LOCUS       sequence-255273-        8949 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  puro exchanged to mCherry.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8949)
  TITLE     Emory Custom Cloning Core Plasmids - Oskar Laur
REFERENCE   2  (bases 1 to 8949)
  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"
COMMENT     Sequence Label: pLKO.1 mCherry
FEATURES             Location/Qualifiers
     source          1..8949
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     426..444
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             522..1232
                     /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(667..685)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     935..954
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     LTR             1304..1537
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    1609..1730
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1646..1665)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1700..1719
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      1770..1905
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(1925..1944)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1926..1944)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1954..1971)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1954..1970)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1963..1985)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2112..2567
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2199..2218)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2409..2430
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2593..2697
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2698..2766,2767..3558)
                     /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(2916..2935)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3729..4317
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4218..4237
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4471..4488
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    4605..4626
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4641..4646,4647..4664,4665..4671)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4679..4695
                     /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     4684..4706
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     4703..4719
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4703..4719
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     4738..4758
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        4740..4758
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        4786..5012
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             5013..5193
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    5240..5365
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    5858..6091
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             6276..6320
                     /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        6618..6866
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     6618..6638
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     6789..6808
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_feature    8780..8897
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        join(8946..8949,1..507)
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
ORIGIN
        1 ttggggttgc gccttttcca aggcagccct gggtttgcgc agggacgcgg ctgctctggg
       61 cgtggttccg ggaaacgcag cggcgccgac cctgggtctc gcacattctt cacgtccgtt
      121 cgcagcgtca cccggatctt cgccgctacc cttgtgggcc ccccggcgac gcttcctgct
