LOCUS       sequence-364103-        7361 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Stable expression of TMEM192-mGFP in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7361)
  AUTHORS   Yim WW, Yamamoto H, Mizushima N
  TITLE     Annexins A1 and A2 are recruited to larger lysosomal injuries 
            independently of ESCRTs to promote repair.
  JOURNAL   FEBS Lett. 2022 Mar 11. doi: 10.1002/1873-3468.14329.
  PUBMED    35274304
REFERENCE   2  (bases 1 to 7361)
  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: "FEBS Lett. 
            2022 Mar 11. doi: 10.1002/1873-3468.14329."
COMMENT     Sequence Label: pMRX-IB-TMEM192-mGFP
FEATURES             Location/Qualifiers
     source          1..7361
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        9..388
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     misc_feature    747..1104
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             1169..1585
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1515..1537
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1553..1569
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     misc_feature    1595..1969
                     /label=pol region
                     /note="Moloney murine leukemia virus (MMLV) pol region 
                     containing the splice acceptor site"
     primer_bind     1907..1923
                     /label=pBMN 5'
                     /note="MMLV sequence, for inserts in pBMN retroviral 
                     vector"
     regulatory      1996..2005
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      2000..2009
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2704..2706,2707..3420)
                     /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(2746..2767)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3007..3026)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3354..3375
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    3451..4017
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     misc_feature    3467..4019
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3611..3628)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3838..3857
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     LTR             4695..5164
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     primer_bind     complement(5294..5311)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5465..6053)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5545..5564)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6217..7008,7009..7077))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6840..6859
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7078..7182)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7250..7268
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 gccatgttga cattgattat tgactagtta ttaatagtaa tcaattacgg ggtcattagt
       61 tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg
      121 accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc
      181 aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc
      241 agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg
      301 gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat
      361 ctacgtatta gtcatcgcta ttaccatgct agagaaccat cagatgtttc cagggtgccc
      421 caaggacctg aaatgaccct gtgccttatt tgaactaacc aatcagttcg cttctcgctt
      481 ctgttcgcgc gcttctgctc cccgagctca ataaaagagc ccacaacccc tcactcggcg
      541 cgccagtcct ccgattgact gagtcgcccg ggtacccgtg tatccaataa accctcttgc
      601 agttgcatcc gacttgtggt ctcgctgttc cttgggaggg tctcctctga gtgattgact
      661 acccgtcagc gggggtcttt catttggggg ctcgtccggg atcgggagac ccctgcccag
      721 ggaccaccga cccaccaccg ggaggtaagc tggccagcaa cttatctgtg tctgtccgat
      781 tgtctagtgt ctatgactga ttttatgcgc ctgcgtcggt actagttagc taactagctc
      841 tgtatctggc ggacccgtgg tggaactgac gagttcggaa cacccggccg caaccctggg
