LOCUS       sequence-127970-        6988 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes (N to C): GASTM mutated glycosylation motif, 3 residue 
            spacer, RVG peptide,10 residue spacer, Lamp2b (exosomal membrane 
            protein), 3 residue spacer, HA tag..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6988)
  AUTHORS   Hung ME, Leonard JN
  TITLE     Stabilization of exosome-targeting peptides via engineered 
            glycosylation.
  JOURNAL   J Biol Chem. 2015 Mar 27;290(13):8166-72. doi: 
            10.1074/jbc.M114.621383. Epub 2015 Feb 5.
  PUBMED    25657008
REFERENCE   2  (bases 1 to 6988)
  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"; doi: 
            "10.1074/jbc.M114.621383"; journalName: "J Biol Chem"; date: 
            "2015-03-27- 27"; volume: "290"; issue: "13"; pages: "8166-72"
COMMENT     Sequence Label: pcDNA GASTM-3-RVG-10-Lamp2b-HA
FEATURES             Location/Qualifiers
     source          1..6988
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        110..489
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        490..693
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     644..664
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     738..757
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        738..756
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      779..788
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2165..2191
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     primer_bind     complement(2288..2305)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2294..2518
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2564..2992
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2651..2670)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2861..2882
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3001..3021)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3006..3335
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3186..3321
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3248..3267
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3384..4409
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     CDS             3384..4409
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     polyA_signal    4539..4660
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4576..4595)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4630..4649
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4709..4725)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4709..4725)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4722..4744)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4733..4749
                     /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(4757..4763,4764..4781,4782..4787))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4802..4823
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4940..4957)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5111..5696)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5191..5210)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5867..6658,6659..6727))
                     /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     6490..6509
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6728..6832)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6907..6926)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 aaaatttaag ctacaacaag gcaaggcttg accgacaatt gcatgaagaa tctgcttagg
       61 gttaggcgtt ttgcgctgct tcgcgatgta cgggccagat atacgcgttg acattgatta
      121 ttgactagtt attaatagta atcaattacg gggtcattag ttcatagccc atatatggag
      181 ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc
      241 ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac tttccattga
      301 cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca agtgtatcat
      361 atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc
      421 cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt agtcatcgct
      481 attaccatgg tgatgcggtt ttggcagtac atcaatgggc gtggatagcg gtttgactca
      541 cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg gcaccaaaat
      601 caacgggact ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat gggcggtagg
      661 cgtgtacggt gggaggtcta tataagcaga gctctctggc taactagaga acccactgct
      721 tactggctta tcgaaattaa tacgactcac tatagggaga cccaagctgg ctagtgccgc
      781 cgccatggtg tgcttccgcc tcttcccggt tccgggctca gggctcgttc tggtctgcct
      841 agtcctggga gctgtgcggt cttatgcagg tgctagcact atgggcagtg gatacaccat
      901 ttggatgccc gagaatccga gaccagggac accttgtgac atttttacca atagcagagg
      961 gaagagagca tccaacgggg gcagtggatc tggatccggt ggctcgagtt tggaacttaa
     1021 tttgacagat tcagaaaatg ccacttgcct ttatgcaaaa tggcagatga atttcacagt
     1081 tcgctatgaa actacaaata aaacttataa aactgtaacc atttcagacc atggcactgt
     1141 gacatataat ggaagcattt gtggggatga tcagaatggt cccaaaatag cagtgcagtt
     1201 cggacctggc ttttcctgga ttgcgaattt taccaaggca gcatctactt attcaattga
     1261 cagcgtctca ttttcctaca acactggtga taacacaaca tttcctgatg ctgaagataa
     1321 aggaattctt actgttgatg aacttttggc catcagaatt ccattgaatg acctttttag
     1381 atgcaatagt ttatcaactt tggaaaagaa tgatgttgtc caacactact gggatgttct
     1441 tgtacaagct tttgtccaaa atggcacagt gagcacaaat gagttcctgt gtgataaaga
     1501 caaaacttca acagtggcac ccaccataca caccactgtg ccatctccta ctacaacacc
     1561 tactccaaag gaaaaaccag aagctggaac ctattcagtt aataatggca atgatacttg
     1621 cctgctggct accatggggc tgcagctgaa catcactcag gataaggttg cttcagttat
     1681 taacatcaac cccaatacaa ctcactccac aggcagctgc cgttctcaca ctgctctact
     1741 tagactcaat agcagcacta ttaagtatct agactttgtc tttgctgtga aaaatgaaaa
     1801 ccgattttat ctgaaggaag tgaacatcag catgtatttg gttaatggct ccgttttcag
     1861 cattgcaaat aacaatctca gctactggga tgcccccctg ggaagttctt atatgtgcaa
     1921 caaagagcag actgtttcag tgtctggagc atttcagata aatacctttg atctaagggt