      181 ccgcccctaa gtcgggaagg ttccttgcgg ttcgcggcgt gccggacgtg acaaacggaa
      241 gccgcacgtc tcactagtac cctcgcagac ggacagcgcc agggagcaat ggcagcgcgc
      301 cgaccgcgat gggctgtggc caatagcggc tgctcagcag ggcgcgccga gagcagcggc
      361 cgggaagggg cggtgcggga ggcggggtgt ggggcggtag tgtgggccct gttcctgccc
      421 gcgcggtgtt ccgcattctg caagcctccg gagcgcacgt cggcagtcgg ctccctcgtt
      481 gaccgaatca ccgacctctc tccccagggg gatcccttac catggtgagc aagggcgagg
      541 aggataacat ggccatcatc aaggagttca tgcgcttcaa ggtgcacatg gagggctccg
      601 tgaacggcca cgagttcgag atcgagggcg agggcgaggg ccgcccctac gagggcaccc
      661 agaccgccaa gctgaaggtg accaagggtg gccccctgcc cttcgcctgg gacatcctgt
      721 cccctcagtt catgtacggc tccaaggcct acgtgaagca ccccgccgac atccccgact
      781 acttgaagct gtccttcccc gagggcttca agtgggagcg cgtgatgaac ttcgaggacg
      841 gcggcgtggt gaccgtgacc caggactcct ccctgcagga cggcgagttc atctacaagg
      901 tgaagctgcg cggcaccaac ttcccctccg acggccccgt aatgcagaag aagaccatgg
      961 gctgggaggc ctcctccgag cggatgtacc ccgaggacgg cgccctgaag ggcgagatca
     1021 agcagaggct gaagctgaag gacggcggcc actacgacgc tgaggtcaag accacctaca
     1081 aggccaagaa gcccgtgcag ctgcccggcg cctacaacgt caacatcaag ttggacatca
     1141 cctcccacaa cgaggactac accatcgtgg aacagtacga acgcgccgag ggccgccact
     1201 ccaccggcgg catggacgag ctgtacaagt agggtacctt taagaccaat gacttacaag
     1261 gcagctgtag atcttagcca ctttttaaaa gaaaaggggg gactggaagg gctaattcac
     1321 tcccaacgaa gacaagatct gctttttgct tgtactgggt ctctctggtt agaccagatc
     1381 tgagcctggg agctctctgg ctaactaggg aacccactgc ttaagcctca ataaagcttg
     1441 ccttgagtgc ttcaagtagt gtgtgcccgt ctgttgtgtg actctggtaa ctagagatcc
     1501 ctcagaccct tttagtcagt gtggaaaatc tctagcagta gtagttcatg tcatcttatt
     1561 attcagtatt tataacttgc aaagaaatga atatcagaga gtgagaggaa cttgtttatt
     1621 gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt
     1681 ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgg
     1741 ctctagctat cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc
     1801 attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctcgg
     1861 cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggg acgtacccaa
     1921 ttcgccctat agtgagtcgt attacgcgcg ctcactggcc gtcgttttac aacgtcgtga
     1981 ctgggaaaac cctggcgtta cccaacttaa tcgccttgca gcacatcccc ctttcgccag
     2041 ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc caacagttgc gcagcctgaa
     2101 tggcgaatgg gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg
     2161 cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc
     2221 ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg
     2281 gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc
     2341 acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt
     2401 ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc
     2461 ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta
     2521 acaaaaattt aacgcgaatt ttaacaaaat attaacgctt acaatttagg tggcactttt
     2581 cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat
     2641 ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtatg
     2701 agtattcaac atttccgtgt cgcccttatt cccttttttg cggcattttg ccttcctgtt
     2761 tttgctcacc cagaaacgct ggtgaaagta aaagatgctg aagatcagtt gggtgcacga
     2821 gtgggttaca tcgaactgga tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa
     2881 gaacgttttc caatgatgag cacttttaaa gttctgctat gtggcgcggt attatcccgt
     2941 attgacgccg ggcaagagca actcggtcgc cgcatacact attctcagaa tgacttggtt
     3001 gagtactcac cagtcacaga aaagcatctt acggatggca tgacagtaag agaattatgc
     3061 agtgctgcca taaccatgag tgataacact gcggccaact tacttctgac aacgatcgga
