LOCUS       Exported                9151 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  There is no gene/insert. It is a backbone plasmid that others can 
            use to insert sgRNAs of interest. The cloning site for this BsmBI.
ACCESSION   .
VERSION     .
KEYWORDS    pLenti-sgRNA
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9151)
  TITLE     pLenti-sgRNA
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 9151)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
FEATURES             Location/Qualifiers
     source          1..9151
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     138..157
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     391..408
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    525..546
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        561..591
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    599..615
                     /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     604..626
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     623..639
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     623..639
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     658..678
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        660..678
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        706..932
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             933..1113
                     /label=5 LTR (truncated)
                     /note="truncated 5 long terminal repeat (LTR) from HIV-1"
     misc_feature    1160..1285
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1778..2011
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2196..2240
                     /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        2418..2658
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA"
     primer_bind     2418..2438
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2589..2608
                     /label=LKO.1 5
                     /note="Human U6 promoter, forward primer"
     misc_RNA        4548..4623
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     misc_feature    4671..4788
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        4857..5068
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     CDS             5094..5693
                     /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(5094..5113)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5590..5610
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    5709..6297
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(5762..5782)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(6180..6191)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             6369..6602
                     /label=3 LTR (Delta-U3)
                     /note="self-inactivating 3 long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    6680..6801
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6717..6736)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6771..6790
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      6841..6976
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6903..6922
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     complement(6996..7015)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(6997..7015)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(7025..7042)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(7025..7041)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7034..7056)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      7183..7638
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7270..7289)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7480..7501
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        7664..7768
                     /gene="bla"
                     /label=AmpR promoter
     CDS             7769..8629
                     /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(7987..8006)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(8800..9151,1..237)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"