LOCUS       sequence-364237-        7208 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral construct for expression of mEmerald targeted to 
            peroxisomes.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7208)
  AUTHORS   Greiner D, Scott TM, Olson GS, Aderem A, Roh-Johnson M, Johnson JS
  TITLE     Genetic Modification of Primary Human Myeloid Cells to Study Cell 
            Migration, Activation, and Organelle Dynamics.
  JOURNAL   Curr Protoc. 2022 Aug;2(8):e514. doi: 10.1002/cpz1.514.
  PUBMED    36018279
REFERENCE   2  (bases 1 to 7208)
  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: "Curr 
            Protoc."; date: "2022-08"; volume: "2(8):e514. doi"; pages: " 
            10.1002/cpz1.514"
COMMENT     Sequence Label: pLKO-perox-mEm
FEATURES             Location/Qualifiers
     source          1..7208
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     14..33
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(252..356)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(382..837)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(519..540)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     731..750
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     964..986
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     978..995
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     979..995
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1005..1024
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1005..1023
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     rep_origin      1044..1179
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    1219..1340
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1230..1249)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1284..1303
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     LTR             1412..1645
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     CDS             complement(join(1808..2518,2519..2521,2522..2524))
                     /codon_start=1
                     /product="brightly fluorescent monomeric variant of GFP"
                     /label=mEmerald
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHKVYITADKQKNGIK
                     VNFKTRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1850..1871)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     2199..2218
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2458..2479
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     promoter        complement(2552..3062)
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     complement(2615..2633)
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     misc_feature    3111..3228
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        complement(3291..3539)
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     complement(3349..3368)
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(3519..3539)
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     CDS             complement(3837..3881)
                     /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    4066..4299
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     misc_feature    4792..4917
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     LTR             4964..5144
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     promoter        complement(5145..5371)
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     primer_bind     complement(5399..5419)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(5399..5417)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5438..5454)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5438..5454)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5451..5473)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5462..5478
                     /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(5486..5492,5493..5510,5511..5516))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5531..5552
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5669..5686)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5840..6428)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5920..5939)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6599..7208,1..182,183..251))
                     /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"
ORIGIN
