LOCUS       sequence-387464-        8938 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral guide RNA backbone derived from pSB700 with puromycin.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8938)
  TITLE     Various CRISPR/Cas9 backbones
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 8938)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pGMC00017 (aka pRC0162)
FEATURES             Location/Qualifiers
     source          1..8938
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             1..634
                     /label=3' LTR
                     /note="3' long terminal repeat (LTR) from HIV-1"
     misc_feature    681..806
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1303..1536
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1721..1765
                     /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    2028..2143
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1 (lacking the first T)"
     promoter        2246..2494
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     2246..2266
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2417..2436
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        2516..2591
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     enhancer        2665..2968
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2970..3243
                     /label=chicken beta-actin promoter
     intron          3244..4260
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     4184..4206
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     4268..4287
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             4294..4893
                     /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(4294..4313)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4790..4810
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    4906..5494
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4959..4979)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5377..5388)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             5701..6334
                     /label=3' LTR
                     /note="3' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(6463..6479)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6463..6479)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6476..6498)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6487..6503
                     /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(6511..6517,6518..6535,6536..6541))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6556..6577
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6694..6711)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6865..7453)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6945..6964)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7624..8415,8416..8484))
                     /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     8247..8266
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8485..8589)
                     /gene="bla"
                     /label=AmpR promoter
     polyA_signal    8637..8771
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(8674..8693)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     8728..8747
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
ORIGIN
        1 tggaagggct aattcactcc caaagaagac aagatatcct tgatctgtgg atctaccaca
       61 cacaaggcta cttccctgat tagcagaact acacaccagg gccaggggtc agatatccac
      121 tgacctttgg atggtgctac aagctagtac cagttgagcc agataaggta gaagaggcca
      181 ataaaggaga gaacaccagc ttgttacacc ctgtgagcct gcatgggatg gatgacccgg
      241 agagagaagt gttagagtgg aggtttgaca gccgcctagc atttcatcac gtggcccgag
      301 agctgcatcc ggagtacttc aagaactgct gatatcgagc ttgctacaag ggactttccg
      361 ctggggactt tccagggagg cgtggcctgg gcgggactgg ggagtggcga gccctcagat
      421 cctgcatata agcagctgct ttttgcctgt actgggtctc tctggttaga ccagatctga
      481 gcctgggagc tctctggcta actagggaac ccactgctta agcctcaata aagcttgcct
      541 tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta gagatccctc
      601 agaccctttt agtcagtgtg gaaaatctct agcagtggcg cccgaacagg gacttgaaag
      661 cgaaagggaa accagaggag ctctctcgac gcaggactcg gcttgctgaa gcgcgcacgg
      721 caagaggcga ggggcggcga ctggtgagta cgccaaaaat tttgactagc ggaggctaga
      781 aggagagaga tgggtgcgag agcgtcagta ttaagcgggg gagaattaga tcgcgatggg
      841 aaaaaattcg gttaaggcca gggggaaaga aaaaatataa attaaaacat atagtatggg
      901 caagcaggga gctagaacga ttcgcagtta atcctggcct gttagaaaca tcagaaggct
      961 gtagacaaat actgggacag ctacaaccat cccttcagac aggatcagaa gaacttagat
     1021 cattatataa tacagtagca accctctatt gtgtgcatca aaggatagag ataaaagaca
     1081 ccaaggaagc tttagacaag atagaggaag agcaaaacaa aagtaagacc accgcacagc