     1981 tcagcctttc aatgtgacac aaggaaagta ttctacagcc caagagtgtt cgctggatga
     2041 tgacaccatt ctaatcccaa ttatagttgg tgctggtctt tcaggcttga ttatcgttat
     2101 agtgattgct tacgtaattg gcagaagaaa aagttatgct ggatatcaga ctctgggaag
     2161 cggataccca tacgatgtgc cagattacgc ttaaggatcc actagtccag tgtggtggaa
     2221 ttctgcagat atccagcaca gtggcggccg ctcgagtcta gagggcccgt ttaaacccgc
     2281 tgatcagcct cgactgtgcc ttctagttgc cagccatctg ttgtttgccc ctcccccgtg
     2341 ccttccttga ccctggaagg tgccactccc actgtccttt cctaataaaa tgaggaaatt
     2401 gcatcgcatt gtctgagtag gtgtcattct attctggggg gtggggtggg gcaggacagc
     2461 aagggggagg attgggaaga caatagcagg catgctgggg atgcggtggg ctctatggct
     2521 tctgaggcgg aaagaaccag ctggggctct agggggtatc cccacgcgcc ctgtagcggc
     2581 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
     2641 ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc
     2701 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc
     2761 gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg
     2821 gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact
     2881 ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt
     2941 tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttaattctgt
     3001 ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
     3061 aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag
     3121 gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc
     3181 cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa
     3241 ttttttttat ttatgcagag gccgaggccg cctctgcctc tgagctattc cagaagtagt
     3301 gaggaggctt ttttggaggc ctaggctttt gcaaaaagct cccgggagct tgtatatcca
     3361 ttttcggatc tgatcagcac gtgatgaaaa agcctgaact caccgcgacg tctgtcgaga
     3421 agtttctgat cgaaaagttc gacagcgtct ccgacctgat gcagctctcg gagggcgaag
     3481 aatctcgtgc tttcagcttc gatgtaggag ggcgtggata tgtcctgcgg gtaaatagct
     3541 gcgccgatgg tttctacaaa gatcgttatg tttatcggca ctttgcatcg gccgcgctcc
     3601 cgattccgga agtgcttgac attggggaat tcagcgagag cctgacctat tgcatctccc
     3661 gccgtgcaca gggtgtcacg ttgcaagacc tgcctgaaac cgaactgccc gctgttctgc
     3721 agccggtcgc ggaggccatg gatgcgatcg ctgcggccga tcttagccag acgagcgggt
     3781 tcggcccatt cggaccgcaa ggaatcggtc aatacactac atggcgtgat ttcatatgcg
     3841 cgattgctga tccccatgtg tatcactggc aaactgtgat ggacgacacc gtcagtgcgt
     3901 ccgtcgcgca ggctctcgat gagctgatgc tttgggccga ggactgcccc gaagtccggc
     3961 acctcgtgca cgcggatttc ggctccaaca atgtcctgac ggacaatggc cgcataacag
     4021 cggtcattga ctggagcgag gcgatgttcg gggattccca atacgaggtc gccaacatct
     4081 tcttctggag gccgtggttg gcttgtatgg agcagcagac gcgctacttc gagcggaggc
     4141 atccggagct tgcaggatcg ccgcggctcc gggcgtatat gctccgcatt ggtcttgacc
     4201 aactctatca gagcttggtt gacggcaatt tcgatgatgc agcttgggcg cagggtcgat
     4261 gcgacgcaat cgtccgatcc ggagccggga ctgtcgggcg tacacaaatc gcccgcagaa
     4321 gcgcggccgt ctggaccgat ggctgtgtag aagtactcgc cgatagtgga aaccgacgcc
     4381 ccagcactcg tccgagggca aaggaatagc acgtgctacg agatttcgat tccaccgccg
     4441 ccttctatga aaggttgggc ttcggaatcg ttttccggga cgccggctgg atgatcctcc
     4501 agcgcgggga tctcatgctg gagttcttcg cccaccccaa cttgtttatt gcagcttata
     4561 atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc
     4621 attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgt ataccgtcga
     4681 cctctagcta gagcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc
     4741 cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct
     4801 aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
     4861 acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta
     4921 ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     4981 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     5041 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     5101 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     5161 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     5221 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     5281 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     5341 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     5401 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     5461 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     5521 agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga
     5581 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     5641 gtagcggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
     5701 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
     5761 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa
     5821 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa
     5881 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc
     5941 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga
     6001 taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa
     6061 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt
     6121 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg
     6181 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc
     6241 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg
     6301 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag
     6361 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt
     6421 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt
     6481 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac
     6541 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac
     6601 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag
     6661 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa
     6721 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga
     6781 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc
     6841 cccgaaaagt gccacctgac gtcgacggat cgggagatct cccgatcccc tatggtgcac
     6901 tctcagtaca atctgctctg atgccgcata gttaagccag tatctgctcc ctgcttgtgt
     6961 gttggaggtc gctgagtagt gcgcgagc
//
