LOCUS       sequence-456028-        4894 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 4894)
  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 4894)
  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: pML390_YWHAQ-C-term_XTEN80-mEGFP-3xHA_pTMPLT
FEATURES             Location/Qualifiers
     source          1..4894
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(122..226)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     325..347
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     339..356
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     340..356
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             1448..1471
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             join(1487..1489,1490..2200)
                     /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(1529..1550)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1790..1809)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2137..2158
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2213..2293
                     /codon_start=1
                     /product="three tandem HA epitope tags"
                     /label=3xHA
                     /translation="YPYDVPDYAYPYDVPDYAYPYDVPDYA"
     primer_bind     complement(3138..3154)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3138..3154)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3151..3173)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     rep_origin      complement(3326..3914)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3406..3425)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    complement(4031..4055)
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             complement(join(4155..4894,1..52,53..121))
                     /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     4778..4797
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
       61 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
      121 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
      181 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
      241 aagtgccaga tacctgaaac aaaacccatc gtacggccaa ggaagtctcc aataactgtg
      301 atccaccaca agcgccaggg ttttcccagt cacgacgttg taaaacgacg gccagtcatg
      361 cataatccgc acgcatctgg aataaggaag tgccattccg cctgacctgg agcttactgg
      421 tgtcacttct gtaataataa tgcagtagtg ctttcaagaa gagcattggg ctgctgggaa
      481 tagagactat cactttgctt attctgaaag tagagtgcct ttttaaaaaa gtttctcttt
      541 tttttttttt ttttttgaga aaggatctca ccctgttgcc taggctggag tgcagtgtca
      601 taatcatgga tcactccagc ctcaacctcc tggattcaag cagtcctcct gcctcagcct
      661 cctgagtagt taggactgta ggcacacacc accccgtacc tggccaattt tgtttttgaa
      721 attttttgta gacgtgggat cttgctatgt tgtccaggct gttgaactcc tggcctcatg
      781 cgatcttctc atcttggcct ccctacgtgc tgagattaca ggcgtgagcc actgtgcctg
      841 gccaaatgct ttttaaataa aaaggaagct ataaaaccat gctccagtgg tatatgtatg
      901 tatgtatgta tgtcattttg taagtgcata gtgaaaggtc tgcaaggatg tactttggga
      961 aggtgatggt ctttggggtt ttgagagaga acttcagtct tagtcaagat ctccattgtt
     1021 tgaggaattt actactcttg tagttaattt tgatataaca atactactta gtgtaattgc
     1081 ctaatctaga aatagtaact taatatgatt cttctttttc tagctttgga catcagacag
     1141 tgcaggagaa gaatgtgatg cggcagaagg ggctgaaaac agtggtggta cttccggggg
     1201 agggccgagc tctggcgcac ccccaccaag tggagggtca cctgccgggt ccccaacatc
     1261 tactgaagaa ggcaccagcg aatccgcaac gcccgagtca ggccctggta cctccacaga
     1321 accatctgaa ggtagtgcgc ctggttcccc agctggaagc cctacttcca ccgaagaagg
     1381 cacgtcaacc gaaccaagtg aaggatctgc ccctgggacc agcactgaac catctgaggg
     1441 tggcggattg gaagttttgt ttcaaggtcc aggatctaac ggcagcgtga gcaagggcga
     1501 ggagctgttc accggggtgg tgcccatcct ggtcgagctg gacggcgacg taaacggcca
     1561 caagttcagc gtgtccggcg agggcgaggg cgatgccacc tacggcaagc tgaccctgaa
     1621 gttcatctgc accaccggca agctgcccgt gccctggccc accctcgtga ccaccctgac
     1681 ctacggcgtg cagtgcttca gccgctaccc cgaccacatg aagcagcacg acttcttcaa
     1741 gtccgccatg cccgaaggct acgtccagga gcgcaccatc ttcttcaagg acgacggcaa
     1801 ctacaagacc cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc gcatcgagct
     1861 gaagggcatc gacttcaagg aggacggcaa catcctgggg cacaagctgg agtacaacta
     1921 caacagccac aacgtctata tcatggccga caagcagaag aacggcatca aggtgaactt
     1981 caagatccgc cacaacatcg aggacggcag cgtgcagctc gccgaccact accagcagaa
     2041 cacccccatc ggcgacggcc ccgtgctgct gcccgacaac cactacctga gcacccagtc
     2101 caagctgagc aaagacccca acgagaagcg cgatcacatg gtcctgctgg agttcgtgac
     2161 cgccgccggg atcactctcg gcatggacga gctgtacaag gctggagctg gctacccata
     2221 cgatgttcca gattacgctt atccgtatga cgtcccggac tatgcatacc cctacgatgt
     2281 acccgattac gccggtgcag ggagtggaag ttaaatccat acagagtgtc atccttcttt
     2341 ccttcaagaa acctttttac acatctccat tccttattcc acttggattt cctatagcaa
     2401 agaaacccat tcatgtgtat ggaatcaact gtttatagtc ttttcacact gcagctttgg
     2461 gaaaacttca ttccttgatt tgtgtttgtc ttggccttcc tggtgtgcag tactgctgta
     2521 gaaaagtatt aatagcttca tttcatataa acataagtaa ctcccaaaca cttatgtaga
     2581 ggactaaaaa tgtatctggt atttaagtaa tctgaaccag ttctgcaagt gactgtgttt
     2641 tgtattactg tgaaaataag aaaatgtagt taattacaat ttaaagagta ttccacataa
     2701 cttcttaatt tctacattcc ctcccttact cttcgggggt ttcctttcag taagcaactt
     2761 ttccatgctc ttaatgtatt cctttttagt aggaatccgg aagtattaga ttgaatggaa
     2821 aagcacttgc catctctgtc taggggtcac aaattgaaat ggctcctgta tcacatacgg
     2881 aggtcttgtg tatctgtggc aacagggagt ttccttattc actctttatt tgctgctgtt
     2941 taagttgcca acctcccctc ccaataaaaa ttcacttaca cctcctgcct ttgtagttct
     3001 ggtattcact ttactatgtg atagaagtag catgttgctg ccagaataca agcattgctt
     3061 tcgtcagaag accgtcagtg aaggctaggt ggaggctcag tgatgataag tctgcgatgg
     3121 tggatgcatg tgtcatggtc atagctgttt cctgtgtgaa attgttatcc gctcagaggg
     3181 cacaatccta ttccgcgcta tccgacaatc tccaagacat taggtggagt tcagttcggc
     3241 gtatggcata tgtcgctgga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
     3301 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
     3361 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
     3421 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     3481 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     3541 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     3601 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     3661 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     3721 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta
     3781 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     3841 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     3901 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct ctattcaaca
     3961 aagccgccgt cccgtcaagt cagcgtaaat gggtaggggg cttcaaatcg tcctcgtgat
     4021 accaattcgg agcctgcttt tttgtacaaa cttgttgata atggcaattc aaggatcttc
     4081 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     4141 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     4201 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     4261 ttaccatctg gccccagtgc tgcaatgata ccgcgagagc cacgctcacc ggctccagat
     4321 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     4381 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     4441 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
     4501 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     4561 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     4621 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     4681 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     4741 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
     4801 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     4861 ccgctgttga gatccagttc gatgtaaccc actc
//
