LOCUS       sequence-453984-        4193 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Homology repair template for genome engineering. Organelle 
            endogenous marker expressing GFP and epitope tags for mass 
            spectrometry..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4193)
  AUTHORS   Hein MY, Peng D, Todorova V, McCarthy F, Kim K, Liu C, Savy L, 
            Januel C, Baltazar-Nunez R, Sekhar M, Vaid S, Bax S, Vangipuram M, 
            Burgess J, Njoya L, Wang E, Ivanov IE, Byrum JR, Pradeep S, Gonzalez
            CG, Aniseia Y, Creery JS, McMorrow AH, Sunshine S, Yeung-Levy S, 
            DeFelice BC, Mehta SB, Itzhak DN, Elias JE, Leonetti MD
  TITLE     Global organelle profiling reveals subcellular localization and 
            remodeling at proteome scale.
  JOURNAL   Cell. 2024 Dec 26:S0092-8674(24)01344-8. doi: 
            10.1016/j.cell.2024.11.028.
  PUBMED    39742809
REFERENCE   2  (bases 1 to 4193)
  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: "Cell. 2024 
            Dec 26:S0092-8674(24)01344-8. doi: 10.1016/j.cell.2024.11.028."
COMMENT     Sequence Label: pML454_G3BP1-N-term_XTEN80-mEGFP-3xHA_pTMPLT
FEATURES             Location/Qualifiers
     source          1..4193
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(57..73)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(57..73)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(70..92)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     rep_origin      complement(245..833)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(325..344)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    complement(950..974)
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             complement(join(1074..1865,1866..1934))
                     /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     1697..1716
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1935..2039)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2138..2160
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2152..2169
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2153..2169
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             2661..2741
                     /codon_start=1
                     /product="three tandem HA epitope tags"
                     /label=3xHA
                     /translation="YPYDVPDYAYPYDVPDYAYPYDVPDYA"
     CDS             join(2754..2756,2757..3467)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(2796..2817)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3057..3076)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3404..3425
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             3477..3500
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
ORIGIN
        1 aggctaggtg gaggctcagt gatgataagt ctgcgatggt ggatgcatgt gtcatggtca
       61 tagctgtttc ctgtgtgaaa ttgttatccg ctcagagggc acaatcctat tccgcgctat
      121 ccgacaatct ccaagacatt aggtggagtt cagttcggcg tatggcatat gtcgctggaa
      181 agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg
      241 cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga
      301 ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg
      361 tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg
      421 gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc
      481 gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg
      541 gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca
      601 ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt
      661 ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag
      721 ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg
      781 gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc
      841 ctttgatctt ttctacgggg tctgacgctc tattcaacaa agccgccgtc ccgtcaagtc
      901 agcgtaaatg ggtagggggc ttcaaatcgt cctcgtgata ccaattcgga gcctgctttt
      961 ttgtacaaac ttgttgataa tggcaattca aggatcttca cctagatcct tttaaattaa
     1021 aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa
     1081 tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc
     1141 tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct
     1201 gcaatgatac cgcgagagcc acgctcaccg gctccagatt tatcagcaat aaaccagcca
     1261 gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt
     1321 aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt
     1381 gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc
     1441 ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc
     1501 tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt
     1561 atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact
     1621 ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc
     1681 ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt
     1741 ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg
     1801 atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct
     1861 gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa
     1921 tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt
     1981 ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc
     2041 acatttcccc gaaaagtgcc agatacctga aacaaaaccc atcgtacggc caaggaagtc
     2101 tccaataact gtgatccacc acaagcgcca gggttttccc agtcacgacg ttgtaaaacg
     2161 acggccagtc atgcataatc cgcacgcatc tggaataagg aagtgccatt ccgcctgacc
     2221 tggagcttac tggtgtcact tcggttgtgg tgagccgaga tcgcaccact gcactccagc
     2281 ctgggcaaca gagagactct gtctcaaaaa caaaacaaga aaattatggt gggaagtatg
     2341 ttggaaatat atacctttga ggttcttacg tcttgaatat tggggataat gatgaattct
     2401 tcataatgat ttcccccttt tgtattcata tgattgcaat acatttgtat aaaatctcaa
     2461 ctggtctgta tgggctctgt ttgtttttat gatgtttgtt catgttttaa acgacttgct
     2521 gaaatgatct tgtctctgag ttggtttcag ctaaatgatt cggtcttttc ccctgtgttt
     2581 gatccttcag gtttggacat atttgactct tttcccccca ggttgaattg accaaagcaa
     2641 tgtctggtgg tacttccggg tatccatacg atgttccaga ttacgcttat ccgtatgacg
     2701 tcccggacta tgcatacccc tatgatgtac ccgactacgc cgctggagct ggcgtgagca
     2761 agggcgagga gctgttcacc ggggtggtgc ccatcctggt cgagctggac ggcgacgtaa
     2821 acggccacaa gttcagcgtg tccggcgagg gcgagggcga tgccacctac ggcaagctga
     2881 ccctgaagtt catctgcacc accggcaagc tgcccgtgcc ctggcccacc ctcgtgacca
     2941 ccctgaccta cggcgtgcag tgcttcagcc gctaccccga ccacatgaag cagcacgact
     3001 tcttcaagtc cgccatgccc gaaggctacg tccaggagcg caccatcttc ttcaaggacg
     3061 acggcaacta caagacccgc gccgaggtga agttcgaggg cgacaccctg gtgaaccgca
     3121 tcgagctgaa gggcatcgac ttcaaggagg acggcaacat cctggggcac aagctggagt
     3181 acaactacaa cagccacaac gtctatatca tggccgacaa gcagaagaac ggcatcaagg
     3241 tgaacttcaa gatccgccac aacatcgagg acggcagcgt gcagctcgcc gaccactacc
     3301 agcagaacac ccccatcggc gacggccccg tgctgctgcc cgacaaccac tacctgagca
     3361 cccagtccaa gctgagcaaa gaccccaacg agaagcgcga tcacatggtc ctgctggagt
     3421 tcgtgaccgc cgccgggatc actctcggca tggacgagct gtacaagggt ggcggattgg
     3481 aagttttgtt tcaaggtcca ggatctaacg gcagcggagg gccgagctct ggcgcacccc
     3541 caccaagtgg agggtcacct gccgggtccc caacatctac tgaagaaggc accagcgaat
     3601 ccgcaacgcc cgagtcaggc cctggtacct ccacagaacc atctgaaggt agtgcgcctg
     3661 gttccccagc tggaagccct acttccaccg aagaaggcac gtcaaccgaa ccaagtgaag
     3721 gatctgcccc tgggaccagc actgaaccat ctgagggtgc agggagtgga agtgtcatgg
     3781 aaaaaccttc accgttactc gtcgggcggg aatttgtgag acagtattac acactgctga
     3841 accaggcccc agacatgctg cataggtaag acatttttct cctgcatcat ctaatgctgt
     3901 cttttagtat gtggtactta tctttgagat tctcccactc atcatcttat gctttgtagg
     3961 gttgttgcat aatacttatt tacatataaa tatcccccaa ttttttattt gagacagagt
     4021 ctcgccctgt tgtccagact ggagtgcagt ggtgcgatct ggactgactg caacgtccac
     4081 ctccccagtt taagtgattc ttctgcccca gcctcctgag tagctgggac cacaggtgcc
     4141 cacaaccatg cccggctaat ttttgtattt tttcgtcaga agaccgtcag tga
//
