LOCUS       sequence-350166-        6850 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of lysosome-associated membrane protein 2b 
            fused to a C-terminal HA tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6850)
  AUTHORS   Hung ME, Leonard JN
  TITLE     A platform for actively loading cargo RNA to elucidate limiting 
            steps in EV-mediated delivery.
  JOURNAL   J Extracell Vesicles. 2016 May 13;5:31027. doi: 
            10.3402/jev.v5.31027. eCollection 2016.
  PUBMED    27189348
REFERENCE   2  (bases 1 to 6850)
  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 Extracell
            Vesicles."; date: "2016-05-13"; volume: "5:31027. doi"; pages: " 
            10.3402/jev.v5.31027. eCollection 2016"
COMMENT     Sequence Label: pMEH31 Lamp2b HA
FEATURES             Location/Qualifiers
     source          1..6850
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(70..89)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        261..640
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        641..844
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     795..815
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     889..908
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        889..907
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      930..939
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2175..2201
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     primer_bind     complement(2298..2315)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2304..2528
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2574..3002
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2661..2680)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2871..2892
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3011..3031)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3016..3345
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3196..3331
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3258..3277
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3394..4419
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     CDS             3394..4419
                     /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    4549..4670
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4586..4605)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4640..4659
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4719..4735)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4719..4735)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4732..4754)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4743..4759
                     /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(4767..4773,4774..4791,4792..4797))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4812..4833
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4950..4967)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5121..5709)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5201..5220)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5880..6671,6672..6740))
                     /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     6503..6522
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6741..6845)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ttccccgaaa agtgccacct gacgtcgacg gatcgggaga tctcccgatc ccctatggtg
       61 cactctcagt acaatctgct ctgatgccgc atagttaagc cagtatctgc tccctgcttg
      121 tgtgttggag gtcgctgagt agtgcgcgag caaaatttaa gctacaacaa ggcaaggctt
      181 gaccgacaat tgcatgaaga atctgcttag ggttaggcgt tttgcgctgc ttcgcgatgt
      241 acgggccaga tatacgcgtt gacattgatt attgactagt tattaatagt aatcaattac
      301 ggggtcatta gttcatagcc catatatgga gttccgcgtt acataactta cggtaaatgg
      361 cccgcctggc tgaccgccca acgacccccg cccattgacg tcaataatga cgtatgttcc
      421 catagtaacg ccaataggga ctttccattg acgtcaatgg gtggagtatt tacggtaaac
      481 tgcccacttg gcagtacatc aagtgtatca tatgccaagt acgcccccta ttgacgtcaa
      541 tgacggtaaa tggcccgcct ggcattatgc ccagtacatg accttatggg actttcctac
      601 ttggcagtac atctacgtat tagtcatcgc tattaccatg gtgatgcggt tttggcagta
      661 catcaatggg cgtggatagc ggtttgactc acggggattt ccaagtctcc accccattga
      721 cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa
      781 ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct atataagcag
      841 agctctctgg ctaactagag aacccactgc ttactggctt atcgaaatta atacgactca
      901 ctatagggag acccaagctg gctagtgccg ccgccatggt gtgcttccgc ctcttcccgg
      961 ttccgggctc agggctcgtt ctggtctgcc tagtcctggg agctgtgcgg tcttatgcat
     1021 tggaacttaa tttgacagat tcagaaaatg ccacttgcct ttatgcaaaa tggcagatga
     1081 atttcacagt tcgctatgaa actacaaata aaacttataa aactgtaacc atttcagacc
     1141 atggcactgt gacatataat ggaagcattt gtggggatga tcagaatggt cccaaaatag
     1201 cagtgcagtt cggacctggc ttttcctgga ttgcgaattt taccaaggca gcatctactt
     1261 attcaattga cagcgtctca ttttcctaca acactggtga taacacaaca tttcctgatg
     1321 ctgaagataa aggaattctt actgttgatg aacttttggc catcagaatt ccattgaatg
     1381 acctttttag atgcaatagt ttatcaactt tggaaaagaa tgatgttgtc caacactact
     1441 gggatgttct tgtacaagct tttgtccaaa atggcacagt gagcacaaat gagttcctgt
     1501 gtgataaaga caaaacttca acagtggcac ccaccataca caccactgtg ccatctccta
     1561 ctacaacacc tactccaaag gaaaaaccag aagctggaac ctattcagtt aataatggca
     1621 atgatacttg cctgctggct accatggggc tgcagctgaa catcactcag gataaggttg
     1681 cttcagttat taacatcaac cccaatacaa ctcactccac aggcagctgc cgttctcaca
     1741 ctgctctact tagactcaat agcagcacta ttaagtatct agactttgtc tttgctgtga
     1801 aaaatgaaaa ccgattttat ctgaaggaag tgaacatcag catgtatttg gttaatggct
     1861 ccgttttcag cattgcaaat aacaatctca gctactggga tgcccccctg ggaagttctt
     1921 atatgtgcaa caaagagcag actgtttcag tgtctggagc atttcagata aatacctttg
