LOCUS       sequence-391775-        8657 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral vector for expression of sgRNA with mCherry, zeocin 
            resistance, BlpI + BstXI cloning sites.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8657)
  AUTHORS   McCollum CO, Didychuk AL, Liu D, Murray-Nerger LA, Cristea IM, 
            Glaunsinger BA
  TITLE     The viral packaging motor potentiates Kaposi's sarcoma-associated 
            herpesvirus gene expression late in infection.
  JOURNAL   PLoS Pathog. 2023 Apr 17;19(4):e1011163. doi: 
            10.1371/journal.ppat.1011163. eCollection 2023 Apr.
  PUBMED    37068108
REFERENCE   2  (bases 1 to 8657)
  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: "PLoS 
            Pathog."; date: "2023-04-17"; volume: "19(4):e1011163. doi"; pages: 
            " 10.1371/journal.ppat.1011163. eCollection 2023 Apr"
COMMENT     Sequence Label: pDM002-sgRNA-lenti-ZeoR
FEATURES             Location/Qualifiers
     source          1..8657
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(47..66)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        238..617
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        619..817
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     772..792
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             835..1015
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1062..1187
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1684..1917
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2102..2146
                     /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"
     misc_feature    2444..2561
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     protein_bind    complement(2814..2847)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     2851..2871
                     /label=mU6-F
                     /note="Mouse U6 promoter, forward primer"
     promoter        3116..3327
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     intron          3328..4266
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     4223..4243
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             4281..4652
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     CDS             4662..4715
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     regulatory      4722..4731
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             4728..5438
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(4873..4891)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     5141..5160
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     protein_bind    complement(5457..5490)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    5546..6134
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(5599..5619)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(6017..6028)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(6137..6153)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6138..6154)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             6663..6843
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(6905..7493)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6985..7004)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7664..8455,8456..8524))
                     /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     8287..8306
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8525..8629)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgacggat cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg
       61 atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt
      121 gcgcgagcaa aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc
      181 tgcttagggt taggcgtttt gcgctgcttc gcgatgtacg ggccagatat acgcgttgac
      241 attgattatt gactagttat taatagtaat caattacggg gtcattagtt catagcccat
      301 atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg
      361 acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt
      421 tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag
      481 tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc
      541 attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag
      601 tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt
      661 ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc
      721 accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg
      781 gcggtaggcg tgtacggtgg gaggtctata taagcagcgc gttttgcctg tactgggtct
      841 ctctggttag accagatctg agcctgggag ctctctggct aactagggaa cccactgctt
      901 aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac
      961 tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtggc
     1021 gcccgaacag ggacttgaaa gcgaaaggga aaccagagga gctctctcga cgcaggactc
     1081 ggcttgctga agcgcgcacg gcaagaggcg aggggcggcg actggtgagt acgccaaaaa
     1141 ttttgactag cggaggctag aaggagagag atgggtgcga gagcgtcagt attaagcggg
     1201 ggagaattag atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag aaaaaatata
     1261 aattaaaaca tatagtatgg gcaagcaggg agctagaacg attcgcagtt aatcctggcc
     1321 tgttagaaac atcagaaggc tgtagacaaa tactgggaca gctacaacca tcccttcaga
     1381 caggatcaga agaacttaga tcattatata atacagtagc aaccctctat tgtgtgcatc
     1441 aaaggataga gataaaagac accaaggaag ctttagacaa gatagaggaa gagcaaaaca
     1501 aaagtaagac caccgcacag caagcggccg gccgctgatc ttcagacctg gaggaggaga
     1561 tatgagggac aattggagaa gtgaattata taaatataaa gtagtaaaaa ttgaaccatt
     1621 aggagtagca cccaccaagg caaagagaag agtggtgcag agagaaaaaa gagcagtggg
     1681 aataggagct ttgttccttg ggttcttggg agcagcagga agcactatgg gcgcagcgtc
     1741 aatgacgctg acggtacagg ccagacaatt attgtctggt atagtgcagc agcagaacaa
     1801 tttgctgagg gctattgagg cgcaacagca tctgttgcaa ctcacagtct ggggcatcaa
     1861 gcagctccag gcaagaatcc tggctgtgga aagataccta aaggatcaac agctcctggg
     1921 gatttggggt tgctctggaa aactcatttg caccactgct gtgccttgga atgctagttg
     1981 gagtaataaa tctctggaac agatttggaa tcacacgacc tggatggagt gggacagaga
     2041 aattaacaat tacacaagct taatacactc cttaattgaa gaatcgcaaa accagcaaga
     2101 aaagaatgaa caagaattat tggaattaga taaatgggca agtttgtgga attggtttaa
     2161 cataacaaat tggctgtggt atataaaatt attcataatg atagtaggag gcttggtagg
     2221 tttaagaata gtttttgctg tactttctat agtgaataga gttaggcagg gatattcacc
     2281 attatcgttt cagacccacc tcccaacccc gaggggaccc gacaggcccg aaggaataga
     2341 agaagaaggt ggagagagag acagagacag atccattcga ttagtgaacg gatcggcact
     2401 gcgtgcgcca attctgcaga caaatggcag tattcatcca caattttaaa agaaaagggg
     2461 ggattggggg gtacagtgca ggggaaagaa tagtagacat aatagcaaca gacatacaaa
     2521 ctaaagaatt acaaaaacaa attacaaaaa ttcaaaattt tcgggtttat tacagggaca
     2581 gcagagatcc agtttggtta gtaccgggcc cgctctagag atccgacgcg ccatctctag
     2641 gcccgcgccg gccccctcgc acggacttgt gggagaagct cggctactcc cctgccccgg
     2701 ttaatttgca tataatattt cctagtaact atagaggctt aatgtgcgat aaaagacaga
     2761 taatctgttc tttttaatac tagctacatt ttacatgata ggcttggatt tctataactt
     2821 cgtatagcat acattatacg aagttataaa cagcacaaaa ggaaactcac cctaactgta
     2881 aagtaattgt gtgttttgag actataagta tcccttggag aaccaccttg ttggaccagg
     2941 atgggcacca cccgtttaag agctaagctg gaaacagcat agcaagttta aataaggcta
     3001 gtccgttatc aacttgaaaa agtggcaccg agtcggtgct ttttttctcg agtactagga
     3061 tccattaggc ggccgcgtgg ataaccgtat taccgccatg catgtgcccg tcagtgggca
     3121 gagcgcacat cgcccacagt ccccgagaag ttggggggag gggtcggcaa ttgaaccggt
     3181 gcctagagaa ggtggcgcgg ggtaaactgg gaaagtgatg tcgtgtactg gctccgcctt
     3241 tttcccgagg gtgggggaga accgtatata agtgcagtag tcgccgtgaa cgttcttttt
     3301 cgcaacgggt ttgccgccag aacacaggta agtgccgtgt gtggttcccg cgggcctggc
     3361 ctctttacgg gttatggccc ttgcgtgcct tgaattactt ccacctggct gcagtacgtg
     3421 attcttgatc ccgagcttcg ggttggaagt gggtgggaga gttcgaggcc ttgcgcttaa
     3481 ggagcccctt cgcctcgtgc ttgagttgag gcctggcctg ggcgctgggg ccgccgcgtg
     3541 cgaatctggt ggcaccttcg cgcctgtctc gctgctttcg ataagtctct agccatttaa
     3601 aatttttgat gacctgctgc gacgcttttt ttctggcaag atagtcttgt aaatgcgggc
     3661 caagatctgc acactggtat ttcggttttt ggggccgcgg gcggcgacgg ggcccgtgcg
     3721 tcccagcgca catgttcggc gaggcggggc ctgcgagcgc ggccaccgag aatcggacgg
     3781 gggtagtctc aagctggccg gcctgctctg gtgcctggcc tcgcgccgcc gtgtatcgcc
     3841 ccgccctggg cggcaaggct ggcccggtcg gcaccagttg cgtgagcgga aagatggccg
     3901 cttcccggcc ctgctgcagg gagctcaaaa tggaggacgc ggcgctcggg agagcgggcg
     3961 ggtgagtcac ccacacaaag gaaaagggcc tttccgtcct cagccgtcgc ttcatgtgac
     4021 tccacggagt accgggcgcc gtccaggcac ctcgattagt tctcgcgctt ttggagtacg
     4081 tcgtctttag gttgggggga ggggttttat gcgatggagt ttccccacac tgagtgggtg
     4141 gagactgaag ttaggccagc ttggcacttg atgtaattct ccttggaatt tgcccttttt
     4201 gagtttggat cttggttcat tctcaagcct cagacagtgg ttcaaagttt ttttcttcca
     4261 tttcaggtgt cgtgagctag atggccaagt tgaccagtgc cgttccggtg ctcaccgcgc
     4321 gcgacgtcgc cggagcggtc gagttctgga ccgaccggct cgggttctcc cgggacttcg
     4381 tggaggacga cttcgccggt gtggtccggg acgacgtgac cctgttcatc agcgcggtcc
     4441 aggaccaggt ggtgccggac aacaccctgg cctgggtgtg ggtgcgcggc ctggacgagc
     4501 tgtacgccga gtggtcggag gtcgtgtcca cgaacttccg ggacgcctcc gggccggcca
     4561 tgaccgagat cggcgagcag ccgtgggggc gggagttcgc cctgcgcgac ccggccggca
     4621 actgcgtgca cttcgtggcc gaggagcagg acggatcggg agagggcaga ggaagtctgc
     4681 taacatgcgg tgacgtcgag gagaatcctg gcccaccggt cgccaccatg gtgagcaagg
