LOCUS       sequence-244374-        7059 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Gateway destination vector for mammalian expression without tag, or 
            for adding custom tags by conventional cloning. Distinct multiple 
            cloning sites 5' and 3' of gateway cassette..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7059)
  AUTHORS   Agrotis A, Pengo N, Burden JJ, Ketteler R
  TITLE     Redundancy of human ATG4 protease isoforms in autophagy and 
            LC3/GABARAP processing revealed in cells.
  JOURNAL   Autophagy. 2019 Jan 21. doi: 10.1080/15548627.2019.1569925.
  PUBMED    30661429
REFERENCE   2  (bases 1 to 7059)
  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. 
            2019 Jan 21. doi: 10.1080/15548627.2019.1569925."
COMMENT     Sequence Label: pDEST-CMV-polysite
FEATURES             Location/Qualifiers
     source          1..7059
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(46..65)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        237..616
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        617..820
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     771..791
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    890..1014
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(1039..1044,1045..1062,1063..1069)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             1123..1803
                     /codon_start=1
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQAGRNLE
                     DPAY"
     primer_bind     complement(1189..1208)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             2123..2428
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VPRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     2307..2326
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(2469..2593)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(2652..2669)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2658..2882
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2928..3356
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3015..3034)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3225..3246
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3365..3385)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3370..3699
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3550..3685
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3612..3631
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3766..4560
                     /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(3820..3839)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4430..4449
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4734..4855
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4771..4790)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4825..4844
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4904..4920)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4904..4920)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4917..4939)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4928..4944
                     /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(4952..4958,4959..4976,4977..4982))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4997..5018
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5135..5152)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5306..5894)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5386..5405)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6065..6856,6857..6925))
                     /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     6688..6707
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6926..7030)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tcgacggatc gggagatctc ccgatcccct atggtgcact ctcagtacaa tctgctctga
       61 tgccgcatag ttaagccagt atctgctccc tgcttgtgtg ttggaggtcg ctgagtagtg
      121 cgcgagcaaa atttaagcta caacaaggca aggcttgacc gacaattgca tgaagaatct
      181 gcttagggtt aggcgttttg cgctgcttcg cgatgtacgg gccagatata cgcgttgaca
      241 ttgattattg actagttatt aatagtaatc aattacgggg tcattagttc atagcccata
      301 tatggagttc cgcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga
      361 cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt
      421 ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt
      481 gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca
      541 ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt
      601 catcgctatt accatggtga tgcggttttg gcagtacatc aatgggcgtg gatagcggtt
      661 tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt tgttttggca
      721 ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga cgcaaatggg
      781 cggtaggcgt gtacggtggg aggtctatat aagcagagct cgtttagtga accgtcagaa
      841 ttgatctggt accggcgtta acgacgctag cggagttaag cttgatcaaa caagtttgta
      901 caaaaaagct gaacgagaaa cgtaaaatga tataaatatc aatatattaa attagatttt
      961 gcataaaaaa cagactacat aatactgtaa aacacaacat atccagtcac tatggcggcc
     1021 gcattaggca ccccaggctt tacactttat gcttccggct cgtataatgt gtggattttg
     1081 agttaggatc cgtcgagatt ttcaggagct aaggaagcta aaatggagaa aaaaatcact
     1141 ggatatacca ccgttgatat atcccaatgg catcgtaaag aacattttga ggcatttcag
     1201 tcagttgctc aatgtaccta taaccagacc gttcagctgg atattacggc ctttttaaag
     1261 accgtaaaga aaaataagca caagttttat ccggccttta ttcacattct tgcccgcctg
     1321 atgaatgctc atccggaatt ccgtatggca atgaaagacg gtgagctggt gatatgggat
     1381 agtgttcacc cttgttacac cgttttccat gagcaaactg aaacgttttc atcgctctgg
     1441 agtgaatacc acgacgattt ccggcagttt ctacacatat attcgcaaga tgtggcgtgt
     1501 tacggtgaaa acctggccta tttccctaaa gggtttattg agaatatgtt tttcgtctca
     1561 gccaatccct gggtgagttt caccagtttt gatttaaacg tggccaatat ggacaacttc
     1621 ttcgcccccg ttttcaccat gggcaaatat tatacgcaag gcgacaaggt gctgatgccg
     1681 ctggcgattc aggttcatca tgccgtttgt gatggcttcc atgtcggcag aatgcttaat
     1741 gaattacaac agtactgcga tgagtggcag gcggggcgta atctagagga tccggcttac
     1801 taaaagccag ataacagtat gcgtatttgc gcgctgattt ttgcggtata agaatatata
     1861 ctgatatgta tacccgaagt atgtcaaaaa gaggtatgct atgaagcagc gtattacagt