     3121 ggaccgaagg agctaaccgc ttttttgcac aacatggggg atcatgtaac tcgccttgat
     3181 cgttgggaac cggagctgaa tgaagccata ccaaacgacg agcgtgacac cacgatgcct
     3241 gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg aactacttac tctagcttcc
     3301 cggcaacaat taatagactg gatggaggcg gataaagttg caggaccact tctgcgctcg
     3361 gcccttccgg ctggctggtt tattgctgat aaatctggag ccggtgagcg tgggtctcgc
     3421 ggtatcattg cagcactggg gccagatggt aagccctccc gtatcgtagt tatctacacg
     3481 acggggagtc aggcaactat ggatgaacga aatagacaga tcgctgagat aggtgcctca
     3541 ctgattaagc attggtaact gtcagaccaa gtttactcat atatacttta gattgattta
     3601 aaacttcatt tttaatttaa aaggatctag gtgaagatcc tttttgataa tctcatgacc
     3661 aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa
     3721 ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca
     3781 ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta
     3841 actggcttca gcagagcgca gataccaaat actgttcttc tagtgtagcc gtagttaggc
     3901 caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca
     3961 gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta
     4021 ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag
     4081 cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt
     4141 cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc
     4201 acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac
     4261 ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac
     4321 gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc
     4381 tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgcctttga gtgagctgat
     4441 accgctcgcc gcagccgaac gaccgagcgc agcgagtcag tgagcgagga agcggaagag
     4501 cgcccaatac gcaaaccgcc tctccccgcg cgttggccga ttcattaatg cagctggcac
     4561 gacaggtttc ccgactggaa agcgggcagt gagcgcaacg caattaatgt gagttagctc
     4621 actcattagg caccccaggc tttacacttt atgcttccgg ctcgtatgtt gtgtggaatt
     4681 gtgagcggat aacaatttca cacaggaaac agctatgacc atgattacgc caagcgcgca
     4741 attaaccctc actaaaggga acaaaagctg gagctgcaag cttaatgtag tcttatgcaa
     4801 tactcttgta gtcttgcaac atggtaacga tgagttagca acatgcctta caaggagaga
     4861 aaaagcaccg tgcatgccga ttggtggaag taaggtggta cgatcgtgcc ttattaggaa
     4921 ggcaacagac gggtctgaca tggattggac gaaccactga attgccgcat tgcagagata
     4981 ttgtatttaa gtgcctagct cgatacataa acgggtctct ctggttagac cagatctgag
     5041 cctgggagct ctctggctaa ctagggaacc cactgcttaa gcctcaataa agcttgcctt
     5101 gagtgcttca agtagtgtgt gcccgtctgt tgtgtgactc tggtaactag agatccctca
     5161 gaccctttta gtcagtgtgg aaaatctcta gcagtggcgc ccgaacaggg acttgaaagc
     5221 gaaagggaaa ccagaggagc tctctcgacg caggactcgg cttgctgaag cgcgcacggc
     5281 aagaggcgag gggcggcgac tggtgagtac gccaaaaatt ttgactagcg gaggctagaa
     5341 ggagagagat gggtgcgaga gcgtcagtat taagcggggg agaattagat cgcgatggga
     5401 aaaaattcgg ttaaggccag ggggaaagaa aaaatataaa ttaaaacata tagtatgggc
     5461 aagcagggag ctagaacgat tcgcagttaa tcctggcctg ttagaaacat cagaaggctg
     5521 tagacaaata ctgggacagc tacaaccatc ccttcagaca ggatcagaag aacttagatc
     5581 attatataat acagtagcaa ccctctattg tgtgcatcaa aggatagaga taaaagacac
     5641 caaggaagct ttagacaaga tagaggaaga gcaaaacaaa agtaagacca ccgcacagca
     5701 agcggccgct gatcttcaga cctggaggag gagatatgag ggacaattgg agaagtgaat
     5761 tatataaata taaagtagta aaaattgaac cattaggagt agcacccacc aaggcaaaga
     5821 gaagagtggt gcagagagaa aaaagagcag tgggaatagg agctttgttc cttgggttct
     5881 tgggagcagc aggaagcact atgggcgcag cgtcaatgac gctgacggta caggccagac
     5941 aattattgtc tggtatagtg cagcagcaga acaatttgct gagggctatt gaggcgcaac
     6001 agcatctgtt gcaactcaca gtctggggca tcaagcagct ccaggcaaga atcctggctg