     1141 aagcggccgg ccgctgatct tcagacctgg aggaggagat atgagggaca attggagaag
     1201 tgaattatat aaatataaag tagtaaaaat tgaaccatta ggagtagcac ccaccaaggc
     1261 aaagagaaga gtggtgcaga gagaaaaaag agcagtggga ataggagctt tgttccttgg
     1321 gttcttggga gcagcaggaa gcactatggg cgcagcgtca atgacgctga cggtacaggc
     1381 cagacaatta ttgtctggta tagtgcagca gcagaacaat ttgctgaggg ctattgaggc
     1441 gcaacagcat ctgttgcaac tcacagtctg gggcatcaag cagctccagg caagaatcct
     1501 ggctgtggaa agatacctaa aggatcaaca gctcctgggg atttggggtt gctctggaaa
     1561 actcatttgc accactgctg tgccttggaa tgctagttgg agtaataaat ctctggaaca
     1621 gatttggaat cacacgacct ggatggagtg ggacagagaa attaacaatt acacaagctt
     1681 aatacactcc ttaattgaag aatcgcaaaa ccagcaagaa aagaatgaac aagaattatt
     1741 ggaattagat aaatgggcaa gtttgtggaa ttggtttaac ataacaaatt ggctgtggta
     1801 tataaaatta ttcataatga tagtaggagg cttggtaggt ttaagaatag tttttgctgt
     1861 actttctata gtgaatagag ttaggcaggg atattcacca ttatcgtttc agacccacct
     1921 cccaaccccg aggggacccg acaggcccga aggaatagaa gaagaaggtg gagagagaga
     1981 cagagacaga tccattcgat tagtgaacgg atctcgacgg tatcgccttt aaaagaaaag
     2041 gggggattgg ggggtacagt gcaggggaaa gaatagtaga cataatagca acagacatac
     2101 aaactaaaga attacaaaaa caaattacaa aaattcaaaa ttttcgggtt tattacaggg
     2161 acagcagaga tccagtttat cattagtgaa cggatctcga cggtatcgat cacgagacta
     2221 gcctcgagcg gccgccccct tcaccgaggg cctatttccc atgattcctt catatttgca
     2281 tatacgatac aaggctgtta gagagataat tggaattaat ttgactgtaa acacaaagat
     2341 attagtacaa aatacgtgac gtagaaagta ataatttctt gggtagtttg cagttttaaa
     2401 attatgtttt aaaatggact atcatatgct taccgtaact tgaaagtatt tcgatttctt
     2461 ggctttatat atcttgtgga aaggacgaaa caccggagac gattaatgcg tctcggtttt
     2521 agagctagaa atagcaagtt aaaataaggc tagtccgtta tcaacttgaa aaagtggcac
     2581 cgagtcggtg ctttttttga attctcgacc tcgagacaaa tggcagtatt cgtcattagt
     2641 tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg
     2701 accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc
     2761 aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc
     2821 agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg
     2881 gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat
     2941 ctacgtatta gtcatcgcta ttaccatggt cgaggtgagc cccacgttct gcttcactct
     3001 ccccatctcc cccccctccc cacccccaat tttgtattta tttatttttt aattattttg
     3061 tgcagcgatg ggggcggggg gggggggcgc gcgccaggcg gggcggggcg gggcgrgggg
     3121 cggggcgggg cgaggcggag aggtgcggcg gcagccaatc agagcggcgc gctccgaaag
     3181 tttcctttta tggcgaggcg gcggcggcgg cggccctata aaaagcgaag cgcgcggcgg
     3241 gcgggagtcg ctgcgcgctg ccttcgcccc gtgccccgct ccgccgccgc ctcgcgccgc
     3301 ccgccccggc tctgactgac cgcgttactc ccacaggtga gcgggcggga cggcccttct
     3361 cctccgggct gtaattagcg cttggtttaa tgacggcttg tttcttttct gtggctgcgt
     3421 gaaagccttg aggggctccg ggagggccct ttgtgcgggg ggagcggctc ggggggtgcg
     3481 tgcgtgtgtg tgtgcgtggg gagcgccgcg tgcggctccg cgctgcccgg cggctgtgag
     3541 cgctgcgggc gcggcgcggg gctttgtgcg ctccgcagtg tgcgcgaggg gagcgcggcc
     3601 gggggcggtg ccccgcggtg cggggggggc tgcgagggga acaaaggctg cgtgcggggt
     3661 gtgtgcgtgg gggggtgagc agggggtgtg ggcgcgtcgg tcgggctgca accccccctg
     3721 cacccccctc cccgagttgc tgagcacggc ccggcttcgg gtgcggggct ccgtacgggg
     3781 cgtggcgcgg ggctcgccgt gccgggcggg gggtggcggc aggtgggggt gccgggcggg
     3841 gcggggccgc ctcgggccgg ggagggctcg ggggaggggc gcggcggccc ccggagcgcc
     3901 ggcggctgtc gaggcgcggc gagccgcagc cattgccttt tatggtaatc gtgcgagagg
     3961 gcgcagggac ttcctttgtc ccaaatctgt gcggagccga aatctgggag gcgccgccgc
     4021 accccctcta gcgggcgcgg ggcgaagcgg tgcggcgccg gcaggaagga aatgggcggg
     4081 gagggccttc gtgcgtcgcc gcgccgccgt ccccttctcc ctctccagcc tcggggctgt
     4141 ccgcgggggg acggctgcct tcggggggga cggggcaggg cggggttcgg cttctggcgt
     4201 gtgaccggcg gctctagagc ctctgctaac catgttcatg ccttcttctt tttcctacag
     4261 ctcctgggca acgtgctggt tattgtggcc accatgaccg agtacaagcc cacggtgcgc
     4321 ctcgccaccc gcgacgacgt ccccagggcc gtacgcaccc tcgccgccgc gttcgccgac
     4381 taccccgcca cgcgccacac cgtcgatccg gaccgccaca tcgagcgggt caccgagctg
     4441 caagaactct tcctcacgcg cgtcgggctc gacatcggca aggtgtgggt cgcggacgac
     4501 ggcgccgcgg tggcggtctg gaccacgccg gagagcgtcg aagcgggggc ggtgttcgcc
     4561 gagatcggcc cgcgcatggc cgagttgagc ggttcccggc tggccgcgca gcaacagatg
     4621 gaaggcctcc tggcgccgca ccggcccaag gagcccgcgt ggttcctggc caccgtcgga
     4681 gtctcgcccg accaccaggg caagggtctg ggcagcgccg tcgtgctccc cggagtggag
     4741 gcggccgagc gcgccggggt gcccgccttc ctggagacct ccgcgccccg caacctcccc
     4801 ttctacgagc ggctcggctt caccgtcacc gccgacgtcg aggtgcccga aggaccgcgc
     4861 acctggtgca tgacccgcaa gcccggtgcc tgaagcgtct ggaacaatca acctctggat