        1 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
       61 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
      121 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
      181 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
      241 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
      301 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
      361 tttccccgaa aagtgccacc taaattgtaa gcgttaatat tttgttaaaa ttcgcgttaa
      421 atttttgtta aatcagctca ttttttaacc aataggccga aatcggcaaa atcccttata
      481 aatcaaaaga atagaccgag atagggttga gtgttgttcc agtttggaac aagagtccac
      541 tattaaagaa cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc
      601 cactacgtga accatcaccc taatcaagtt ttttggggtc gaggtgccgt aaagcactaa
      661 atcggaaccc taaagggagc ccccgattta gagcttgacg gggaaagccg gcgaacgtgg
      721 cgagaaagga agggaagaaa gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg
      781 tcacgctgcg cgtaaccacc acacccgccg cgcttaatgc gccgctacag ggcgcgtccc
      841 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
      901 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt
      961 tttcccagtc acgacgttgt aaaacgacgg ccagtgagcg cgcgtaatac gactcactat
     1021 agggcgaatt gggtacgtcc ctaggcctcc aaaaaagcct cctcactact tctggaatag
     1081 ctcagaggcc gaggcggcct cggcctctgc ataaataaaa aaaattagtc agccatgggg
     1141 cggagaatgg gcggaactgg gcggagttag gggcgggatg ggcggagtta ggggcgggat
     1201 agctagagcc agacatgata agatacattg atgagtttgg acaaaccaca actagaatgc
     1261 agtgaaaaaa atgctttatt tgtgaaattt gtgatgctat tgctttattt gtaaccatta
     1321 taagctgcaa taaacaagtt cctctcactc tctgatattc atttctttgc aagttataaa
     1381 tactgaataa taagatgaca tgaactacta ctgctagaga ttttccacac tgactaaaag
     1441 ggtctgaggg atctctagtt accagagtca cacaacagac gggcacacac tacttgaagc
     1501 actcaaggca agctttattg aggcttaagc agtgggttcc ctagttagcc agagagctcc
     1561 caggctcaga tctggtctaa ccagagagac ccagtacaag caaaaagcag atcttgtctt
     1621 cgttgggagt gaattagccc ttccagtccc cccttttctt ttaaaaagtg gctaagatct
     1681 acagctgcct tgtaagtcat tggtcttaaa ggtaccgatg catggggtcg tgcgctcctt
     1741 tcggtcgggc gctgcgggtc gtggggcggg cgttacttgt acagatatcc gtacgttaca
     1801 gcttggactt gtacagctcg tccatgccga gagtgatccc ggcggcggtc acgaactcca
     1861 gcaggaccat gtgatcgcgc ttctcgttgg ggtctttgct cagcttggac tgggtgctca
     1921 ggtagtggtt gtcgggcagc agcacggggc cgtcgccgat gggggtgttc tgctggtagt
     1981 ggtcggcgag ctgcacgctg ccgtcctcga tgttgtggcg ggtcttgaag ttcaccttga
     2041 tgccgttctt ctgcttgtcg gcggtgatat agaccttgtg gctgttgtag ttgtactcca
     2101 gcttgtgccc caggatgttg ccgtcctcct tgaagtcgat gcccttcagc tcgatgcggt
     2161 tcaccagggt gtcgccctcg aacttcacct cggcgcgggt cttgtagttg ccgtcgtcct
     2221 tgaagaagat ggtgcgctcc tggacgtagc cttcgggcat ggcggacttg aagaagtcgt
     2281 gctgcttcat gtggtcgggg tagcgggcga agcactgcac gccgtaggtc aaggtggtca
     2341 cgagggtggg ccagggcacg ggcagcttgc cggtggtgca gatgaacttc agggtcagct
     2401 tgccgtaggt ggcatcgccc tcgccctcgc cggacacgct gaacttgtgg ccgtttacgt
     2461 cgccgtccag ctcgaccagg atgggcacca ccccggtgaa cagctcctcg cccttgctca
     2521 ccatggtaag ctccggtgac gtcggatccc cctggggaga gaggtcggtg attcggtcaa
     2581 cgagggagcc gactgccgac gtgcgctccg gaggcttgca gaatgcggaa caccgcgcgg
     2641 gcaggaacag ggcccacact accgccccac accccgcctc ccgcaccgcc ccttcccggc
     2701 cgctgctctc ggcgcgccct gctgagcagc cgctattggc cacagcccat cgcggtcggc
     2761 gcgctgccat tgctccctgg cgctgtccgt ctgcgagggt actagtgaga cgtgcggctt
     2821 ccgtttgtca cgtccggcac gccgcgaacc gcaaggaacc ttcccgactt aggggcggag
     2881 caggaagcgt cgccgggggg cccacaaggg tagcggcgaa gatccgggtg acgctgcgaa
     2941 cggacgtgaa gaatgtgcga gacccagggt cggcgccgct gcgtttcccg gaaccacgcc
     3001 cagagcagcc gcgtccctgc gcaaacccag ggctgccttg gaaaaggcgc aaccccaacc
     3061 ccctcgagcc gcggccaaag tggatctctg ctgtccctgt aataaacccg aaaattttga
     3121 atttttgtaa tttgtttttg taattcttta gtttgtatgt ctgttgctat tatgtctact
     3181 attctttccc ctgcactgta ccccccaatc cccccttttc ttttaaaatt gtggatgaat
     3241 actgccattt gtctcgaggt cgagaattca cgcgtgcata cctgcggacc ggtgtttcgt
     3301 cctttccaca agatatataa agccaagaaa tcgaaatact ttcaagttac ggtaagcata
     3361 tgatagtcca ttttaaaaca taattttaaa actgcaaact acccaagaaa ttattacttt
     3421 ctacgtcacg tattttgtac taatatcttt gtgtttacag tcaaattaat tccaattatc
     3481 tctctaacag ccttgtatcg tatatgcaaa tatgaaggaa tcatgggaaa taggccctcg