     4741 gcgaggagga taacatggcc atcatcaagg agttcatgcg cttcaaggtg cacatggagg
     4801 gctccgtgaa cggccacgag ttcgagatcg agggcgaggg cgagggccgc ccctacgagg
     4861 gcacccagac cgccaagctg aaggtgacca agggtggccc cctgcccttc gcctgggaca
     4921 tcctgtcccc tcagttcatg tacggctcca aggcctacgt gaagcacccc gccgacatcc
     4981 ccgactactt gaagctgtcc ttccccgagg gcttcaagtg ggagcgcgtg atgaacttcg
     5041 aggacggcgg cgtggtgacc gtgacccagg actcctccct gcaggacggc gagttcatct
     5101 acaaggtgaa gctgcgcggc accaacttcc cctccgacgg ccccgtaatg cagaagaaga
     5161 ccatgggctg ggaggcctcc tccgagcgga tgtaccccga ggacggcgcc ctgaagggcg
     5221 agatcaagca gaggctgaag ctgaaggacg gcggccacta cgacgctgag gtcaagacca
     5281 cctacaaggc caagaagccc gtgcagctgc ccggcgccta caacgtcaac atcaagttgg
     5341 acatcacctc ccacaacgag gactacacca tcgtggaaca gtacgaacgc gccgagggcc
     5401 gccactccac cggcggcatg gacgagctgt acaagtaaaa ttcgtcgagg gacctaataa
     5461 cttcgtatag catacattat acgaagttat acatgtttaa gggttccggt tccactaggt
     5521 acaattcgat atcaagctta tcgataatca acctctggat tacaaaattt gtgaaagatt
     5581 gactggtatt cttaactatg ttgctccttt tacgctatgt ggatacgctg ctttaatgcc
     5641 tttgtatcat gctattgctt cccgtatggc tttcattttc tcctccttgt ataaatcctg
     5701 gttgctgtct ctttatgagg agttgtggcc cgttgtcagg caacgtggcg tggtgtgcac
     5761 tgtgtttgct gacgcaaccc ccactggttg gggcattgcc accacctgtc agctcctttc
     5821 cgggactttc gctttccccc tccctattgc cacggcggaa ctcatcgccg cctgccttgc
     5881 ccgctgctgg acaggggctc ggctgttggg cactgacaat tccgtggtgt tgtcggggaa
     5941 atcatcgtcc tttccttggc tgctcgcctg tgttgccacc tggattctgc gcgggacgtc
     6001 cttctgctac gtcccttcgg ccctcaatcc agcggacctt ccttcccgcg gcctgctgcc
     6061 ggctctgcgg cctcttccgc gtcttcgcct tcgccctcag acgagtcgga tctccctttg
     6121 ggccgcctcc ccgcatcgat accgtcgacc tcgatcgaga cctagaaaaa catggagcaa
     6181 tcacaagtag caatacagca gctaccaatg ctgattgtgc ctggctagaa gcacaagagg
     6241 aggaggaggt gggttttcca gtcacacctc aggtaccttt aagaccaatg acttacaagg
     6301 cagctgtaga tcttagccac tttttaaaag aaaagggggg actggaaggg ctaattcact
     6361 cccaacgaag acaagatatc cttgatctgt ggatctacca cacacaaggc tacttccctg
     6421 attggcagaa ctacacacca gggccaggga tcagatatcc actgaccttt ggatggtgct
     6481 acaagctagt accagttgag caagagaagg tagaagaagc caatgaagga gagaacaccc
     6541 gcttgttaca ccctgtgagc ctgcatggga tggatgaccc ggagagagaa gtattagagt
     6601 ggaggtttga cagccgccta gcatttcatc acatggcccg agagctgcat ccggactgta
     6661 ctgggtctct ctggttagac cagatctgag cctgggagct ctctggctaa ctagggaacc
     6721 cactgcttaa gcctcaataa agcttgcctt gagtgcttca agtagtgtgt gcccgtctgt
     6781 tgtgtgactc tggtaactag agatccctca gaccctttta gtcagtgtgg aaaatctcta
     6841 gcagcatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg
     6901 cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga
     6961 ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg
     7021 tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg
     7081 gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc
     7141 gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg
     7201 gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca
     7261 ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt
     7321 ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag
     7381 ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg
     7441 gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc
     7501 ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt
     7561 tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt
     7621 ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca
     7681 gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg
     7741 tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac
     7801 cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg
     7861 ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc
     7921 gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta
     7981 caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac
     8041 gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc
     8101 ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac
     8161 tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact
     8221 caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa
     8281 tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt
     8341 cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca
     8401 ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa
     8461 aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac
     8521 tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg
     8581 gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc
     8641 gaaaagtgcc acctgac
//