     1981 atctaagggt tcagcctttc aatgtgacac aaggaaagta ttctacagcc caagagtgtt
     2041 cgctggatga tgacaccatt ctaatcccaa ttatagttgg tgctggtctt tcaggcttga
     2101 ttatcgttat agtgattgct tacgtaattg gcagaagaaa aagttatgct ggatatcaga
     2161 ctctgggaag cggataccca tacgatgtgc cagattacgc ttaaggatcc actagtccag
     2221 tgtggtggaa ttctgcagat atccagcaca gtggcggccg ctcgagtcta gagggcccgt
     2281 ttaaacccgc tgatcagcct cgactgtgcc ttctagttgc cagccatctg ttgtttgccc
     2341 ctcccccgtg ccttccttga ccctggaagg tgccactccc actgtccttt cctaataaaa
     2401 tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct attctggggg gtggggtggg
     2461 gcaggacagc aagggggagg attgggaaga caatagcagg catgctgggg atgcggtggg
     2521 ctctatggct tctgaggcgg aaagaaccag ctggggctct agggggtatc cccacgcgcc
     2581 ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact
     2641 tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc
     2701 cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt
     2761 acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc
     2821 ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt
     2881 gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat
     2941 tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa
     3001 ttaattctgt ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc
     3061 agaagtatgc aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc
     3121 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg
     3181 cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat
     3241 ggctgactaa ttttttttat ttatgcagag gccgaggccg cctctgcctc tgagctattc
     3301 cagaagtagt gaggaggctt ttttggaggc ctaggctttt gcaaaaagct cccgggagct
     3361 tgtatatcca ttttcggatc tgatcagcac gtgatgaaaa agcctgaact caccgcgacg
     3421 tctgtcgaga agtttctgat cgaaaagttc gacagcgtct ccgacctgat gcagctctcg
     3481 gagggcgaag aatctcgtgc tttcagcttc gatgtaggag ggcgtggata tgtcctgcgg
     3541 gtaaatagct gcgccgatgg tttctacaaa gatcgttatg tttatcggca ctttgcatcg
     3601 gccgcgctcc cgattccgga agtgcttgac attggggaat tcagcgagag cctgacctat
     3661 tgcatctccc gccgtgcaca gggtgtcacg ttgcaagacc tgcctgaaac cgaactgccc
     3721 gctgttctgc agccggtcgc ggaggccatg gatgcgatcg ctgcggccga tcttagccag
     3781 acgagcgggt tcggcccatt cggaccgcaa ggaatcggtc aatacactac atggcgtgat
     3841 ttcatatgcg cgattgctga tccccatgtg tatcactggc aaactgtgat ggacgacacc
     3901 gtcagtgcgt ccgtcgcgca ggctctcgat gagctgatgc tttgggccga ggactgcccc
     3961 gaagtccggc acctcgtgca cgcggatttc ggctccaaca atgtcctgac ggacaatggc
     4021 cgcataacag cggtcattga ctggagcgag gcgatgttcg gggattccca atacgaggtc
     4081 gccaacatct tcttctggag gccgtggttg gcttgtatgg agcagcagac gcgctacttc
     4141 gagcggaggc atccggagct tgcaggatcg ccgcggctcc gggcgtatat gctccgcatt
     4201 ggtcttgacc aactctatca gagcttggtt gacggcaatt tcgatgatgc agcttgggcg
     4261 cagggtcgat gcgacgcaat cgtccgatcc ggagccggga ctgtcgggcg tacacaaatc
     4321 gcccgcagaa gcgcggccgt ctggaccgat ggctgtgtag aagtactcgc cgatagtgga
     4381 aaccgacgcc ccagcactcg tccgagggca aaggaatagc acgtgctacg agatttcgat
     4441 tccaccgccg ccttctatga aaggttgggc ttcggaatcg ttttccggga cgccggctgg
     4501 atgatcctcc agcgcgggga tctcatgctg gagttcttcg cccaccccaa cttgtttatt
     4561 gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt
     4621 ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgt
     4681 ataccgtcga cctctagcta gagcttggcg taatcatggt catagctgtt tcctgtgtga
     4741 aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc
     4801 tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc
     4861 cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc
     4921 ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt
     4981 cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca
     5041 ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa
     5101 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat
     5161 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc
     5221 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc
     5281 gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt
     5341 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac
     5401 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg
     5461 ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca
     5521 gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc
     5581 gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa
     5641 accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa
     5701 ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac
     5761 tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta
     5821 aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt
     5881 taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata
     5941 gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc
     6001 agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac
     6061 cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag
     6121 tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac
     6181 gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc
     6241 agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg
     6301 gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc
     6361 atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct
     6421 gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc
     6481 tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc
     6541 atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc
     6601 agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc
     6661 gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca
     6721 cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt
     6781 tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt
     6841 ccgcgcacat
//