     1921 gacagttgac agcgacagct atcagttgct caaggcatat atgatgtcaa tatctccggt
     1981 ctggtaagca caaccatgca gaatgaagcc cgtcgtctgc gtgccgaacg ctggaaagcg
     2041 gaaaatcagg aagggatggc tgaggtcgcc cggtttattg aaatgaacgg ctcttttgct
     2101 gacgagaaca ggggctggtg aaatgcagtt taaggtttac acctataaaa gagagagccg
     2161 ttatcgtctg tttgtggatg tacagagtga tattattgac acgcccgggc gacggatggt
     2221 gatccccctg gccagtgcac gtctgctgtc agataaagtc ccccgtgaac tttacccggt
     2281 ggtgcatatc ggggatgaaa gctggcgcat gatgaccacc gatatggcca gtgtgccggt
     2341 ctccgttatc ggggaagaag tggctgatct cagccaccgc gaaaatgaca tcaaaaacgc
     2401 cattaacctg atgttctggg gaatataaat gtcaggctcc cttatacaca gccagtctgc
     2461 aggtcgacca tagtgactgg atatgttgtg ttttacagta ttatgtagtc tgttttttat
     2521 gcaaaatcta atttaatata ttgatattta tatcatttta cgtttctcgt tcagctttct
     2581 tgtacaaagt ggttcgatct agactcgagc ggtgtttaaa ccggcttaat taagataaac
     2641 ccgctgatca gcctcgactg tgccttctag ttgccagcca tctgttgttt gcccctcccc
     2701 cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc ctttcctaat aaaatgagga
     2761 aattgcatcg cattgtctga gtaggtgtca ttctattctg gggggtgggg tggggcagga
     2821 cagcaagggg gaggattggg aagacaatag caggcatgct ggggatgcgg tgggctctat
     2881 ggcttctgag gcggaaagaa ccagctgggg ctctaggggg tatccccacg cgccctgtag
     2941 cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag
     3001 cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt
     3061 tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca
     3121 cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat cgccctgata
     3181 gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca
     3241 aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag ggattttgcc
     3301 gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattaatt
     3361 ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc aggcagaagt
     3421 atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc aggctcccca
     3481 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt cccgccccta
     3541 actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc ccatggctga
     3601 ctaatttttt ttatttatgc agaggccgag gccgcctctg cctctgagct attccagaag
     3661 tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa agctcccggg agcttgtata
     3721 tccattttcg gatctgatca agagacagga tgaggatcgt ttcgcatgat tgaacaagat
     3781 ggattgcacg caggttctcc ggccgcttgg gtggagaggc tattcggcta tgactgggca
     3841 caacagacaa tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg
     3901 gttctttttg tcaagaccga cctgtccggt gccctgaatg aactgcagga cgaggcagcg
     3961 cggctatcgt ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact
     4021 gaagcgggaa gggactggct gctattgggc gaagtgccgg ggcaggatct cctgtcatct
     4081 caccttgctc ctgccgagaa agtatccatc atggctgatg caatgcggcg gctgcatacg
     4141 cttgatccgg ctacctgccc attcgaccac caagcgaaac atcgcatcga gcgagcacgt
     4201 actcggatgg aagccggtct tgtcgatcag gatgatctgg acgaagagca tcaggggctc
     4261 gcgccagccg aactgttcgc caggctcaag gcgcgcatgc ccgacggcga ggatctcgtc
     4321 gtgacccatg gcgatgcctg cttgccgaat atcatggtgg aaaatggccg cttttctgga
     4381 ttcatcgact gtggccggct gggtgtggcg gaccgctatc aggacatagc gttggctacc
     4441 cgtgatattg ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt gctttacggt
     4501 atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga
     4561 gcgggactct ggggttcgaa atgaccgacc aagcgacgcc caacctgcca tcacgagatt
     4621 tcgattccac cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg
     4681 gctggatgat cctccagcgc ggggatctca tgctggagtt cttcgcccac cccaacttgt
     4741 ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag
     4801 catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta tcttatcatg
     4861 tctgtatacc gtcgacctct agctagagct tggcgtaatc atggtcatag ctgtttcctg
     4921 tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta
     4981 aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg
     5041 ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga
     5101 gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg
     5161 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
     5221 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
     5281 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
     5341 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
     5401 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     5461 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     5521 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     5581 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     5641 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     5701 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta
     5761 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     5821 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     5881 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
     5941 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
     6001 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg
     6061 acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat
     6121 ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg
     6181 gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa
     6241 taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca
     6301 tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc
     6361 gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt
     6421 cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa
     6481 aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat
     6541 cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct
     6601 tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga
     6661 gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag
     6721 tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga
     6781 gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca
     6841 ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg
     6901 cgacacggaa atgttgaata ctcatactct tcctttttca atattattga agcatttatc
     6961 agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag
     7021 gggttccgcg cacatttccc cgaaaagtgc cacctgacg
//
