LOCUS       sequence-102260-        7659 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  sgRNA for mammalian cells with mCherry marker.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7659)
  AUTHORS   Zalatan JG, Lee ME, Almeida R, Gilbert LA, Whitehead EH, La Russa M,
            Tsai JC, Weissman JS, Dueber JE, Qi LS, Lim WA
  TITLE     Engineering Complex Synthetic Transcriptional Programs with CRISPR 
            RNA Scaffolds.
  JOURNAL   Cell. 2014 Dec 18. pii: S0092-8674(14)01570-0. doi: 
            10.1016/j.cell.2014.11.052.
  PUBMED    25533786
REFERENCE   2  (bases 1 to 7659)
  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: "Cell. 2014 
            Dec 18. pii: S0092-8674(14)01570-0. doi: 
            10.1016/j.cell.2014.11.052."
COMMENT     Sequence Label: pJZC102
FEATURES             Location/Qualifiers
     source          1..7659
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(56..75)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        247..626
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        628..826
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     LTR             844..1024
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1071..1196
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1695..1928
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2113..2157
                     /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    2455..2572
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     protein_bind    complement(2825..2858)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     2862..2882
                     /label=mU6-F
                     /note="Mouse U6 promoter, forward primer"
     enhancer        3182..3485
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3486..3689
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     CDS             3732..4439
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     complement(3874..3892)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     4142..4161
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     protein_bind    complement(4459..4492)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    4548..5136
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4601..4621)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5019..5030)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(5139..5155)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5140..5156)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             5665..5845
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(5907..6495)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5987..6006)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6666..7457,7458..7526))
                     /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     7289..7308
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7527..7631)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gcttaagcgg tcgacggatc gggagatctc ccgatcccct atggtgcact ctcagtacaa
       61 tctgctctga tgccgcatag ttaagccagt atctgctccc tgcttgtgtg ttggaggtcg
      121 ctgagtagtg cgcgagcaaa atttaagcta caacaaggca aggcttgacc gacaattgca
      181 tgaagaatct gcttagggtt aggcgttttg cgctgcttcg cgatgtacgg gccagatata
      241 cgcgttgaca ttgattattg actagttatt aatagtaatc aattacgggg tcattagttc
      301 atagcccata tatggagttc cgcgttacat aacttacggt aaatggcccg cctggctgac
      361 cgcccaacga cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa
      421 tagggacttt ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag
      481 tacatcaagt gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc
      541 ccgcctggca ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct
      601 acgtattagt catcgctatt accatggtga tgcggttttg gcagtacatc aatgggcgtg
      661 gatagcggtt tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt
      721 tgttttggca ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga
      781 cgcaaatggg cggtaggcgt gtacggtggg aggtctatat aagcagcgcg ttttgcctgt
      841 actgggtctc tctggttaga ccagatctga gcctgggagc tctctggcta actagggaac
      901 ccactgctta agcctcaata aagcttgcct tgagtgcttc aagtagtgtg tgcccgtctg
