LOCUS       sequence-452480-        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: pML307_SEC61B-N-term_XTEN80-mEGFP-3xHA_pTMPLT
FEATURES             Location/Qualifiers
     source          1..4893
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(26..817,818..886))
                     /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     649..668
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(887..991)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     1090..1112
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1104..1121
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1105..1121
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             1963..2043
                     /codon_start=1
                     /product="three tandem HA epitope tags"
                     /label=3xHA
                     /translation="YPYDVPDYAYPYDVPDYAYPYDVPDYA"
     CDS             join(2056..2058,2059..2769)
                     /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(2098..2119)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2359..2378)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2706..2727
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2779..2802
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     primer_bind     complement(3902..3918)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3902..3918)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3915..3937)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     rep_origin      complement(4090..4678)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4170..4189)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    complement(4795..4819)
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
ORIGIN
        1 tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag
       61 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga
      121 tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagag ccacgctcac
      181 cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc
      241 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta
      301 gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac
      361 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat
      421 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa
      481 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg
      541 tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag
      601 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc
      661 cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct
      721 caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat
      781 cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg
      841 ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc
      901 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
      961 tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccagatacct
     1021 gaaacaaaac ccatcgtacg gccaaggaag tctccaataa ctgtgatcca ccacaagcgc
     1081 cagggttttc ccagtcacga cgttgtaaaa cgacggccag tcatgcataa tccgcacgca
     1141 tctggaataa ggaagtgcca ttccgcctga cctggagctt actggtgtca cttccagccg
     1201 ctcagcgccg cccacgccca ggtctgggtg gaggaacagc accgacggct tgggaaccga
     1261 gtcccgttcc cggccctgct cctccgccat ggccctggag ccgcaactgc accccgcacc
     1321 ctgatggggg tcttctgcgc aagctccgcg ctcgtagctc ccagctggcc actgcgggcc
     1381 gaccccgccc tgccgtacgt gcgtcagtta ggccacatca gcgcaaatct gtgagggtct
     1441 agtaactgcc tgagaaaata tcttgtctga ccccggttat atttttcctt cggtagggat
     1501 tggactttct gaaggacgtt gtgatccaaa ggaaggaggc cggaggactc tacttcccat
     1561 acagcaggta actaagttgt ctgtagcaga ctgtctacag gcatatcgtg agacgaccca
     1621 ggcgtccctg gggtcagaga ggaccttgcc tgcaagtccg ggggcggggc ctgagtcagt
     1681 ctcgccagct gccggtcttt cgggggctcc gtaactttct atccgtccgc gtcagcgcct
     1741 tgccaccctc atctccaata tggtatggcg gcccttccat gatccccgcc tctcccagaa
     1801 gccctgactc ctcctgcttt gcgccgtgct tttcctctgt agctcccttg cttcccccag
     1861 cctcgggtgt gggtgtctag gccggggttc tggggcaggc ctgccgcgct cacccgtctg
     1921 tctgcttgtc tccctctaca gcctagtggt ggtacttccg ggtacccata cgatgttcca
     1981 gattacgctt atccgtatga cgtcccggac tatgcatacc cctacgatgt acccgattac
     2041 gccgctggag ctggcgtgag caagggcgag gagctgttca ccggggtggt gcccatcctg
     2101 gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc
     2161 gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg
     2221 ccctggccca ccctcgtgac caccctgacc tacggcgtgc agtgcttcag ccgctacccc
     2281 gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag
     2341 cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag
     2401 ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac
     2461 atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat catggccgac
     2521 aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga ggacggcagc
     2581 gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg
     2641 cccgacaacc actacctgag cacccagtcc aagctgagca aagaccccaa cgagaagcgc
     2701 gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag
     2761 ctgtacaagg gtggcggatt ggaagttttg tttcaaggtc caggatctaa cggcagcgga
     2821 gggccgagct ctggcgcacc cccaccaagt ggagggtcac ctgccgggtc cccaacatct
     2881 actgaagaag gcaccagcga atccgcaacg cccgagtcag gccctggtac ctccacagaa
     2941 ccatctgaag gtagtgcgcc tggttcccca gctggaagcc ctacttccac cgaagaaggc
     3001 acgtcaaccg aaccaagtga aggatctgcc cctgggacca gcactgaacc atctgagggt
     3061 gcagggagtg gaagtggtcc gacccccagt ggcactaacg tgggatcctc agggcgctct
     3121 cccagcaaag cagtggccgc ccgggcggcg ggatccactg tccggcagag gtaaggaacc
     3181 ctgcagttcg ttcgcttcca gactcggaga taggacccag aacctcgctg attctggggt
     3241 ggagtcccta gcatgtgaag attgacaaag gcaaaatgag cttctagtga cgtggccgtg
     3301 ggagtagtta aaggcctttt gggaggaagg cgacattttt tttctcgttg ctcagtttag
     3361 ggcactactc ttaaaaaagg aaagttaaca aactggaata gagtcagaga taactttgag
     3421 aaaaccgatg tcattaaact ggtgtctctg gacctgaggt ttgcactcac atttccatct
     3481 ggcggcccca taagcaatct gtcctacaga taactcgtcc tacacaaaac ttagtctctt
     3541 ttcagctcag ctctctcact ctcaattata tctccttact tccatatggc actgttgtac
     3601 actcatttac tcagagccag aaacgtcagc gtcatcttgg atttttctta tgctctttct
     3661 ctctctagtc atatgccaga ctttaaactc tgcttgaaag ctttctcata agctctttcc
     3721 ttttcccttt ctactgcttt gcatttgcta cttaaccctt ttcttcaggc tgtttgcttt
     3781 ccagtccatc gttcgctctg ctgttactct tctgcgtagt ttctgcgtca gaagaccgtc
     3841 agtgaaggct aggtggaggc tcagtgatga taagtctgcg atggtggatg catgtgtcat
     3901 ggtcatagct gtttcctgtg tgaaattgtt atccgctcag agggcacaat cctattccgc
     3961 gctatccgac aatctccaag acattaggtg gagttcagtt cggcgtatgg catatgtcgc
     4021 tggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt
     4081 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag
     4141 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc
     4201 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc
     4261 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt
     4321 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt
     4381 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc
     4441 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa
     4501 gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa
     4561 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg
     4621 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga
     4681 agatcctttg atcttttcta cggggtctga cgctctattc aacaaagccg ccgtcccgtc
     4741 aagtcagcgt aaatgggtag ggggcttcaa atcgtcctcg tgataccaat tcggagcctg
     4801 cttttttgta caaacttgtt gataatggca attcaaggat cttcacctag atccttttaa
     4861 attaaaaatg aagttttaaa tcaatctaaa gta
//
