LOCUS       sequence-443428-        7499 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Stable expression of Evectin-2 in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7499)
  AUTHORS   Shinoda S, Sakai Y, Matsui T, Uematsu M, Koyama-Honda I, Sakamaki 
            JI, Yamamoto H, Mizushima N
  TITLE     Syntaxin 17 recruitment to mature autophagosomes is temporally 
            regulated by PI4P accumulation.
  JOURNAL   Elife. 2024 Jun 4;12:RP92189. doi: 10.7554/eLife.92189.
  PUBMED    38831696
REFERENCE   2  (bases 1 to 7499)
  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: "Elife."; 
            date: "2024-06-4"; volume: "12:RP92189. doi"; pages: " 
            10.7554/eLife.92189"
COMMENT     Sequence Label: pMRXIP-GFP-Evectin-2
FEATURES             Location/Qualifiers
     source          1..7499
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    667..1024
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             1089..1505
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1435..1457
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1473..1489
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     misc_feature    1515..1889
                     /label=pol region
                     /note="Moloney murine leukemia virus (MMLV) pol region 
                     containing the splice acceptor site"
     primer_bind     1827..1843
                     /label=pBMN 5'
                     /note="MMLV sequence, for inserts in pBMN retroviral 
                     vector"
     CDS             join(1920..1922,1923..1925,1926..2636)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1920..1922,1923..1925,1926..2636)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1965..1986)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2226..2245)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2573..2594
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    3350..3916
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     misc_feature    3366..3918
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3510..3527)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3737..3756
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             3930..4529
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(3930..3949)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4426..4446
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             4753..5222
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     primer_bind     complement(5352..5369)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5523..6111)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5603..5622)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6275..7066,7067..7135))
                     /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     6898..6917
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7136..7240)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7308..7326
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     enhancer        join(7428..7499,1..308)
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac gacccccgcc
       61 cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact ttccattgac
      121 gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa gtgtatcata
      181 tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg cattatgccc
      241 agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta gtcatcgcta
      301 ttaccatgct agagaaccat cagatgtttc cagggtgccc caaggacctg aaatgaccct
      361 gtgccttatt tgaactaacc aatcagttcg cttctcgctt ctgttcgcgc gcttctgctc
      421 cccgagctca ataaaagagc ccacaacccc tcactcggcg cgccagtcct ccgattgact
      481 gagtcgcccg ggtacccgtg tatccaataa accctcttgc agttgcatcc gacttgtggt
      541 ctcgctgttc cttgggaggg tctcctctga gtgattgact acccgtcagc gggggtcttt
      601 catttggggg ctcgtccggg atcgggagac ccctgcccag ggaccaccga cccaccaccg
      661 ggaggtaagc tggccagcaa cttatctgtg tctgtccgat tgtctagtgt ctatgactga
      721 ttttatgcgc ctgcgtcggt actagttagc taactagctc tgtatctggc ggacccgtgg
      781 tggaactgac gagttcggaa cacccggccg caaccctggg agacgtccca gggacttcgg
