LOCUS       sequence-435429-        6484 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses HaloTag-LC3B, an autophagosome marker.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6484)
  AUTHORS   Tudorica DA, Basak B, Puerta Cordova AS, Khuu G, Rose K, Lazarou M, 
            Holzbaur ELF, Hurley JH
  TITLE     A RAB7A phosphoswitch coordinates Rubicon Homology protein 
            regulation of Parkin-dependent mitophagy.
  JOURNAL   J Cell Biol. 2024 Jul 1;223(7):e202309015. doi: 
            10.1083/jcb.202309015. Epub 2024 May 10.
  PUBMED    38728007
REFERENCE   2  (bases 1 to 6484)
  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: "J Cell 
            Biol."; date: "2024-07-1"; volume: "223(7):e202309015. doi"; pages: 
            " 10.1083/jcb.202309015. Epub 2024 May 10"
COMMENT     Sequence Label: pHTN-LC3B
FEATURES             Location/Qualifiers
     source          1..6484
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      848..857
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(866..885)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(867..885)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(927..946)
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     intron          929..1061
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     promoter        complement(1197..1400)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(1226..1246)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        1401..1780
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     misc_feature    1971..2254
                     /label=cer region
                     /note="ColE1-derived recombination site that helps to 
                     maintain plasmids as monomers"
     rep_origin      2370..2958
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2859..2878
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3116..3907,3908..3976))
                     /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     3739..3758
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3977..4081)
                     /gene="bla"
                     /label=AmpR promoter
     polyA_signal    4237..4285
                     /note="synthetic polyadenylation signal"
     CDS             complement(4349..5143)
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4460..4479)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     5070..5089
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(5194..5551)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5208..5343
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     enhancer        5360..5551
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     5508..5528
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     polyA_signal    5796..5917
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5833..5852)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5887..5906
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5949..5967
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(6013..6435)
                     /codon_start=1
                     /gene="Map1lc3b"
                     /product="rat LC3 protein"
                     /label=LC3
                     /note="Atg8 family member found on autophagosomes"
                     /translation="PSEKTFKQRRSFEQRVEDVRLIREQHPTKIPVIIERYKGEKQLPV
                     LDKTKFLVPDHVNMSELIKIIRRRLQLNANQAFFLLVNGHSMVSVSTPISEVYESERDE
                     DGFLYMVYASQETFGTALAVTYMSALKATATGREPCL"
     CDS             complement(join(6445..6484,1..851))
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /translation="MAEIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSS
                     YVWRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVI
                     HDWGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLII
                     DQNVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALV
                     EEYMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDL
                     IGSEIARWLSTLEGSG"
ORIGIN
        1 ctgccgatca ggtccgggtt gtcttcttgc agcagattca gacccgggcc gatgtccaca
       61 gccttgcagt taggcaggct tttggccagg cgagcggctt cggccggtgg gatcagaacg
      121 cctggggtgc cccagaacag cagcttcggg acaggggact ggtgcagcca gtccatgtat
      181 tcttcgacca gcgcgacgat gttcgctggc tcaccggcga ttggcagctc gtttgggaag
      241 cgccacagtg gctcgcggtc aacaggattc aggaacggct cgcggtaatg gtccatctcg
      301 acttcagtca gcgggcggac gacacccatc ggcagcgtac cctcgataaa aacgttctga
      361 tcgatgatca gcttgcggcc gacgtcggtg gtgcggaagg cctggaaggt ctcgcgggca
      421 aattctggcc attcgtccca ggtcgggata gggcggatga actccataaa tgcaatacct
      481 ttgacgcgct ctggattgcg cttggcccag tggaaaccca gagcggagcc ccagtcgtga
      541 atgaccagga cgacctcttc cagacccagg gcttcgatga aggcatccat gaagcggacg
      601 tggtcgtcga agaaataacc caggtctggt ttgtcggatt tgcccatacc gatcaggtct
      661 ggagcaatgc agcgatgggt cggtgcaaca tgcgggatga tgttgcgcca cacgtaggag
      721 gaggtcgggt taccgtgcag gaacagcaca ggggtgccat cgcgcggacc aacatcgacg
      781 tagtgcatgc gctcgcccag gacttccaca taatgggggt cgaatggaaa gccagtaccg
      841 atttctgcca tggtggcttt gctagcccta tagtgagtcg tattaatact ctagccttaa
      901 gagctgtaat tgaactggga gtggacacct gtggagagaa aggcaaagtg gatgtcagta
      961 agaccaatag gtgcctatca gaaacgcaag agtcttctct gtctcgacaa gcccagtttc
     1021 tattggtctc cttaaacctg tcttgtaacc ttgatactta cctgcccagt gcctcacgac
     1081 caacttctgc agcttaagtt cgagactgtt gtgtcagaag cactgactgc gttagcaatt
     1141 taactgtgat aaactaccgc aataaagctt ctagtgatct gacggttcac taaaccagct
     1201 ctgcttatat agacctccca ccgtacacgc ctaccgccca tttgcgtcaa cggggcgggg
     1261 ttattacgac attttggaaa gtcccgttga ttttggtgcc aaaacaaact cccattgacg
     1321 tcaatggggt ggagacttgg aaatccccgt gagtcaaacc gctatccacg cccattggtg
     1381 tactgccaaa accgcatcac catggtaata gcgatgacta atacgtagat gtactgccaa
     1441 gtaggaaagt cccgtaaggt catgtactgg gcataatgcc aggcgggcca tttaccgtca
     1501 ttgacgtcaa tagggggcgg acttggcata tgatacactt gatgtactgc caagtgggca
     1561 gtttaccgta aatactccac ccattgacgt caatggaaag tccctattgg cgttactatg
     1621 ggaacatacg tcattattga cgtcaatggg cgggggtcgt tgggcggtca gccaggcggg
     1681 ccatttaccg taagttatgt aacgcggaac tccatatatg ggctatgaac taatgacccc
     1741 gtaattgatt actattaata actagtcaat aatcaatgcc aacatggcgg tcatattgga
     1801 catgagccaa tataaatgta catattatga tatagataca acgtatgcaa tggccaatag
     1861 ccaatattga tttatgctat ataaccaatg aataatatgg ctaatggcca atattgaaga
     1921 tctcgatcct ctacgccgga cgcatcgtgg ccggagtact gcggtttatc tcgatggcta
     1981 cgagggcaga cagtaagtgg atttaccata atcccttaat tgtacgcacc gctaaaacgc
     2041 gttcagcgcg atcacggcag cagacaggta aaaatggcaa caaaccaccc gaaaaactgc
     2101 cgcaatcgcg cctgataaat tttaaccgta tgaataccta tgcaaccaga gggtacaggc
     2161 cacattaccc ccacttaatc cactgaagct gccatttttc atggtttcac catcccagcg
     2221 aagggccatc cagcgtgcgt tcctgtattt ccggactata aaaggatcta ggtgaagatc
     2281 ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca
     2341 gaccccgtag aaaagatcaa aggatcttct tgaaatcctt tttttctgcg cgtaatctgc
     2401 tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta
     2461 ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgtcctt
     2521 ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc
     2581 gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg
     2641 ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg
     2701 tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag
     2761 ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc
     2821 agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat
     2881 agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg
     2941 gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc
     3001 tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt
     3061 accgcctttg agtgagctga taccggttgg gcgtcgcttg gtcggtcatt tcgaattacc
     3121 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
     3181 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg
