LOCUS       sequence-420141-        6135 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of N-terminally tagged GABARAP with mTagBFP2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6135)
  AUTHORS   Uffing A, Gold L, Gensch T, Weiergraber OH, Hoffmann S, Willbold D
  TITLE     Highlighting the hidden: monitoring the avidity-driven association 
            of a fluorescent GABARAP tandem with microtubules in living cells.
  JOURNAL   Autophagy Rep. 2024 May 16;3(1):2348899. doi: 
            10.1080/27694127.2024.2348899. eCollection 2024.
  PUBMED    40395516
REFERENCE   2  (bases 1 to 6135)
  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: "Autophagy 
            Rep."; date: "2024-05-16"; volume: "3(1):2348899. doi"; pages: " 
            10.1080/27694127.2024.2348899. eCollection 2024"
COMMENT     Sequence Label: pcDNA5/FRT/TO mTagBFP2-GABARAP
FEATURES             Location/Qualifiers
     source          1..6135
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    820..838
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    841..859
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     869..893
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             976..1686
                     /codon_start=1
                     /product="enhanced monomeric blue fluorescent protein 
                     (Subach et al., 2011)"
                     /label=mTagBFP2
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTM
                     RIKVVEGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGG
                     VLTATQDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRND
                     MALKLVGGSHLIANAKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAV
                     ARYCDLPSKLGHKLN"
     primer_bind     complement(2087..2104)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2093..2317
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2450..2469)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    2601..2648
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             2656..3678
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             2656..3678
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     polyA_signal    3808..3929
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3845..3864)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3899..3918
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3978..3994)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3978..3994)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3991..4013)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4002..4018
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(4026..4032,4033..4050,4051..4056))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4071..4092
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4209..4226)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4380..4968)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4460..4479)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5139..5930,5931..5999))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     PPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5762..5781
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6000..6104)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc
      841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga
      901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga
      961 ctctagcgtt taaacatggt gtctaagggc gaagagctga ttaaggagaa catgcacatg
     1021 aagctgtaca tggagggcac cgtggacaac catcacttca agtgcacatc cgagggcgaa
     1081 ggcaagccct acgagggcac ccagaccatg agaatcaagg tggtcgaggg cggccctctc
     1141 cccttcgcct tcgacatcct ggctactagc ttcctctacg gcagcaagac cttcatcaac
     1201 cacacccagg gcatccccga cttcttcaag cagtccttcc ctgagggctt cacatgggag
     1261 agagtcacca catacgaaga cgggggcgtg ctgaccgcta cccaggacac cagcctccag
     1321 gacggctgcc tcatctacaa cgtcaagatc agaggggtga acttcacatc caacggccct
     1381 gtgatgcaga agaaaacact cggctgggag gccttcaccg agacgctgta ccccgctgac
     1441 ggcggcctgg aaggcagaaa cgacatggcc ctgaagctcg tgggcgggag ccatctgatc
     1501 gcaaacgcca agaccacata tagatccaag aaacccgcta agaacctcaa gatgcctggc
     1561 gtctactatg tggactacag actggaaaga atcaaggagg ccaacaacga gacctacgtc
     1621 gagcagcacg aggtggcagt ggccagatac tgcgacctcc ctagcaaact ggggcacaag
     1681 cttaattccg gactcagatc tcgagctatg aagttcgtgt acaaagaaga gcatccgttc
     1741 gagaagcgcc gctctgaggg cgagaagatc cgaaagaaat acccggaccg ggtgccggtg
     1801 atagtagaaa aggctcccaa agctcggata ggagacctgg acaaaaagaa atacctggtg
     1861 ccttctgatc tcacagttgg tcagttctac ttcttgatcc ggaagcgaat tcatctccga
     1921 gctgaggatg ccttgttttt ctttgtcaac aatgtcattc cacccaccag tgccacaatg
     1981 ggtcagctgt accaggaaca ccatgaagaa gacttctttc tctacattgc ctacagtgac
     2041 gaaagtgtct acggtctgta agggcccact taaacccgct gatcagcctc gactgtgcct
     2101 tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt
     2161 gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg
     2221 tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac
     2281 aatagcaggc atgctgggga tgcggtgggc tctatggctt ctgaggcgga aagaaccagc
     2341 tggggctcta gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg
     2401 gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct
     2461 ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg
     2521 ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag
     2581 ggtgatggtt cacgtaccta gaagttccta ttccgaagtt cctattctct agaaagtata
     2641 ggaacttcct tggccaaaaa gcctgaactc accgcgacgt ctgtcgagaa gtttctgatc
     2701 gaaaagttcg acagcgtctc cgacctgatg cagctctcgg agggcgaaga atctcgtgct
     2761 ttcagcttcg atgtaggagg gcgtggatat gtcctgcggg taaatagctg cgccgatggt
     2821 ttctacaaag atcgttatgt ttatcggcac tttgcatcgg ccgcgctccc gattccggaa
     2881 gtgcttgaca ttggggaatt cagcgagagc ctgacctatt gcatctcccg ccgtgcacag
     2941 ggtgtcacgt tgcaagacct gcctgaaacc gaactgcccg ctgttctgca gccggtcgcg
     3001 gaggccatgg atgcgatcgc tgcggccgat cttagccaga cgagcgggtt cggcccattc
     3061 ggaccgcaag gaatcggtca atacactaca tggcgtgatt tcatatgcgc gattgctgat
     3121 ccccatgtgt atcactggca aactgtgatg gacgacaccg tcagtgcgtc cgtcgcgcag
     3181 gctctcgatg agctgatgct ttgggccgag gactgccccg aagtccggca cctcgtgcac
     3241 gcggatttcg gctccaacaa tgtcctgacg gacaatggcc gcataacagc ggtcattgac
     3301 tggagcgagg cgatgttcgg ggattcccaa tacgaggtcg ccaacatctt cttctggagg
     3361 ccgtggttgg cttgtatgga gcagcagacg cgctacttcg agcggaggca tccggagctt
     3421 gcaggatcgc cgcggctccg ggcgtatatg ctccgcattg gtcttgacca actctatcag
     3481 agcttggttg acggcaattt cgatgatgca gcttgggcgc agggtcgatg cgacgcaatc
     3541 gtccgatccg gagccgggac tgtcgggcgt acacaaatcg cccgcagaag cgcggccgtc
     3601 tggaccgatg gctgtgtaga agtactcgcc gatagtggaa accgacgccc cagcactcgt
     3661 ccgagggcaa aggaatagca cgtactacga gatttcgatt ccaccgccgc cttctatgaa
     3721 aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat
     3781 ctcatgctgg agttcttcgc ccaccccaac ttgtttattg cagcttataa tggttacaaa
     3841 taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt
     3901 ggtttgtcca aactcatcaa tgtatcttat catgtctgta taccgtcgac ctctagctag
     3961 agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt
     4021 ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc
     4081 taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc
     4141 cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct
     4201 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca
     4261 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac
     4321 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt
     4381 ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg
     4441 cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc
     4501 tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc
     4561 gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc
     4621 aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac
     4681 tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt
     4741 aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct
     4801 aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc
     4861 ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt
     4921 ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg
     4981 atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc
     5041 atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa
     5101 tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag
     5161 gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg
     5221 tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga
     5281 gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg aggggccgag
     5341 cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa
     5401 gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc
     5461 atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca
     5521 aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg
     5581 atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat
     5641 aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc
     5701 aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg
     5761 gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg
     5821 gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt
     5881 gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca
     5941 ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata
     6001 ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac
     6061 atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa
     6121 gtgccacctg acgtc
//
