LOCUS       sequence-261777-        8074 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  The puromycin selection marker was replaced by blasticidin in 
            SHC003. This construct could be used together with other plasmid 
            contains puromycin selection marker..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8074)
  AUTHORS   Wang S, Crisman L, Miller J, Datta I, Gulbranson DR, Tian Y, Yin Q, 
            Yu H, Shen J
  TITLE     Inducible Exoc7/Exo70 knockout reveals a critical role of the 
            exocyst in insulin-regulated GLUT4 exocytosis.
  JOURNAL   J Biol Chem. 2019 Dec 27;294(52):19988-19996. doi: 
            10.1074/jbc.RA119.010821. Epub 2019 Nov 18.
  PUBMED    31740584
REFERENCE   2  (bases 1 to 8074)
  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.RA119.010821"; journalName: "J Biol Chem"; date: 
            "2019-12-27- 27"; volume: "294"; issue: "52"; pages: "19988-19996"
COMMENT     Sequence Label: SHC003BSD
FEATURES             Location/Qualifiers
     source          1..8074
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..227
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             228..408
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    455..580
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1073..1306
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1491..1535
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     promoter        1833..2081
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     1833..1853
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2004..2023
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     enhancer        2144..2447
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2448..2651
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2602..2622
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      2690..2699
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2696..3394
                     /codon_start=1
                     /product="improved green fluorescent protein from 
                     Pontellina plumata (Evdokimov et al., 2006)"
                     /label=TurboGFP
                     /note="mammalian codon-optimized"
                     /translation="MESDESGLPAMEIECRITGTLNGVEFELVGGGEGTPEQGRMTNKM
                     KSTKGALTFSPYLLSHVMGYGFYHFGTYPSGYENPFLHAINNGGYTNTRIEKYEDGGVL
                     HVSFSYRYEAGRVIGDFKVMGTGFPEDSVIFTDKIIRSNATVEHLHPMGDNDLDGSFTR
                     TFSLRDGGYYSSVVDSHMHFKSAIHPSILQNGGPMFAFRRVEEDHSNTELGIVEYQHAF
                     KTPDADAGEE"
     misc_feature    3435..3552
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        3601..4111
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     4030..4048
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     LTR             4593..4826
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    4898..5019
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4935..4954)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4989..5008
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      5059..5194
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(5214..5233)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(5215..5233)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5243..5260)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5243..5259)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5252..5274)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5401..5856
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5488..5507)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5698..5719
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5882..5986
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5987..6055,6056..6847)
                     /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     complement(6205..6224)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7018..7606
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7507..7526
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7760..7777
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7894..7915
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7930..7935,7936..7953,7954..7960)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7968..7984
                     /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)."
     primer_bind     7973..7995
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7992..8008
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7992..8008
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     8027..8047
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        8029..8047
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
ORIGIN
        1 tgtagtctta tgcaatactc ttgtagtctt gcaacatggt aacgatgagt tagcaacatg
       61 ccttacaagg agagaaaaag caccgtgcat gccgattggt ggaagtaagg tggtacgatc
      121 gtgccttatt aggaaggcaa cagacgggtc tgacatggat tggacgaacc actgaattgc