      901 agacgtccca gggacttcgg gggccgtttt tgtggcccga cctgagtcca aaaatcccga
      961 tcgttttgga ctctttggtg cacccccctt agaggaggga tatgtggttc tggtaggaga
     1021 cgagaaccta aaacagttcc cgcctccgtc tgaatttttg ctttcggttt gggaccgaag
     1081 ccgcgccgcg cgtcttgtct gctgcagcat cgttctgtgt tgtctctgtc tgactgtgtt
     1141 tctgtatttg tctgaaaata tgggcccggg ccagactgtt accactccct taagtttgac
     1201 cttaggtcac tggaaagatg tcgagcggat cgctcacaac cagtcggtag atgtcaagaa
     1261 gagacgttgg gttaccttct gctctgcaga atggccaacc tttaacgtcg gatggccgcg
     1321 agacggcacc tttaaccgag acctcatcac ccaggttaag atcaaggtct tttcacctgg
     1381 cccgcatgga cacccagacc aggtccccta catcgtgacc tgggaagcct tggcttttga
     1441 cccccctccc tgggtcaagc cctttgtaca ccctaagcct ccgcctcctc ttcctccatc
     1501 cgccccgtct ctcccccttg aacctcctcg ttcgaccccg cctcgatcct ccctttatcc
     1561 agccctcact ccttctctag gcgcccccat atggccatat gagatcttat atggggcacc
     1621 cccgcccctt gtaaacttcc ctgaccctga catgacaaga gttactaaca gcccctctct
     1681 ccaagctcac ttacaggctc tctacttagt ccagcacgaa gtctggagac ctctggcggc
     1741 agcctaccaa gaacaactgg accgaccggt ggtacctcac ccttaccgag tcggcgacac
     1801 agtgtgggtc cgccgacacc agactaagaa cctagaacct cgctggaaag gaccttacac
     1861 agtcctgctg accaccccca ccgccctcaa agtagacggc atcgcagctt ggatacacgc
     1921 cgcccacgtg aaggctgccg accccggggg tggaccatcc tctagactgc cggatctgcc
     1981 atcgatataa agcttgccac catggcggcg gggggcagga tggaggacgg ttccttggat
     2041 atcacccaga gtattgaaga cgacccactt ctggatgccc agcttctccc acaccactca
     2101 ttacaagctc actttagacc ccgattccat cctcttccta cagtcatcat agtgaatctt
     2161 ctgtggttta ttcatctcgt gtttgttgtt ttagcatttt taacaggtgt gctttgttct
     2221 tatcctaatc caaatgagga caagtgccca ggaaattaca caaacccatt gaaagttcag
     2281 acggttataa tccttgggaa agttattttg tggattctcc atttactcct tgaatgctac
     2341 atccagtatc accacagcaa aatcagaaac cgaggctata acttgatcta ccgatcaaca
     2401 aggcatctca agagacttgc gttgatgata cagtcctctg gcaacacagt gcttctcctc
     2461 atactgtgca tgcagcactc cttcccagag cctggcagat tgtatcttga cctcattctg
     2521 gccatcttgg cactggaact catctgttcc ctgatatgtc tcctcattta cacagtgaaa
     2581 atccggagat ttaataaagc taaaccagag cctgatatac ttgaagaaga aaaaatctat
     2641 gcttacccca gcaatattac ctcggagact ggattcaggg ggaattcggg ctctggttca
     2701 ggcgtgagca agggcgagga gctgttcacc ggggtggtgc ccatcctggt cgagctggac
     2761 ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg gcgagggcga tgccacctac
     2821 ggcaagctga ccctgaagtt catctgcacc accggcaagc tgcccgtgcc ctggcccacc
     2881 ctcgtgacca ccctgaccta cggcgtgcag tgcttcagcc gctaccccga ccacatgaag
     2941 cagcacgact tcttcaagtc cgccatgccc gaaggctacg tccaggagcg caccatcttc
     3001 ttcaaggacg acggcaacta caagacccgc gccgaggtga agttcgaggg cgacaccctg
     3061 gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg acggcaacat cctggggcac
     3121 aagctggagt acaactacaa cagccacaac gtctatatca tggccgacaa gcagaagaac
     3181 ggcatcaagg tgaacttcaa gatccgccac aacatcgagg acggcagcgt gcagctcgcc
     3241 gaccactacc agcagaacac ccccatcggc gacggccccg tgctgctgcc cgacaaccac
     3301 tacctgagca cccagtccaa actgagcaaa gaccccaacg agaagcgcga tcacatggtc
     3361 ctgctggagt tcgtgaccgc cgccgggatc actctcggca tggacgagct gtacaagtaa
     3421 tgatttctcg agagcggccg ctacgtaaat tccgcccccc cccctaacgt tactggccga
     3481 agccgcttgg aataaggccg gtgtgcgttt gtctatatgt tattttccac catattgccg
     3541 tcttttggca atgtgagggc ccggaaacct ggccctgtct tcttgacgag cattcctagg
     3601 ggtctttccc ctctcgccaa aggaatgcaa ggtctgttga atgtcgtgaa ggaagcagtt
     3661 cctctggaag tttcttgaag acaaacaacg tctgtagcga ccctttgcag gcagcggaac
     3721 cccccacctg gcgacaggtg cctctgcggc caaaagccac gtgtataaga tacacctgca
     3781 aaggcggcac aaccccagtg ccacgttgtg agttggatag ttgtggaaag agtcaaatgg
     3841 ctctcctcaa gcgtattcaa caaggggctg aaggatgccc agaaggtacc ccattgtatg
     3901 ggatctgatc tggggcctcg gtgcacatgc tttacatgtg tttagtcgag gttaaaaaaa
     3961 cgtctaggcc ccccgaacca cggggacgtg gttttccttt gaaaaacacg atgataatat