      961 ttgtgtgact ctggtaacta gagatccctc agaccctttt agtcagtgtg gaaaatctct
     1021 agcagtggcg cccgaacagg gacttgaaag cgaaagggaa accagaggag ctctctcgac
     1081 gcaggactcg gcttgctgaa gcgcgcacgg caagaggcga ggggcggcga ctggtgagta
     1141 cgccaaaaat tttgactagc ggaggctaga aggagagaga tgggtgcgag agcgtcagta
     1201 ttaagcgggg gagaattaga tcgcgatggg aaaaaattcg gttaaggcca gggggaaaga
     1261 aaaaatataa attaaaacat atagtatggg caagcaggga gctagaacga ttcgcagtta
     1321 atcctggcct gttagaaaca tcagaaggct gtagacaaat actgggacag ctacaaccat
     1381 cccttcagac aggatcagaa gaacttagat cattatataa tacagtagca accctctatt
     1441 gtgtgcatca aaggatagag ataaaagaca ccaaggaagc tttagacaag atagaggaag
     1501 agcaaaacaa aagtaagacc accgcacagc aagcggccgg ccgcgctgat cttcagacct
     1561 ggaggaggag atatgaggga caattggaga agtgaattat ataaatataa agtagtaaaa
     1621 attgaaccat taggagtagc acccaccaag gcaaagagaa gagtggtgca gagagaaaaa
     1681 agagcagtgg gaataggagc tttgttcctt gggttcttgg gagcagcagg aagcactatg
     1741 ggcgcagcgt caatgacgct gacggtacag gccagacaat tattgtctgg tatagtgcag
     1801 cagcagaaca atttgctgag ggctattgag gcgcaacagc atctgttgca actcacagtc
     1861 tggggcatca agcagctcca ggcaagaatc ctggctgtgg aaagatacct aaaggatcaa
     1921 cagctcctgg ggatttgggg ttgctctgga aaactcattt gcaccactgc tgtgccttgg
     1981 aatgctagtt ggagtaataa atctctggaa cagatttgga atcacacgac ctggatggag
     2041 tgggacagag aaattaacaa ttacacaagc ttaatacact ccttaattga agaatcgcaa
     2101 aaccagcaag aaaagaatga acaagaatta ttggaattag ataaatgggc aagtttgtgg
     2161 aattggttta acataacaaa ttggctgtgg tatataaaat tattcataat gatagtagga
     2221 ggcttggtag gtttaagaat agtttttgct gtactttcta tagtgaatag agttaggcag
     2281 ggatattcac cattatcgtt tcagacccac ctcccaaccc cgaggggacc cgacaggccc
     2341 gaaggaatag aagaagaagg tggagagaga gacagagaca gatccattcg attagtgaac
     2401 ggatcggcac tgcgtgcgcc aattctgcag acaaatggca gtattcatcc acaattttaa
     2461 aagaaaaggg gggattgggg ggtacagtgc aggggaaaga atagtagaca taatagcaac
     2521 agacatacaa actaaagaat tacaaaaaca aattacaaaa attcaaaatt ttcgggttta
     2581 ttacagggac agcagagatc cagtttggtt agtaccgggc ccgctctaga gatccgacgc
     2641 gccatctcta ggcccgcgcc ggccccctcg cacagacttg tgggagaagc tcggctactc
     2701 ccctgccccg gttaatttgc atataatatt tcctagtaac tatagaggct taatgtgcga
     2761 taaaagacag ataatctgtt ctttttaata ctagctacat tttacatgat aggcttggat
     2821 ttctataact tcgtatagca tacattatac gaagttataa acagcacaaa aggaaactca
     2881 ccctaactgt aaagtaattg tgtgttttga gactataagt atcccttgga gaaccacctt
     2941 gttggcatac ttctgcctgc tggggagcct ggtttaagag ctatgctgga aacagcatag
     3001 caagtttaaa taaggctagt ccgttatcaa cttgaaaaag tggcaccgag tcggtgcttt
     3061 ttttctcgag tactaggatc cattaggcgg ccgcgtggat aaccgtatta ccgccatgca
     3121 ttagttatta atagtaatca attacggggt cattagttca tagcccatat atggagttcc
     3181 gcgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac ccccgcccat
     3241 tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc cattgacgtc
     3301 aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg tatcatatgc
     3361 caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt
     3421 acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc atcgctatta
     3481 ccatggtgat gcggttttgg cagtacatca atgggcgtgg atagcggttt gactcacggg
     3541 gatttccaag tctccacccc attgacgtca atgggagttt gttttggcac caaaatcaac
     3601 gggactttcc aaaatgtcgt aacaactccg ccccattgac gcaaatgggc ggtaggcgtg
     3661 tacggtggga ggtctatata agcagagctg gtttagtgaa ccgtcagatc cgctagccca
     3721 ccatgcccgg ggtgagcaag ggcgaggagg ataacatggc catcatcaag gagttcatgc
     3781 gcttcaaggt gcacatggag ggctccgtga acggccacga gttcgagatc gagggcgagg
     3841 gcgagggccg cccctacgag ggcacccaga ccgccaagct gaaggtgacc aagggtggcc
     3901 ccctgccctt cgcctgggac atcctgtccc ctcagttcat gtacggctcc aaggcctacg
     3961 tgaagcaccc cgccgacatc cccgactact tgaagctgtc cttccccgag ggcttcaagt
     4021 gggagcgcgt gatgaacttc gaggacggcg gcgtggtgac cgtgacccag gactcctccc
     4081 tgcaggacgg cgagttcatc tacaaggtga agctgcgcgg caccaacttc ccctccgacg
     4141 gccccgtaat gcagaagaag accatgggct gggaggcctc ctccgagcgg atgtaccccg
