LOCUS Exported 7659 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION sgRNA for mammalian cells with mCherry marker. ACCESSION . VERSION . KEYWORDS pJZC102 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 May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7659 /organism="synthetic DNA construct" /mol_type="other DNA" 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 2825..2858 /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." 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 4459..4492 /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." 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(6666..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 //