      841 gggccgtttt tgtggcccga cctgagtcca aaaatcccga tcgttttgga ctctttggtg
      901 cacccccctt agaggaggga tatgtggttc tggtaggaga cgagaaccta aaacagttcc
      961 cgcctccgtc tgaatttttg ctttcggttt gggaccgaag ccgcgccgcg cgtcttgtct
     1021 gctgcagcat cgttctgtgt tgtctctgtc tgactgtgtt tctgtatttg tctgaaaata
     1081 tgggcccggg ccagactgtt accactccct taagtttgac cttaggtcac tggaaagatg
     1141 tcgagcggat cgctcacaac cagtcggtag atgtcaagaa gagacgttgg gttaccttct
     1201 gctctgcaga atggccaacc tttaacgtcg gatggccgcg agacggcacc tttaaccgag
     1261 acctcatcac ccaggttaag atcaaggtct tttcacctgg cccgcatgga cacccagacc
     1321 aggtccccta catcgtgacc tgggaagcct tggcttttga cccccctccc tgggtcaagc
     1381 cctttgtaca ccctaagcct ccgcctcctc ttcctccatc cgccccgtct ctcccccttg
     1441 aacctcctcg ttcgaccccg cctcgatcct ccctttatcc agccctcact ccttctctag
     1501 gcgcccccat atggccatat gagatcttat atggggcacc cccgcccctt gtaaacttcc
     1561 ctgaccctga catgacaaga gttactaaca gcccctctct ccaagctcac ttacaggctc
     1621 tctacttagt ccagcacgaa gtctggagac ctctggcggc agcctaccaa gaacaactgg
     1681 accgaccggt ggtacctcac ccttaccgag tcggcgacac agtgtgggtc cgccgacacc
     1741 agactaagaa cctagaacct cgctggaaag gaccttacac agtcctgctg accaccccca
     1801 ccgccctcaa agtagacggc atcgcagctt ggatacacgc cgcccacgtg aaggctgccg
     1861 accccggggg tggaccatcc tctagactgc cggatcccag tgtggtggta cgggaattca
     1921 tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc catcctggtc gagctggacg
     1981 gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg
     2041 gcaagctgac cctgaagttc atctgcacca ccggcaagct gcccgtgccc tggcccaccc
     2101 tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg ctaccccgac cacatgaagc
     2161 agcacgactt cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc accatcttct
     2221 tcaaggacga cggcaactac aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg
     2281 tgaaccgcat cgagctgaag ggcatcgact tcaaggagga cggcaacatc ctggggcaca
     2341 agctggagta caactacaac agccacaacg tctatatcat ggccgacaag cagaagaacg
     2401 gcatcaaggt gaacttcaag atccgccaca acatcgagga cggcagcgtg cagctcgccg
     2461 accactacca gcagaacacc cccatcggcg acggccccgt gctgctgccc gacaaccact
     2521 acctgagcac ccagtccgcc ctgagcaaag accccaacga gaagcgcgat cacatggtcc
     2581 tgctggagtt cgtgaccgcc gccgggatca ctctcggcat ggacgagctg tacaagtccg
     2641 gccggactca gatctcgatg gcgtttgtga agagtggctg gttgctgcga cagagtacta
     2701 ttttgaagcg ctggaagaag aactggtttg atctgtggtc ggatggtcac ctgatctatt
     2761 atgatgacca gactcggcag aatatcgagg ataaggtcca catgccaatg gactgcatca
     2821 acatccgcac ggggcaggaa tgtcgggata ctcagccccc ggatggaaag tcaaaagact
     2881 gcatgctcca gattgtttgt cgagatggga aaacaattag tctttgtgca gaaagcacag
     2941 atgattgctt ggcctggaaa tttacactcc aagattctag gacaaacgtc gacggtacca
     3001 tggcgtttgt gaagagtggc tggttgctgc gacagagtac tattttgaag cgctggaaga
     3061 agaactggtt tgatctgtgg tcggatggtc acctgatcta ttatgatgac cagactcggc
     3121 agaatatcga ggataaggtc cacatgccaa tggactgcat caacatccgc acggggcagg
     3181 aatgtcggga tactcagccc ccggatggaa agtcaaaaga ctgcatgctc cagattgttt
     3241 gtcgagatgg gaaaacaatt agtctttgtg cagaaagcac agatgattgc ttggcctgga
     3301 aatttacact ccaagattct aggacaaact aggcggccgc tacgtaaatt cckccccccc
     3361 ccctaacgtt actggccgaa gccgcttgga ataaggccgg tgtgcgtttg tctatatgtt
     3421 attttccacc atattgccgt cttttggcaa tgtgagggcc cggaaacctg gccctgtctt
     3481 cttgacgagc attcctaggg gtctttcccc tctcgccaaa ggaatgcaag gtctgttgaa
     3541 tgtcgtgaag gaagcagttc ctctggaagc ttcttgaaga caaacaacgt ctgtagcgac
     3601 cctttgcagg cagcggaacc ccccacctgg cgacaggtgc ctctgcggcc aaaagccacg
     3661 tgtataagat acacctgcaa aggcggcaca accccagtgc cacgttgtga gttggatagt
     3721 tgtggaaaga gtcaaatggc tctcctcaag cgtattcaac aaggggctga aggatgccca
     3781 gaaggtaccc cattgtatgg gatctgatct ggggcctcgg tgcacatgct ttacatgtgt
     3841 ttagtcgagg ttaaaaaaac gtctaggccc cccgaaccac ggggacgtgg ttttcctttg
     3901 aaaaacacga tgataatatg gccacaacca tgaccgagta caagcccacg gtgcgcctcg
     3961 ccacccgcga cgacgtcccc cgggccgtac gcaccctcgc cgccgcgttc gccgactacc