      181 cgcattgcag agatattgta tttaagtgcc tagctcgata cataaacggg tctctctggt
      241 tagaccagat ctgagcctgg gagctctctg gctaactagg gaacccactg cttaagcctc
      301 aataaagctt gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt gactctggta
      361 actagagatc cctcagaccc ttttagtcag tgtggaaaat ctctagcagt ggcgcccgaa
      421 cagggacttg aaagcgaaag ggaaaccaga ggagctctct cgacgcagga ctcggcttgc
      481 tgaagcgcgc acggcaagag gcgaggggcg gcgactggtg agtacgccaa aaattttgac
      541 tagcggaggc tagaaggaga gagatgggtg cgagagcgtc agtattaagc gggggagaat
      601 tagatcgcga tgggaaaaaa ttcggttaag gccaggggga aagaaaaaat ataaattaaa
      661 acatatagta tgggcaagca gggagctaga acgattcgca gttaatcctg gcctgttaga
      721 aacatcagaa ggctgtagac aaatactggg acagctacaa ccatcccttc agacaggatc
      781 agaagaactt agatcattat ataatacagt agcaaccctc tattgtgtgc atcaaaggat
      841 agagataaaa gacaccaagg aagctttaga caagatagag gaagagcaaa acaaaagtaa
      901 gaccaccgca cagcaagcgg ccgctgatct tcagacctgg aggaggagat atgagggaca
      961 attggagaag tgaattatat aaatataaag tagtaaaaat tgaaccatta ggagtagcac
     1021 ccaccaaggc aaagagaaga gtggtgcaga gagaaaaaag agcagtggga ataggagctt
     1081 tgttccttgg gttcttggga gcagcaggaa gcactatggg cgcagcgtca atgacgctga
     1141 cggtacaggc cagacaatta ttgtctggta tagtgcagca gcagaacaat ttgctgaggg
     1201 ctattgaggc gcaacagcat ctgttgcaac tcacagtctg gggcatcaag cagctccagg
     1261 caagaatcct ggctgtggaa agatacctaa aggatcaaca gctcctgggg atttggggtt
     1321 gctctggaaa actcatttgc accactgctg tgccttggaa tgctagttgg agtaataaat
     1381 ctctggaaca gatttggaat cacacgacct ggatggagtg ggacagagaa attaacaatt
     1441 acacaagctt aatacactcc ttaattgaag aatcgcaaaa ccagcaagaa aagaatgaac
     1501 aagaattatt ggaattagat aaatgggcaa gtttgtggaa ttggtttaac ataacaaatt
     1561 ggctgtggta tataaaatta ttcataatga tagtaggagg cttggtaggt ttaagaatag
     1621 tttttgctgt actttctata gtgaatagag ttaggcaggg atattcacca ttatcgtttc
     1681 agacccacct cccaaccccg aggggacccg acaggcccga aggaatagaa gaagaaggtg
     1741 gagagagaga cagagacaga tccattcgat tagtgaacgg atctcgacgg tatcgatcac
     1801 gagactagcc tcgagcggcc gcccccttca ccgagggcct atttcccatg attccttcat
     1861 atttgcatat acgatacaag gctgttagag agataattgg aattaatttg actgtaaaca
     1921 caaagatatt agtacaaaat acgtgacgta gaaagtaata atttcttggg tagtttgcag
     1981 ttttaaaatt atgttttaaa atggactatc atatgcttac cgtaacttga aagtatttcg
     2041 atttcttggc tttatatatc ttgtggaaag gacgaaacac cggtagttat taatagtaat
     2101 caattacggg gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg
     2161 taaatggccc gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt
     2221 atgttcccat agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac
     2281 ggtaaactgc ccacttggca gtacatcaag tgtatcatat gccaagtacg ccccctattg
     2341 acgtcaatga cggtaaatgg cccgcctggc attatgccca gtacatgacc ttatgggact
     2401 ttcctacttg gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt
     2461 ggcagtacat caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc
     2521 ccattgacgt caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc
     2581 gtaacaactc cgccccattg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata
     2641 taagcagagc tggtttagtg aaccgtcaga tccgctagcg ctaccggtcg ccaccatgga
     2701 gagcgacgag agcggcctgc ccgccatgga gatcgagtgc cgcatcaccg gcaccctgaa
     2761 cggcgtggag ttcgagctgg tgggcggcgg agagggcacc cccgagcagg gccgcatgac
     2821 caacaagatg aagagcacca aaggcgccct gaccttcagc ccctacctgc tgagccacgt
     2881 gatgggctac ggcttctacc acttcggcac ctaccccagc ggctacgaga accccttcct
     2941 gcacgccatc aacaacggcg gctacaccaa cacccgcatc gagaagtacg aggacggcgg
     3001 cgtgctgcac gtgagcttca gctaccgcta cgaggccggc cgcgtgatcg gcgacttcaa
     3061 ggtgatgggc accggcttcc ccgaggacag cgtgatcttc accgacaaga tcatccgcag
     3121 caacgccacc gtggagcacc tgcaccccat gggcgataac gatctggatg gcagcttcac
     3181 ccgcaccttc agcctgcgcg acggcggcta ctacagctcc gtggtggaca gccacatgca
     3241 cttcaagagc gccatccacc ccagcatcct gcagaacggg ggccccatgt tcgccttccg
     3301 ccgcgtggag gaggatcaca gcaacaccga gctgggcatc gtggagtacc agcacgcctt
     3361 caagaccccg gatgcagatg ccggtgaaga ataaaattct cgacctcgag acaaatggca
     3421 gtattcatcc acaattttaa aagaaaaggg gggattgggg ggtacagtgc aggggaaaga
     3481 atagtagaca taatagcaac agacatacaa actaaagaat tacaaaaaca aattacaaaa
     3541 attcaaaatt ttcgggttta ttacagggac agcagagatc cactttggcc gcggctcgag
     3601 ggggttgggg ttgcgccttt tccaaggcag ccctgggttt gcgcagggac gcggctgctc
     3661 tgggcgtggt tccgggaaac gcagcggcgc cgaccctggg tctcgcacat tcttcacgtc
     3721 cgttcgcagc gtcacccgga tcttcgccgc tacccttgtg ggccccccgg cgacgcttcc
     3781 tgctccgccc ctaagtcggg aaggttcctt gcggttcgcg gcgtgccgga cgtgacaaac
     3841 ggaagccgca cgtctcacta gtaccctcgc agacggacag cgccagggag caatggcagc
     3901 gcgccgaccg cgatgggctg tggccaatag cggctgctca gcagggcgcg ccgagagcag
     3961 cggccgggaa ggggcggtgc gggaggcggg gtgtggggcg gtagtgtggg ccctgttcct