     3541 gtgaaggggg cggccgctcg aggctagtct cgtgatcgat accgtcgaga tccgttcact
     3601 aatcgaatgg atctgtctct gtctctctct ccaccttctt cttctattcc ttcgggcctg
     3661 tcgggtcccc tcggggttgg gaggtgggtc tgaaacgata atggtgaata tccctgccta
     3721 actctattca ctatagaaag tacagcaaaa actattctta aacctaccaa gcctcctact
     3781 atcattatga ataattttat ataccacagc caatttgtta tgttaaacca attccacaaa
     3841 cttgcccatt tatctaattc caataattct tgttcattct tttcttgctg gttttgcgat
     3901 tcttcaatta aggagtgtat taagcttgtg taattgttaa tttctctgtc ccactccatc
     3961 caggtcgtgt gattccaaat ctgttccaga gatttattac tccaactagc attccaaggc
     4021 acagcagtgg tgcaaatgag ttttccagag caaccccaaa tccccaggag ctgttgatcc
     4081 tttaggtatc tttccacagc caggattctt gcctggagct gcttgatgcc ccagactgtg
     4141 agttgcaaca gatgctgttg cgcctcaata gccctcagca aattgttctg ctgctgcact
     4201 ataccagaca ataattgtct ggcctgtacc gtcagcgtca ttgacgctgc gcccatagtg
     4261 cttcctgctg ctcccaagaa cccaaggaac aaagctccta ttcccactgc tcttttttct
     4321 ctctgcacca ctcttctctt tgccttggtg ggtgctactc ctaatggttc aatttttact
     4381 actttatatt tatataattc acttctccaa ttgtccctca tatctcctcc tccaggtctg
     4441 aagatcagcg gccgcttgct gtgcggtggt cttacttttg ttttgctctt cctctatctt
     4501 gtctaaagct tccttggtgt cttttatctc tatcctttga tgcacacaat agagggttgc
     4561 tactgtatta tataatgatc taagttcttc tgatcctgtc tgaagggatg gttgtagctg
     4621 tcccagtatt tgtctacagc cttctgatgt ttctaacagg ccaggattaa ctgcgaatcg
     4681 ttctagctcc ctgcttgccc atactatatg ttttaattta tattttttct ttccccctgg
     4741 ccttaaccga attttttccc atcgcgatct aattctcccc cgcttaatac tgacgctctc
     4801 gcacccatct ctctccttct agcctccgct agtcaaaatt tttggcgtac tcaccagtcg
     4861 ccgcccctcg cctcttgccg tgcgcgcttc agcaagccga gtcctgcgtc gagagagctc
     4921 ctctggtttc cctttcgctt tcaagtccct gttcgggcgc cactgctaga gattttccac
     4981 actgactaaa agggtctgag ggatctctag ttaccagagt cacacaacag acgggcacac
     5041 actacttgaa gcactcaagg caagctttat tgaggcttaa gcagtgggtt ccctagttag
     5101 ccagagagct cccaggctca gatctggtct aaccagagag acccgtttat gtatcgagct
     5161 aggcacttaa atacaatatc tctgcaatgc ggcaattcag tggttcgtcc aatccatgtc
     5221 agacccgtct gttgccttcc taataaggca cgatcgtacc accttacttc caccaatcgg
     5281 catgcacggt gctttttctc tccttgtaag gcatgttgct aactcatcgt taccatgttg
     5341 caagactaca agagtattgc ataagactac attaagcttg cagctccagc ttttgttccc
     5401 tttagtgagg gttaattgcg cgcttggcgt aatcatggtc atagctgttt cctgtgtgaa
     5461 attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct
     5521 ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc
     5581 agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg
     5641 gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
     5701 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag
     5761 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
     5821 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
     5881 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     5941 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     6001 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
     6061 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     6121 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     6181 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     6241 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg
     6301 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     6361 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     6421 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     6481 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa
     6541 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
     6601 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
     6661 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
     6721 gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc
     6781 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt
     6841 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg
     6901 ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca
     6961 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg
     7021 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca
     7081 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg
     7141 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct
     7201 cttgcccg
//