     4021 ccgccacgcg ccacaccgtc gacccggacc gccacatcga gcgggtcacc gagctgcaag
     4081 aactcttcct cacgcgcgtc gggctcgaca tcggcaaggt gtgggtcgcg gacgacggcg
     4141 ccgcggtggc ggtctggacc acgccggaga gcgtcgaagc gggggcggtg ttcgccgaga
     4201 tcggcccgcg catggccgag ttgagcggtt cccggctggc cgcgcagcaa cagatggaag
     4261 gcctcctggc gccgcaccgg cccaaggagc ccgcgtggtt cctggccacc gtcggcgtct
     4321 cgcccgacca ccagggcaag ggtctgggca gcgccgtcgt gctccccgga gtggaggcgg
     4381 ccgagcgcgc cggggtgccc gccttcctgg agacctccgc gccccgcaac ctccccttct
     4441 acgagcggct cggcttcacc gtcaccgccg acgtcgaggt gcccgaagga ccgcgcacct
     4501 ggtgcatgac ccgcaagccc ggtgcctgac gcccgcccca cgacccgcag cgcccgaccg
     4561 aaaggagcgc acgaccccat ggctcgacga taaaataaaa gattttattt agtctccaga
     4621 aaaagggggg aatgaaagac cccacctgta ggtttggcaa gctagcttaa gtaacgccat
     4681 tttgcaaggc atggaaaaat acataactga gaatagagaa gttcagatca aggtcaggaa
     4741 cagatggaac agctgaatat gggccaaaca ggatatctgt ggtaagcagt tcctgccccg
     4801 gctcagggcc aagaacagat ggaacagctg aatatgggcc aaacaggata tctgtggtaa
     4861 gcagttcctg ccccggctca gggccaagaa cagatggtcc ccagatgcgg tccagccctc
     4921 agcagtttct agagaaccat cagatgtttc cagggtgccc caaggacctg aaatgaccct
     4981 gtgccttatt tgaactaacc aatcagttcg cttctcgctt ctgttcgcgc gcttctgctc
     5041 cccgagctca ataaaagagc ccacaacccc tcactcgggg cgccagtcct ccgattgact
     5101 gagtcgcccg ggtacccgtg tatccaataa accctcttgc agttgcatcc gacttgtggt
     5161 ctcgctgttc cttgggaggg tctcctctga gtgattgact acccgtcagc gggggtcttt
     5221 cacatgcagc atgtatcaaa attaatttgg ttttttttct taagtattta cattaaatgg
     5281 ccatagttgc attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc
     5341 tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta
     5401 tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag
     5461 aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg
     5521 tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg
     5581 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg
     5641 cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga
     5701 agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc
     5761 tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt
     5821 aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact
     5881 ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg
     5941 cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt
     6001 accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt
     6061 ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct
     6121 ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg
     6181 gtcatgagat tatcaaaaag gatcttcacc tagatccttt tgcggccggc cgcaaatcaa
     6241 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     6301 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     6361 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
     6421 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     6481 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     6541 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
     6601 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     6661 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     6721 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     6781 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     6841 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca acacgggata
     6901 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     6961 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     7021 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     7081 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
     7141 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     7201 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
     7261 cacctgacgt ctaagaaacc attattatca tgacattaac ctataaaaat aggcgtatca
     7321 cgaggccctt tcgtcttcac tcgaggagct tggcccattg catacgttgt atccatatca
     7381 taatatgtac atttatattg gctcatgtcc aacattaccg ccatgttgac attgattatt
     7441 gactagttat taatagtaat caattacggg gtcattagtt catagcccat atatggagt
//
