LOCUS       sequence-456037-        4382 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 4382)
  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 4382)
  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: pML484_RPL36-C-term_XTEN80-mEGFP-3xHA_pTMPLT
FEATURES             Location/Qualifiers
     source          1..4382
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     60..79
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(298..402)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     501..523
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     515..532
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     516..532
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             1382..1405
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             join(1421..1423,1424..2134)
                     /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(1463..1484)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1724..1743)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2071..2092
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2147..2227
                     /codon_start=1
                     /product="three tandem HA epitope tags"
                     /label=3xHA
                     /translation="YPYDVPDYAYPYDVPDYAYPYDVPDYA"
     primer_bind     complement(2802..2818)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2802..2818)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2815..2837)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     rep_origin      complement(2990..3578)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3070..3089)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    complement(3695..3719)
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             complement(join(3819..4382,1..228,229..297))
                     /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"
ORIGIN
        1 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga
       61 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg
      121 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc
      181 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
      241 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact
      301 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat
      361 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt
      421 gccagatacc tgaaacaaaa cccatcgtac ggccaaggaa gtctccaata actgtgatcc
      481 accacaagcg ccagggtttt cccagtcacg acgttgtaaa acgacggcca gtcatgcata
      541 atccgcacgc atctggaata aggaagtgcc attccgcctg acctggagct tactggtgtc
      601 acttcgcgtc ctgccagtgt tgcggagact gggacttagg gaggatggga gctgctccgg
      661 gcgcggcgaa aagcgctagg cgttaatacc ttggttctca cagctaccca cagaggggag
      721 gaagtagcca gggaaatcgc ggcagcgcga gagaagctgc ttaactagaa tgcagggatc
      781 cccctaccct gacggccgcc cctttccccc ctagcgtctg accaaacaca ccaagttcgt
      841 gcgggacatg attcgggagg tgtgtggctt tgccccgtac gagcggcgcg ccatggagtt
      901 actgaaggtc tccaaggaca aacgggccct caaatttatc aagaaaaggg taggtgggcg
      961 ctgccggccg aggggcgggg tgggatggga gtcagggccg ggctgacggc ggcctcgtcc
     1021 ctggcaggtg gggacgcaca tccgcgccaa gaggaagcgg gaggagctga gcaacgtact
     1081 ggccgccatg aggaaagctg ctgccaagaa agactctggt ggtacttccg ggggagggcc
     1141 gagctctggc gcacccccac caagtggagg gtcacctgcc gggtccccaa catctactga
     1201 agaaggcacc agcgaatccg caacgcccga gtcaggccct ggtacctcca cagaaccatc
     1261 tgaaggtagt gcgcctggtt ccccagctgg aagccctact tccaccgaag aaggcacgtc
     1321 aaccgaacca agtgaaggat ctgcccctgg gaccagcact gaaccatctg agggtggcgg
     1381 attggaagtt ttgtttcaag gtccaggatc taacggcagc gtgagcaagg gcgaggagct
     1441 gttcaccggg gtggtgccca tcctggtcga gctggacggc gacgtaaacg gccacaagtt
     1501 cagcgtgtcc ggcgagggcg agggcgatgc cacctacggc aagctgaccc tgaagttcat
     1561 ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc gtgaccaccc tgacctacgg
     1621 cgtgcagtgc ttcagccgct accccgacca catgaagcag cacgacttct tcaagtccgc
     1681 catgcccgaa ggctacgtcc aggagcgcac catcttcttc aaggacgacg gcaactacaa
     1741 gacccgcgcc gaggtgaagt tcgagggcga caccctggtg aaccgcatcg agctgaaggg
     1801 catcgacttc aaggaggacg gcaacatcct ggggcacaag ctggagtaca actacaacag
     1861 ccacaacgtc tatatcatgg ccgacaagca gaagaacggc atcaaggtga acttcaagat
     1921 ccgccacaac atcgaggacg gcagcgtgca gctcgccgac cactaccagc agaacacccc
     1981 catcggcgac ggccccgtgc tgctgcccga caaccactac ctgagcaccc agtccaagct
     2041 gagcaaagac cccaacgaga agcgcgatca catggtcctg ctggagttcg tgaccgccgc
     2101 cgggatcact ctcggcatgg acgagctgta caaggctgga gctggctatc catacgatgt
     2161 tccagattac gcttatccgt atgacgtccc ggactatgca tacccctatg atgtacccga
     2221 ctacgccggt gcagggagtg gaagttgagc cggtccggtg cggtctggct gttatattga
     2281 agtgctacac actagccctt tctctcctgc tgtccgtggg tgggtgtggg tgtgtcgggg
     2341 gcccgcagtc ccctgtctgg tgcccgctct gagccacacc ctctccgggt gctgcctggt
     2401 cgtgaatcaa aagccgtggc ccgcccaccc ttcccggggc agcaggtgag gaagccgccg
     2461 tactgcaaat gactttaatc attaaatagc ttctatgcca cactctgatt aagccgactg
     2521 aggtccctgg gatctgggtc actggaccga gctgctcgct cggtggctcc actgccaggt
     2581 ccgggcgcgc tccccacagc gctcagttct ggcccagaca gggcctgaca tccagttctg
     2641 gcccagacag ggcctgacat ccgccgcctg cagtcccggg gtggccgtca ccgttccacg
     2701 gccagcgcct ctgcctgctc gtccgcgtca gaagaccgtc agtgaaggct aggtggaggc
     2761 tcagtgatga taagtctgcg atggtggatg catgtgtcat ggtcatagct gtttcctgtg
     2821 tgaaattgtt atccgctcag agggcacaat cctattccgc gctatccgac aatctccaag
     2881 acattaggtg gagttcagtt cggcgtatgg catatgtcgc tggaaagaac atgtgagcaa
     2941 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
     3001 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
     3061 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
     3121 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
     3181 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
     3241 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
     3301 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
     3361 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     3421 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     3481 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
     3541 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
     3601 cggggtctga cgctctattc aacaaagccg ccgtcccgtc aagtcagcgt aaatgggtag
     3661 ggggcttcaa atcgtcctcg tgataccaat tcggagcctg cttttttgta caaacttgtt
     3721 gataatggca attcaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa
     3781 tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag
     3841 gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg
     3901 tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga
     3961 gagccacgct caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag
     4021 cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa
     4081 gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc
     4141 atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca
     4201 aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg
     4261 atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat
     4321 aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc
     4381 aa
//