     4921 tacaaaattt gtgaaagatt gactggtatt cttaactatg ttgctccttt tacgctatgt
     4981 ggatacgctg ctttaatgcc tttgtatcat gctattgctt cccgtatggc tttcattttc
     5041 tcctccttgt ataaatcctg gttgctgtct ctttatgagg agttgtggcc cgttgtcagg
     5101 caacgtggcg tggtgtgcac tgtgtttgct gacgcaaccc ccactggttg gggcattgcc
     5161 accacctgtc agctcctttc cgggactttc gctttccccc tccctattgc cacggcggaa
     5221 ctcatcgccg cctgccttgc ccgctgctgg acaggggctc ggctgttggg cactgacaat
     5281 tccgtggtgt tgtcggggaa gctgacgtcc tttccatggc tgctcgcctg tgttgccacc
     5341 tggattctgc gcgggacgtc cttctgctac gtcccttcgg ccctcaatcc agcggacctt
     5401 ccttcccgcg gcctgctgcc ggctctgcgg cctcttccgc gtcttcgcct tcgccctcag
     5461 acgagtcgga tctccctttg ggccgcctcc ccgcctggaa ttaattctgc agtcgagacc
     5521 tagaaaaaca tggagcaatc acaagtagca atacagcagc taccaatgct gattgtgcct
     5581 ggctagaagc acaagaggag gaggaggtgg gttttccagt cacacctcag gtacctttaa
     5641 gaccaatgac ttacaaggca gctgtagatc ttagccactt tttaaaagaa aagaggggac
     5701 tggaagggct aattcactcc caacgaagac aagatatcct tgatctgtgg atctaccaca
     5761 cacaaggcta cttccctgat tagcagaact acacaccagg gccaggggtc agatatccac
     5821 tgacctttgg atggtgctac aagctagtac cagttgagcc agataaggta gaagaggcca
     5881 ataaaggaga gaacaccagc ttgttacacc ctgtgagcct gcatgggatg gatgacccgg
     5941 agagagaagt gttagagtgg aggtttgaca gccgcctagc atttcatcac gtggcccgag
     6001 agctgcatcc ggagtacttc aagaactgct gatatcgagc ttgctacaag ggactttccg
     6061 ctggggactt tccagggagg cgtggcctgg gcgggactgg ggagtggcga gccctcagat
     6121 cctgcatata agcagctgct ttttgcctgt actgggtctc tctggttaga ccagatctga
     6181 gcctgggagc tctctggcta actagggaac ccactgctta agcctcaata aagcttgcct
     6241 tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta gagatccctc
     6301 agaccctttt agtcagtgtg gaaaatctct agcagtagta gttcatgtca tcttattatt
     6361 cagtatttat aacttgcaaa gaaatgaata tcagagagtg agaggccttg acattgctag
     6421 cgttttaccg tcgacctcta gctagagctt ggcgtaatca tggtcatagc tgtttcctgt
     6481 gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa
     6541 agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc
     6601 tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag
     6661 aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt
     6721 cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga
     6781 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg
     6841 taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa
     6901 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt
     6961 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct
     7021 gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct
     7081 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc
     7141 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt
     7201 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc
     7261 tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat
     7321 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa
     7381 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa
     7441 aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga
     7501 aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct
     7561 tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga
     7621 cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc
     7681 catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg
     7741 ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat
     7801 aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat
     7861 ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg
     7921 caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc
     7981 attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa
     8041 agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc
     8101 actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt
     8161 ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag
     8221 ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt
     8281 gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag
     8341 atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac
     8401 cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc
     8461 gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca
     8521 gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg
     8581 ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc gacggatcgg gagatcaact
     8641 tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata
     8701 aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc
     8761 atgtctggat caactggata actcaagcta accaaaatca tcccaaactt cccaccccat
     8821 accctattac cactgccaat tacctgtggt ttcatttact ctaaacctgt gattcctctg
     8881 aattattttc attttaaaga aattgtattt gttaaatatg tactacaaac ttagtagt
//
