661 agcccccgat ttagagcttg acggggaaag ccggcgaacg tggcgagaaa ggaagggaag
     3721 aaagcgaaag gagcgggcgc tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc
     3781 accacacccg ccgcgcttaa tgcgccgcta cagggcgcgt cccattcgcc attcaggctg
     3841 cgcaactgtt gggaagggcg atcggtgcgg gcctcttcgc tattacgcca gtcgaccgcc
     3901 aattcaatat ggcgtatatg gactcatgcc aattcaatat ggtggatctg gacctgtgcc
     3961 aattcaatat ggcgtatatg gactcgtgcc aattcaatat ggtggatctg gaccccagcc
     4021 aattcaatat ggcggacttg gcaccatgcc aattcaatat ggcggacctg gcactgtgcc
     4081 aactggggag gggtctactt ggcacggtgc caagtttgag gaggggtctt ggccctgtgc
     4141 caagtccgcc atattgaatt ggcatggtgc caataatggc ggccatattg gctatatgcc
     4201 aggatcaata tataggcaat atccaatatg gccctatgcc aatatggcta ttggccaggt
     4261 tcaatactat gtattggccc tatgccatat agtattccat atatgggttt tcctattgac
     4321 gtagatagcc cctcccaatg ggcggtccca tataccatat atggggcttc ctaataccgc
     4381 ccatagccac tcccccattg acgtcaatgg tctctatata tggtctttcc tattgacgtc
     4441 atatgggcgg tcctattgac gtatatggcg cctcccccat tgacgtcaat tacggtaaat
     4501 ggcccgcctg gctcaatgcc cattgacgtc aataggacca cccaccattg acgtcaatgg
     4561 gatggctcat tgcccattca tatccgttct cacgccccct attgacgtca atgacggtaa
     4621 atggcccact tggcagtaca tcaatatcta ttaatagtaa cttggcaagt acattactat
     4681 tggaagtacg ccagggtaca ttggcagtac tcccattgac gtcaatggcg gtaaatggcc
     4741 cgcgatggct gccaagtaca tccccattga cgtcaatggg gaggggcaat gacgcaaatg
     4801 ggcgttccat tgacgtaaat gggcggtagg cgtgcctaat gggaggtcta tataagcaat
     4861 gctcgtttag ggaac
//
