LOCUS       sequence-451725-        4806 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Donor template for mScarlet insertion into the C-terminus of the 
            MAPRE1 locus. For growing microtubule tip visualization. To be 
            co-transfected with sgRNA plasmid px330-PITCh-MAPRE1 (Addgene 
            #207793).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4806)
  AUTHORS   Philip R, Sharma A, Matellan L, Erpf AC, Hsu WH, Tkach JM, Wyatt 
            HDM, Pelletier L
  TITLE     qTAG: an adaptable plasmid scaffold for CRISPR-based endogenous 
            tagging.
  JOURNAL   EMBO J. 2024 Dec 12. doi: 10.1038/s44318-024-00337-5.
  PUBMED    39668248
REFERENCE   2  (bases 1 to 4806)
  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"; journalName: "EMBO J. 
            2024 Dec 12. doi: 10.1038/s44318-024-00337-5."
COMMENT     Sequence Label: qTAG-C-Solo-mScarlet-MAPRE1
FEATURES             Location/Qualifiers
     source          1..4806
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     364..386
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     378..395
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     379..395
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             1138..1833
                     /codon_start=1
                     /product="bright monomeric red fluorescent protein evolved 
                     from a synthetic template (Bindels et al., 2016)"
                     /label=mScarlet
                     /translation="MVSKGEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFSWDILSPQFMYGSRAFTKHPADIPDYYKQSFPEGFKWERVMNFEDGG
                     AVTVTQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDGVLKGDIK
                     MALRLKDGGRYLADFKTTYKAKKPVQMPGAYNVDRKLDITSHNEDYTVVEQYERSEGRH
                     STGGMDELYK"
     primer_bind     complement(1271..1289)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1539..1558
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     complement(2585..2601)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2585..2601)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2598..2620)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2609..2625
                     /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(2633..2639,2640..2657,2658..2663))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2678..2699
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2816..2833)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2987..3575)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3067..3086)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3746..4537,4538..4606))
                     /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     4369..4388
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4607..4711)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4779..4797
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc
      241 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
      301 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt
      361 tttcccagtc acgacgttgt aaaacgacgg ccagtgaatt ctcaggaatt tttgagggaa
      421 attttttttt ttttttagtg acggggatct cactttattg cccaggctgg agtgcctggt
      481 ggtgcgatca tggctcactg cagctttgac ctcctgggtc aagcagtcct gcctcgaccc
      541 cgctggggag tagctgggac tacaggcatg tcccaccagg cttggctagt tttttttttt
      601 ttcgagacat tgtctcactg tgttacccag gctggtcttg aacttctggg ctcaagcctt
      661 ggccttccaa agtgctagga ttacaggcat gagccatcac acctggcctt ctttcttctt
      721 taaaaaacag ttttattgaa gtataactga cataataaac tgcacatatt taaaatgtac
      781 ctttgatgac atgggtgaaa ttaccaccat aatgaagaca atgtaaatat gcatcacttt
      841 caaaagtttg agggtaaatt tgactgtgct cagtttttgt ctttggtgag atgcaattct
      901 actacccaag aaataaaata gcagtcacct ggcctgtata gagccaggaa gaaccattgt
      961 ttttaagagg ctgtaagtat gaggaaagtg aactcacagt agaaatggat ctttcagtgg
     1021 ctttcccctc tcattttcct atttcaggaa ggctttgtga tacctgatga aggaggccca
     1081 caggaggagc aagaagagta tggaggaagc ggaggatctg gcggtaccgg cggaagcatg
     1141 gtgagcaagg gcgaggcagt gatcaaggag ttcatgcggt tcaaggtgca catggagggc
     1201 tccatgaacg gccacgagtt cgagatcgag ggcgagggcg agggccgccc ctacgagggc
     1261 acccagaccg ccaagctgaa ggtgaccaag ggtggccccc tgcccttctc ctgggacatc
     1321 ctgtcccctc agttcatgta cggctccagg gccttcacca agcaccccgc cgacatcccc
     1381 gactactata agcagtcctt ccccgagggc ttcaagtggg agcgcgtgat gaacttcgag
     1441 gacggcggcg ccgtgaccgt gacccaggac acctccctgg aggacggcac cctgatctac
     1501 aaggtgaagc tccgcggcac caacttccct cctgacggcc ccgtaatgca gaagaagaca
     1561 atgggctggg aagcgtccac cgagcggttg taccccgagg acggcgtgct gaagggcgac
     1621 attaagatgg ccctgcgcct gaaggacggc ggccgctacc tggcggactt caagaccacc
     1681 tacaaggcca agaagcccgt gcagatgccc ggcgcctaca acgtcgaccg caagttggac
     1741 atcacctccc acaacgagga ctacaccgtg gtggaacagt acgaacgctc cgagggccgc
     1801 cactccaccg gcggcatgga cgagctgtac aagggaggtt ccctcgagac cggtaagagc
     1861 aagtgataac agcctggacc agcagagcaa catcggaatt cttcactcca aatcatgtgc
     1921 ttaactgtaa aatactccct tttgttatcc ttagaggact cactggtttc ttttcataag
     1981 caaaaagtac ctcttcttaa agtgcacttt gcagacgttt cactcctttt ccaataagtt
     2041 tgagttagga gcttttacct tgtagcagag cagtattaac acctagttgg ttcacctgga
     2101 aaacagagag gctgaccgtg gggctcacca tgcggatgcg ggtcacaytg aatgctggag
     2161 agatgttatg taatatgctg aggtggcgac ctcagtggag aaatgtaaag actgaattga
     2221 attttaagct aatgtgaaat cagagaatgt tgtaataagt aaatgcctta agagtattta
     2281 aaatatgctt ccacatttca aaatataaaa tgtaacatga caagagattt tgcgtttgac
     2341 attgtgtctg ggaaggaagg gccagacctt ggaacctttg gaacctgctg tcaacaggtc
     2401 ttacagggct gcttgaaccc tcataggcct aggctttggt ctaaaaggaa catttaaaaa
     2461 gttgccctgt aaagttattt ggtgtcattg accaattgca tcccagctaa aaagcaagag
     2521 gcatcgttgc ctggataata gaggatgtgt ttcagccctg agatgtaagc ttggcgtaat
     2581 catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac
     2641 gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa
     2701 ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat
     2761 gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc
     2821 tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg
     2881 cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag
     2941 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
     3001 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
     3061 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
     3121 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
     3181 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
     3241 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
     3301 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
     3361 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
     3421 ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
     3481 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
     3541 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
     3601 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
     3661 aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta
     3721 tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag
     3781 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga
     3841 tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac
     3901 cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc
     3961 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta
     4021 gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac
     4081 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat
     4141 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa
     4201 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg
     4261 tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag
     4321 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc
     4381 cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct
     4441 caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat
     4501 cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg
     4561 ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc
     4621 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
     4681 tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg
     4741 tctaagaaac cattattatc atgacattaa cctataaaaa taggcgtatc acgaggccct
     4801 ttcgtc
//