     3241 ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc
     3301 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta
     3361 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg
     3421 ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct
     3481 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta
     3541 gctccttcgg tcctccgata gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg
     3601 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga
     3661 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt
     3721 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttgaaa gtgctcatca
     3781 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt
     3841 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt
     3901 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga
     3961 aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat cagggttatt
     4021 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
     4081 gcacatttcc ccgaaaagtg ccacctgacg tctaagaaac cattacgtag aaagccagtc
     4141 cgcagaaacg gtgctgaccc cggatgaatg tcagctactg ggctatctgg acaagggaaa
     4201 acgcaagcgc aaagaggatc cttatcgcta tcgattcaca caaaaaacca acacacagat
     4261 gtaatgaaaa taaagatatt ttattgcggc catcgtgatg gcaggttggg cgtcgcttgg
     4321 tcggtcattt cgaaccccag agtcccgctc agaagaactc gtcaagaagg cgatagaagg
     4381 cgatgcgctg cgaatcggga gcggcgatac cgtaaagcac gaggaagcgg tcagcccatt
     4441 cgccgccaag ctcttcagca atatcacggg tagccaacgc tatgtcctga tagcggtccg
     4501 ccacacccag ccggccacag tcgatgaatc cagaaaagcg gccattttcc accatgatat
     4561 tcggcaagca ggcatcgcca tgggtcacga cgagatcctc gccgtcgggc atgcgcgcct
     4621 tgagcctggc gaacagttcg gctggcgcga gcccctgatg ctcttcgtcc agatcatcct
     4681 gatcgacaag accggcttcc atccgagtac gtgctcgctc gatgcgatgt ttcgcttggt
     4741 ggtcgaatgg gcaggtagcc ggatcaagcg tatgcagccg ccgcattgca tcagccatga
     4801 tggatacttt ctcggcagga gcaaggtgag atgacaggag atcctgcccc ggcacttcgc
     4861 ccaatagcag ccagtccctt cccgcttcag tgacaacgtc gagcacagct gcgcaaggaa
     4921 cgcccgtcgt ggccagccac gatagccgcg ctgcctcgtc ctgcagttca ttcagggcac
     4981 cggacaggtc ggtcttgaca aaaagaaccg ggcgcccctg cgctgacagc cggaacacgg
     5041 cggcatcaga gcagccgatt gtctgttgtg cccagtcata gccgaatagc ctctccaccc
     5101 aagcggccgg agaacctgcg tgcaatccat cttgttcaat catggtggct ctagccttaa
     5161 gttcgagact gttgtgtcag aagaatcaag ctttttgcaa aagcctaggc ctccaaaaaa
     5221 gcctcctcac tacttctgga atagctcaga ggccgaggcg gcctcggcct ctgcataaat
     5281 aaaaaaaatt agtcagccat ggggcggaga atgggcggaa ctgggcggag ttaggggcgg
     5341 gatgggcgga gttaggggcg ggactatggt tgctgactaa ttgagatgca tgctttgcat
     5401 acttctgcct gctggggagc ctggggactt tccacacctg gttgctgact aattgagatg
     5461 catgctttgc atacttctgc ctgctgggga gcctggggac tttccacacc ctaactgaca
     5521 cacattccac agctggttct ttccgcctca gaaggtacct aaccaagttc ctctttcaga
     5581 ggttatttca ggccatggtg ctgcgcagat ccgcgtatgc ggtgtgaaat accgcacaga
     5641 tgcgtaagga gaaaataccg catcaggaaa ttgtaagcgt taatattttg ttaaaattcg
     5701 attttaccac atttgtagag gttttacttg cttaaaaaaa cctcccacat ctccccctga
     5761 acctgaaaca taaaatgaat gcaattgttg ttgttaactt gtttattgca gcttataatg
     5821 gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt
     5881 ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctgctcg aagcggccaa
     5941 ggggttatgc tagttattgc tcagcggtgg cagcagccaa ctcagcaagc ggccgctcga
     6001 gtcgacttat cacaagcatg gctctcttcc tgttgctgtt gccttcagag ctgacatgta
     6061 tgtaacagcc agtgctgtcc cgaacgtctc ctgggaggca tagaccatgt acaggaagcc
     6121 gtcttcatct ctctcgctct cgtacacttc agagatgggt gtggacacac tcaccatgct
     6181 gtgcccattc accaggagga agaaggcttg gttagcattg agctgcaggc gccttctaat
     6241 tatcttgatg agttcgctca tattcacgtg atcaggtaca aggaacttgg tcttgtccag
     6301 gacgggcagc tgcttctcac ccttgtatcg ctctataatc actgggatct tggtggggtg
     6361 ctgctcccgg atgagccgga catcttccac tctttgttcg aagctccggc gctgtttgaa
     6421 ggtcttctcg gacggtgatc cgctacctga tccctcgagc gtcgacagcc agcgcgcgat
     6481 ctcg
//