     4021 ggccacaacc gtcgacatgg tcatgaaaac atttaacatt tctcaacaag atctagaatt
     4081 agtagaagta gcgacagaga agattacaat gctttatgag gataataaac atcatgtggg
     4141 agcggcaatt cgtacgaaaa caggagaaat catttcggca gtacatattg aagcgtatat
     4201 aggacgagta actgtttgtg cagaagccat tgcgattggt agtgcagttt cgaatggaca
     4261 aaaggatttt gacacgattg tagctgttag acacccttat tctgacgaag tagatagaag
     4321 tattcgagtg gtaagtcctt gtggtatgtg tagggagttg atttcagact atgcaccaga
     4381 ttgttttgtg ttaatagaaa tgaatggcaa gttagtcaaa actacgattg aagaactcat
     4441 tccactcaaa tatacccgaa attaagtcga ccgcccgccc cacgacccgc agcgcccgac
     4501 cgaaaggagc gcacgacccc atggctcgac gataaaataa aagattttat ttagtctcca
     4561 gaaaaagggg ggaatgaaag accccacctg taggtttggc aagctagctt aagtaacgcc
     4621 attttgcaag gcatggaaaa atacataact gagaatagag aagttcagat caaggtcagg
     4681 aacagatgga acagctgaat atgggccaaa caggatatct gtggtaagca gttcctgccc
     4741 cggctcaggg ccaagaacag atggaacagc tgaatatggg ccaaacagga tatctgtggt
     4801 aagcagttcc tgccccggct cagggccaag aacagatggt ccccagatgc ggtccagccc
     4861 tcagcagttt ctagagaacc atcagatgtt tccagggtgc cccaaggacc tgaaatgacc
     4921 ctgtgcctta tttgaactaa ccaatcagtt cgcttctcgc ttctgttcgc gcgcttctgc
     4981 tccccgagct caataaaaga gcccacaacc cctcactcgg ggcgccagtc ctccgattga
     5041 ctgagtcgcc cgggtacccg tgtatccaat aaaccctctt gcagttgcat ccgacttgtg
     5101 gtctcgctgt tccttgggag ggtctcctct gagtgattga ctacccgtca gcgggggtct
     5161 ttcacatgca gcatgtatca aaattaattt ggtttttttt cttaagtatt tacattaaat
     5221 ggccatagtt gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc
     5281 gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg
     5341 tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa
     5401 agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg
     5461 cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga
     5521 ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg
     5581 tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg
     5641 gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc
     5701 gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg
     5761 gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca
     5821 ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt
     5881 ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag
     5941 ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg
     6001 gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc
     6061 ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt
     6121 tggtcatgag attatcaaaa aggatcttca cctagatcct tttgcggccg gccgcaaatc
     6181 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc
     6241 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta
     6301 gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga
     6361 cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg
     6421 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc
     6481 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat
     6541 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag
     6601 gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat
     6661 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa
     6721 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa
     6781 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caacacggga
     6841 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg
     6901 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc
     6961 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
     7021 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact
     7081 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat
     7141 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt
     7201 gccacctgac gtctaagaaa ccattattat catgacatta acctataaaa ataggcgtat
     7261 cacgaggccc tttcgtcttc actataggag cttggcccat tgcatacgtt gtatccatat
     7321 cataatatgt acatttatat tggctcatgt ccaacattac c
//