     6061 tggaaagata cctaaaggat caacagctcc tggggatttg gggttgctct ggaaaactca
     6121 tttgcaccac tgctgtgcct tggaatgcta gttggagtaa taaatctctg gaacagattt
     6181 ggaatcacac gacctggatg gagtgggaca gagaaattaa caattacaca agcttaatac
     6241 actccttaat tgaagaatcg caaaaccagc aagaaaagaa tgaacaagaa ttattggaat
     6301 tagataaatg ggcaagtttg tggaattggt ttaacataac aaattggctg tggtatataa
     6361 aattattcat aatgatagta ggaggcttgg taggtttaag aatagttttt gctgtacttt
     6421 ctatagtgaa tagagttagg cagggatatt caccattatc gtttcagacc cacctcccaa
     6481 ccccgagggg acccgacagg cccgaaggaa tagaagaaga aggtggagag agagacagag
     6541 acagatccat tcgattagtg aacggatctc gacggtatcg atcacgagac tagcctcgag
     6601 cggccgcccc cttcaccgag ggcctatttc ccatgattcc ttcatatttg catatacgat
     6661 acaaggctgt tagagagata attggaatta atttgactgt aaacacaaag atattagtac
     6721 aaaatacgtg acgtagaaag taataatttc ttgggtagtt tgcagtttta aaattatgtt
     6781 ttaaaatgga ctatcatatg cttaccgtaa cttgaaagta tttcgatttc ttggctttat
     6841 atatcttgtg gaaaggacga aacaccggtg tgttgtaaat gagcacacaa aatacacatg
     6901 ctaaaatatt atattctatg acctttataa aatcaaccaa aatcttcttt ttaataactt
     6961 tagtatcaat aattagaatt tttatgttcc tttttgcaaa cttttaataa aaatgagcaa
     7021 aataaaaaaa cgctagtttt agtaactcgc gttgttttct tcacctttaa taatagctac
     7081 tccaccactt gttcctaagc ggtcagctcc tgcttcaatc attttttgag catcttcaaa
     7141 tgttctaact ccaccagctg ctttaactaa agcattgtct ttaacaactg acttcattag
     7201 tttaacatct tcaaatgttg cacctgattt tgaaaatcct gttgatgttt taacaaattc
     7261 taatccagct tcaacagcta tttcacaagc tttcatgatt tcttcttttg ttaataaaca
     7321 attttccata atacatttaa caacatgtga tccagctgct ttttttacag ctttcatgtc
     7381 ttctaaaact aattcataat ttttgtcttt taatgcacca atatttaata ccatatcaat
     7441 ttctgttgca ccatctttaa ttgcttcaga aacttcgaat gcttttgtag ctgttgtgca
     7501 tgcacctaga ggaaaaccta caacatttgt tattcctaca tttgtgcctt ttaataattc
     7561 tttacaatag cttgttcaat atgaattaac acaaactgtt gcaaaatcaa attcaattgc
     7621 ttcatcacat aattgtttaa tttcagcttt cgtagcatct tgttttaata atgtgtgatc
     7681 tatatatttg tttagtttca ttttttctcc tatatattca tttttaattt taattcttta
     7741 ataatttcgt ctactttaac tttagcgttt tgaacagatt caccaacacc tataaaataa
     7801 atttttagtt taggttcagt tccacttggg cgaacagcaa atcatgactt atcttctaaa
     7861 taaaatttta gtaagtcttg tcctggcata ttatacattc catcgatgta gtcttcaaca
     7921 ttaacaactt taagtccagc aatttgagtt aagggtgttg ctctcaatga tttcattaat
     7981 ggttcaattt ttaatttctt ttcttctggt ttaaaattca agtttaaagt gaaagtgtaa
     8041 tatgcaccca tttctttaaa taaatcttct aaatagtcta ctaatgtttt attttgtttt
     8101 ttataaaatc aagcagcctc tgctattaat atagaagctt gtattccatc tttatctcta
     8161 gctgagtcat caattacata tccataactt tcttcataag caaaaacaaa atttaatccg
     8221 ttatcttctt ctttagcaat ttctctaccc attcatttaa atccagttaa agtttttaca
     8281 atattaactc catatttttc atgagcgatt ctatcaccca aatcacttgt tacaaaactt
     8341 gaatatagag ccggattttt tggaatgcta tttaagcgtt ttagatttga taattttcaa
     8401 tcaattaaaa ttggtcctgt ttgatttcca tctaatctta caaaatgacc atcatgtttt
     8461 attgccattc caaatctgtc agcatctggg tcattcataa taataatatc tgcatcatgt
     8521 ttaataccat attcaagcgg tatttttcat gcaggatcaa attctggatt tggatttaca
     8581 acatttttaa atgtttcatc ttcaaatgca tgctcttcaa cctcaataac gttatatcct
     8641 gattcacgta atatttttgg ggtaaattta gttcctgttc cattaactgc gctaaaaata
     8701 atttttaaat cttttttagc ttcttgctct tttttgtaga attctcgacc tcgagacaaa
     8761 tggcagtatt catccacaat tttaaaagaa aaggggggat tggggggtac agtgcagggg
     8821 aaagaatagt agacataata gcaacagaca tacaaactaa agaattacaa aaacaaatta
     8881 caaaaattca aaattttcgg gtttattaca gggacagcag agatccactt tggccgcggc
     8941 tcgaggggg
//