     4021 gcccgcgcgg tgttccgcat tctgcaagcc tccggagcgc acgtcggcag tcggctccct
     4081 cgttgaccga atcaccgacc tctctcccca gggggatccc tagccaccat gcctttgtct
     4141 caagaagaat ccaccctcat tgaaagagca acggctacaa tcaacagcat ccccatctct
     4201 gaagactaca gcgtcgccag cgcagctctc tctagcgacg gccgcatctt cactggtgtc
     4261 aatgtatatc attttactgg gggaccttgt gcagaactcg tggtgctggg cactgctgct
     4321 gctgcggcag ctggcaacct gacttgtatc gtcgcgatcg gaaatgagaa caggggcatc
     4381 ttgagcccct gcggacggtg ccgacaggtg cttctcgatc tgcatcctgg gatcaaagcc
     4441 atagtgaagg acagtgatgg acagccgacg gcagttggga ttcgtgaatt gctgccctct
     4501 ggttatgtgt gggagggcta aggtaccttt aagaccaatg acttacaagg cagctgtaga
     4561 tcttagccac tttttaaaag aaaagggggg actggaaggg ctaattcact cccaacgaag
     4621 acaagatctg ctttttgctt gtactgggtc tctctggtta gaccagatct gagcctggga
     4681 gctctctggc taactaggga acccactgct taagcctcaa taaagcttgc cttgagtgct
     4741 tcaagtagtg tgtgcccgtc tgttgtgtga ctctggtaac tagagatccc tcagaccctt
     4801 ttagtcagtg tggaaaatct ctagcagtag tagttcatgt catcttatta ttcagtattt
     4861 ataacttgca aagaaatgaa tatcagagag tgagaggaac ttgtttattg cagcttataa
     4921 tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca
     4981 ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctggc tctagctatc
     5041 ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc
     5101 catggctgac taattttttt tatttatgca gaggccgagg ccgcctcggc ctctgagcta
     5161 ttccagaagt agtgaggagg cttttttgga ggcctaggga cgtacccaat tcgccctata
     5221 gtgagtcgta ttacgcgcgc tcactggccg tcgttttaca acgtcgtgac tgggaaaacc
     5281 ctggcgttac ccaacttaat cgccttgcag cacatccccc tttcgccagc tggcgtaata
     5341 gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg cagcctgaat ggcgaatggg
     5401 acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg
     5461 ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca
     5521 cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta
     5581 gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc
     5641 catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg
     5701 gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct tttgatttat
     5761 aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta
     5821 acgcgaattt taacaaaata ttaacgctta caatttaggt ggcacttttc ggggaaatgt
     5881 gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag
     5941 acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga gtattcaaca
     6001 tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt ttgctcaccc
     6061 agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag tgggttacat
     6121 cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag aacgttttcc
     6181 aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta ttgacgccgg
     6241 gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtactcacc
     6301 agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca gtgctgccat
     6361 aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag gaccgaagga
     6421 gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc
     6481 ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg tagcaatggc
     6541 aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc ggcaacaatt
     6601 aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc
     6661 tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg gtatcattgc
     6721 agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga cggggagtca
     6781 ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca
     6841 ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa aacttcattt
     6901 ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta
     6961 acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg
     7021 agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc
     7081 ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag
     7141 cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa
     7201 gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc
     7261 cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc
     7321 gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta
     7381 caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag
     7441 aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct
     7501 tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga
     7561 gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc
     7621 ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt
     7681 atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata ccgctcgccg
     7741 cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc gcccaatacg
     7801 caaaccgcct ctccccgcgc gttggccgat tcattaatgc agctggcacg acaggtttcc
     7861 cgactggaaa gcgggcagtg agcgcaacgc aattaatgtg agttagctca ctcattaggc
     7921 accccaggct ttacacttta tgcttccggc tcgtatgttg tgtggaattg tgagcggata
     7981 acaatttcac acaggaaaca gctatgacca tgattacgcc aagcgcgcaa ttaaccctca
     8041 ctaaagggaa caaaagctgg agctgcaagc ttaa
//
