LOCUS       sequence-452518-        4893 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 4893)
  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 4893)
  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: pML383_CCDC47-C-term_XTEN80-mEGFP-3xHA_pTMPLT
FEATURES             Location/Qualifiers
     source          1..4893
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     22..41
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(260..364)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     463..485
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     477..494
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     478..494
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             1585..1608
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             join(1624..1626,1627..2337)
                     /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(1666..1687)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1927..1946)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2274..2295
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2350..2430
                     /codon_start=1
                     /product="three tandem HA epitope tags"
                     /label=3xHA
                     /translation="YPYDVPDYAYPYDVPDYAYPYDVPDYA"
     primer_bind     complement(3275..3291)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3275..3291)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3288..3310)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     rep_origin      complement(3463..4051)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3543..3562)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    complement(4168..4192)
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             complement(join(4292..4893,1..190,191..259))
                     /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 cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca
       61 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca
      121 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg
      181 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac
      241 ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt
      301 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc
      361 cgcgcacatt tccccgaaaa gtgccagata cctgaaacaa aacccatcgt acggccaagg
      421 aagtctccaa taactgtgat ccaccacaag cgccagggtt ttcccagtca cgacgttgta
      481 aaacgacggc cagtcatgca taatccgcac gcatctggaa taaggaagtg ccattccgcc
      541 tgacctggag cttactggtg tcacttcgtg accactagtg gccaacagtg aagtcatact
      601 ttgacttgac ctctaaggct tctatcagtc cacgtaataa taacctgaac cttcttactc
      661 ttctatgtaa cttcctaatt cctaaataaa aatatgataa atttgataat ttttatcaca
      721 tttgataaat atgatgtctc aaatagagca ttatgaattt aagtattatg tgcaggtatg
      781 gtgtctgtgc taataagaaa tttgcaatga ataacctgat ggaacaagat tcttaaaatt
      841 tattgaactg aagtagaatg tatcttctgg cagttataca tttatgatat aatgccatca
      901 tgaatggtat ggtgaacatc aggctaagtg ttccctgggt acagaaaaaa atcaaaccag
      961 aataacaaag gctggttaat tatagtagtt tctttcaaaa ctgagatttt agtttacctt
     1021 tgcttcattt gaagttgatt taatttttat tcactaatgt agatattggc aattattttt
     1081 ataactatct tctagaatgt tatggtgcaa gcctaagttt ttgaataata ctataggcag
     1141 tattctaatt tggatttttt tttaacatat gtttcaagct taatttgaat tctctcgtcc
     1201 ccactccctt tctctattta ttaccccttc tttctctagg aggctgcatt gaggcgtgag
     1261 caaaagaagt tggaaaagaa gcaaatgaaa atgaaacaaa tcaaagtgaa agccatgagt
     1321 ggtggtactt ccgggggagg gccgagctct ggcgcacccc caccaagtgg agggtcacct
     1381 gccgggtccc caacatctac tgaagaaggc accagcgaat ccgcaacgcc cgagtcaggc
     1441 cctggtacct ccacagaacc atctgaaggt agtgcgcctg gttccccagc tggaagccct
     1501 acttccaccg aagaaggcac gtcaaccgaa ccaagtgaag gatctgcccc tgggaccagc
     1561 actgaaccat ctgagggtgg cggattggaa gttttgtttc aaggtccagg atctaacggc
     1621 agcgtgagca agggcgagga gctgttcacc ggggtggtgc ccatcctggt cgagctggac
     1681 ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg gcgagggcga tgccacctac
     1741 ggcaagctga ccctgaagtt catctgcacc accggcaagc tgcccgtgcc ctggcccacc
     1801 ctcgtgacca ccctgaccta cggcgtgcag tgcttcagcc gctaccccga ccacatgaag
     1861 cagcacgact tcttcaagtc cgccatgccc gaaggctacg tccaggagcg caccatcttc
     1921 ttcaaggacg acggcaacta caagacccgc gccgaggtga agttcgaggg cgacaccctg
     1981 gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg acggcaacat cctggggcac
     2041 aagctggagt acaactacaa cagccacaac gtctatatca tggccgacaa gcagaagaac
     2101 ggcatcaagg tgaacttcaa gatccgccac aacatcgagg acggcagcgt gcagctcgcc
     2161 gaccactacc agcagaacac ccccatcggc gacggccccg tgctgctgcc cgacaaccac
     2221 tacctgagca cccagtccaa gctgagcaaa gaccccaacg agaagcgcga tcacatggtc
     2281 ctgctggagt tcgtgaccgc cgccgggatc actctcggca tggacgagct gtacaaggct
     2341 ggagctggct acccatacga tgttccagat tacgcttatc cgtatgacgt cccggactat
     2401 gcatacccct acgatgtacc cgattacgcc ggtgcaggga gtggaagtta aagccatccc
     2461 agagatttga gttctgatgc cacctgtaag ctctgaattc acaggaaaca tgaaaaacgc
     2521 cagtccattt ctcaacctta aatttcagac agtcttgggc aactgagaaa tccttatttc
     2581 atcatctact ctgtttgggg tttggggttt tacagagatt gaagatacct ggaaagggct
     2641 ctgtttcaag aatttttttt tccagataat caaattattt tgattatttt ataaaaggaa
     2701 tgatctatga aatctgtgta ggttttaaat attttaaaaa ttataataca aatcatcagt
     2761 gcttttagta cttcagtgtt taaagaaata ccgtgaaatt tataggtaga taaccagatt
     2821 gttgcttttt gtttaaacca agcagttgaa atggctataa agactgactc taaaccaaga
     2881 ttctgcaaat aatgattgga attgcacaat aaacattgct tgatgttttc ttgtatgtct
     2941 acattaaact tgagaaaaag taaaaattag aacactgtat gtagtaatga aatttcaggg
     3001 acccagaaca taatgtagta tatgttttta ggtgggagat gctgataaca aaattaatag
     3061 gaagtctgta ggcattagga tactgacatg tacatggaaa attctaggga caggagcatc
     3121 attttttcct tacctgatac cacgaaccag tgacaacgtg aatgctgtat tttaagtggt
     3181 tgtatgttta ttttcttgcg tcagaagacc gtcagtgaag gctaggtgga ggctcagtga
     3241 tgataagtct gcgatggtgg atgcatgtgt catggtcata gctgtttcct gtgtgaaatt
     3301 gttatccgct cagagggcac aatcctattc cgcgctatcc gacaatctcc aagacattag
     3361 gtggagttca gttcggcgta tggcatatgt cgctggaaag aacatgtgag caaaaggcca
     3421 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
     3481 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
     3541 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
     3601 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag
     3661 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca
     3721 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa
     3781 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc
     3841 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag
     3901 aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg
     3961 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca
     4021 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc
     4081 tgacgctcta ttcaacaaag ccgccgtccc gtcaagtcag cgtaaatggg tagggggctt
     4141 caaatcgtcc tcgtgatacc aattcggagc ctgctttttt gtacaaactt gttgataatg
     4201 gcaattcaag gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct
     4261 aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta
     4321 tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa
     4381 ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagagccac
     4441 gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa
     4501 gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag
     4561 taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg
     4621 tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag
     4681 ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg
     4741 tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc
     4801 ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat
     4861 tctgagaata gtgtatgcgg cgaccgagtt gct
//