     4201 aggacggcgc cctgaagggc gagatcaagc agaggctgaa gctgaaggac ggcggccact
     4261 acgacgctga ggtcaagacc acctacaagg ccaagaagcc cgtgcagctg cccggcgcct
     4321 acaacgtcaa catcaagttg gacatcacct cccacaacga ggactacacc atcgtggaac
     4381 agtacgaacg cgccgagggc cgccactcca ccggcggcat ggacgagctg tacaagtaag
     4441 aattcgtcga gggacctaat aacttcgtat agcatacatt atacgaagtt atacatgttt
     4501 aagggttccg gttccactag gtacaattcg atatcaagct tatcgataat caacctctgg
     4561 attacaaaat ttgtgaaaga ttgactggta ttcttaacta tgttgctcct tttacgctat
     4621 gtggatacgc tgctttaatg cctttgtatc atgctattgc ttcccgtatg gctttcattt
     4681 tctcctcctt gtataaatcc tggttgctgt ctctttatga ggagttgtgg cccgttgtca
     4741 ggcaacgtgg cgtggtgtgc actgtgtttg ctgacgcaac ccccactggt tggggcattg
     4801 ccaccacctg tcagctcctt tccgggactt tcgctttccc cctccctatt gccacggcgg
     4861 aactcatcgc cgcctgcctt gcccgctgct ggacaggggc tcggctgttg ggcactgaca
     4921 attccgtggt gttgtcgggg aaatcatcgt cctttccttg gctgctcgcc tgtgttgcca
     4981 cctggattct gcgcgggacg tccttctgct acgtcccttc ggccctcaat ccagcggacc
     5041 ttccttcccg cggcctgctg ccggctctgc ggcctcttcc gcgtcttcgc cttcgccctc
     5101 agacgagtcg gatctccctt tgggccgcct ccccgcatcg ataccgtcga cctcgatcga
     5161 gacctagaaa aacatggagc aatcacaagt agcaatacag cagctaccaa tgctgattgt
     5221 gcctggctag aagcacaaga ggaggaggag gtgggttttc cagtcacacc tcaggtacct
     5281 ttaagaccaa tgacttacaa ggcagctgta gatcttagcc actttttaaa agaaaagggg
     5341 ggactggaag ggctaattca ctcccaacga agacaagata tccttgatct gtggatctac
     5401 cacacacaag gctacttccc tgattggcag aactacacac cagggccagg gatcagatat
     5461 ccactgacct ttggatggtg ctacaagcta gtaccagttg agcaagagaa ggtagaagaa
     5521 gccaatgaag gagagaacac ccgcttgtta caccctgtga gcctgcatgg gatggatgac
     5581 ccggagagag aagtattaga gtggaggttt gacagccgcc tagcatttca tcacatggcc
     5641 cgagagctgc atccggactg tactgggtct ctctggttag accagatctg agcctgggag
     5701 ctctctggct aactagggaa cccactgctt aagcctcaat aaagcttgcc ttgagtgctt
     5761 caagtagtgt gtgcccgtct gttgtgtgac tctggtaact agagatccct cagacccttt
     5821 tagtcagtgt ggaaaatctc tagcagcatg tgagcaaaag gccagcaaaa ggccaggaac
     5881 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac
     5941 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg
     6001 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac
     6061 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat
     6121 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag
     6181 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac
     6241 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt
     6301 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt
     6361 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc
     6421 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga
     6481 aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac
     6541 gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc
     6601 cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct
     6661 gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca
     6721 tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct
     6781 ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca
     6841 ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc
     6901 atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg
     6961 cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct
     7021 tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa
     7081 aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta
     7141 tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc
     7201 ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg
     7261 agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa
     7321 gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg
     7381 agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc
     7441 accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg
     7501 gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat
     7561 cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata
     7621 ggggttccgc gcacatttcc ccgaaaagtg ccacctgac
//
