LOCUS       Exported                8455 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  Third generation lentiviral vector expressing GFP and puromycin.
ACCESSION   .
VERSION     .
KEYWORDS    pLenti-CMV-MCS-GFP-SV-puro
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8455)
  AUTHORS   Witwicka H, Hwang SY, Reyes-Gutierrez P, Jia H, Odgren PE, Donahue 
            LR, Birnbaum MJ, Odgren PR
  TITLE     Studies of OC-STAMP in Osteoclast Fusion: A New Knockout Mouse 
            Model, Rescue of Cell Fusion, and Transmembrane Topology.
  JOURNAL   PLoS One. 2015 Jun 4;10(6):e0128275. doi: 
            10.1371/journal.pone.0128275. eCollection 2015.
  PUBMED    26042409
REFERENCE   2  (bases 1 to 8455)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "PLoS One.";
            date: "2015-06-4"; volume: "10(6):e0128275. doi"; pages: " 
            10.1371/journal.pone.0128275. eCollection 2015"
FEATURES             Location/Qualifiers
     source          1..8455
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     protein_bind    113..134
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        149..179
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    187..203
                     /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     192..214
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     211..227
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     211..227
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     246..266
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        248..266
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        294..520
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             521..701
                     /label=5 LTR (truncated)
                     /note="truncated 5 long terminal repeat (LTR) from HIV-1"
     misc_feature    748..873
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1366..1599
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1784..1828
                     /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    2095..2211
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     enhancer        2234..2537
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2538..2741
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2692..2712
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2738..2762
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             2818..3537
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             2818..3534
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2863..2884)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3124..3143)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3471..3492
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(3582..3602)
                     /label=pBABE 3
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3587..3916
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3767..3902
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3926..4525
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(3926..3945)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4422..4442
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    4592..5180
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4645..4665)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5063..5074)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             5267..5500
                     /label=3 LTR (Delta-U3)
                     /note="self-inactivating 3 long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    5572..5693
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5609..5628)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5663..5682
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      5733..5868
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(5888..5907)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(5889..5907)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5917..5934)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5917..5933)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5926..5948)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      6075..6530
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6162..6181)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6372..6393
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        6556..6660
                     /gene="bla"
                     /label=AmpR promoter
     CDS             6661..7521
                     /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(6879..6898)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7692..8280
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8181..8200
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     8434..8451
                     /label=L4440
                     /note="L4440 vector, forward primer"