LOCUS Exported 4381 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Bacterial SP and NM repression YFP reporter: protospacer B. ACCESSION . VERSION . KEYWORDS SK-YFP-SPNM-B SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 4381) AUTHORS Esvelt KM, Mali P, Braff JL, Moosburner M, Yaung SJ, Church GM TITLE Orthogonal Cas9 proteins for RNA-guided gene regulation and editing. JOURNAL Nat Methods. 2013 Sep 29. doi: 10.1038/nmeth.2681. PUBMED 24076762 REFERENCE 2 (bases 1 to 4381) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..4381 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind complement(165..193) /label=GFP-R /note="GFP, reverse primer. Does NOT anneal to EGFP" primer_bind 788..808 /label=GFP-F /note="GFP, forward primer. Does NOT anneal to EGFP" terminator 910..996 /gene="Escherichia coli rrnB" /label=rrnB T1 terminator /note="transcription terminator T1 from the E. coli rrnB gene" CDS complement(1531..2481) /codon_start=1 /gene="rep101" /product="RepA protein needed for replication with the pSC101 origin" /label=Rep101 /translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFS SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLRNGLRKTLHDALTAKIQLTSFEAKF LSDMQSKHDLNGSFSWLTQKQRTTLENILAKYGRI" rep_origin 2529..2751 /label=pSC101 ori /note="low-copy replication origin that requires the Rep101 protein" terminator 3243..3337 /label=lambda t0 terminator /note="transcription terminator from phage lambda" CDS complement(3368..4162) /codon_start=1 /gene="aph(3')-II (or nptII)" /product="aminoglycoside phosphotransferase from Tn5" /label=NeoR/KanR /note="confers resistance to neomycin, kanamycin, and G418 (Geneticin(R))" /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF" primer_bind complement(3479..3498) /label=Neo-F /note="Neomycin resistance gene, forward primer" primer_bind 4089..4108 /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 4341..4359 /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" ORIGIN 1 gatccgataa atatctaaca ccgtgcgtgt tgactatttt acctctggcg gtgataatgg 61 ttgcatgtac tagaattctt taactttaag aaggagatat acatatgaat cccctatggc 121 gaatactttt aaagtctcgt aaaggagaag aacttttcac tggagttgtc ccaattcttg 181 ttgaattaga tggtgatgtt aatgggcaca aattttctgt cagtggagag ggtgaaggtg 241 atgcaacata cggaaaactt acccttaaat ttatttgcac tactggaaaa ctacctgttc 301 catggccaac acttgtcact actttcggtt atggtctaaa atgctttgct agatacccag 361 atcatatgaa acggcatgac tttttcaaga gtgccatgcc cgaaggttat gtacaggaaa 421 gaactatatt tttcaaagat gacgggaact acaagacacg tgctgaagtc aagtttgaag 481 gtgataccct tgttaataga atcgagttaa aaggtattga ttttaaagaa gatggaaaca 541 ttcttggaca caaattggaa tacaactata actcacacaa tgtatacatc atggcagaca 601 aacaaaagaa tggaatcaaa gttaacttca aaattagaca caacattgaa gatggaagcg 661 ttcaactagc agaccattat caacaaaata ctccaattgg cgatggccct gtccttttac 721 cagacaacca ttacctgtcc tatcaatctg ccctttcgaa agatcccaac gaaaagagag 781 accacatggt ccttcttgag tttgtaacag ctgctgggat tacacatggc atggatgaac 841 tatacaaata agcttaaccg aagcgtttga tagttgatat cctttgcctg cggccgcaac 901 tagaggcatc aaataaaacg aaaggctcag tcgaaagact gggcctttcg ttttatctgt 961 tgtttgtcgg tgaacgctct cctgagtagg acaaatccgc cgccctagac ctagggtacg 1021 ggttttgctg cccgcaaacg ggctgttctg gtgttgctag tttgttatca gaatcgcaga 1081 tccggcttca ggtttgccgg ctgaaagcgc tatttcttcc agaattgcca tgattttttc 1141 cccacgggag gcgtcactgg ctcccgtgtt gtcggcagct ttgattcgat aagcagcatc 1201 gcctgtttca ggctgtctat gtgtgactgt tgagctgtaa caagttgtct caggtgttca 1261 atttcatgtt ctagttgctt tgttttactg gtttcacctg ttctattagg tgttacatgc 1321 tgttcatctg ttacattgtc gatctgttca tggtgaacag ctttaaatgc accaaaaact 1381 cgtaaaagct ctgatgtatc tatctttttt acaccgtttt catctgtgca tatggacagt 1441 tttccctttg atatctaacg gtgaacagtt gttctacttt tgtttgttag tcttgatgct 1501 tcactgatag atacaagagc cataagaacc tcagatcctt ccgtatttag ccagtatgtt 1561 ctctagtgtg gttcgttgtt tttgcgtgag ccatgagaac gaaccattga gatcatgctt 1621 actttgcatg tcactcaaaa attttgcctc aaaactggtg agctgaattt ttgcagttaa 1681 agcatcgtgt agtgtttttc ttagtccgtt acgtaggtag gaatctgatg taatggttgt 1741 tggtattttg tcaccattca tttttatctg gttgttctca agttcggtta cgagatccat 1801 ttgtctatct agttcaactt ggaaaatcaa cgtatcagtc gggcggcctc gcttatcaac 1861 caccaatttc atattgctgt aagtgtttaa atctttactt attggtttca aaacccattg 1921 gttaagcctt ttaaactcat ggtagttatt ttcaagcatt aacatgaact taaattcatc 1981 aaggctaatc tctatatttg ccttgtgagt tttcttttgt gttagttctt ttaataacca 2041 ctcataaatc ctcatagagt atttgttttc aaaagactta acatgttcca gattatattt 2101 tatgaatttt tttaactgga aaagataagg caatatctct tcactaaaaa ctaattctaa 2161 tttttcgctt gagaacttgg catagtttgt ccactggaaa atctcaaagc ctttaaccaa 2221 aggattcctg atttccacag ttctcgtcat cagctctctg gttgctttag ctaatacacc 2281 ataagcattt tccctactga tgttcatcat ctgagcgtat tggttataag tgaacgatac 2341 cgtccgttct ttccttgtag ggttttcaat cgtggggttg agtagtgcca cacagcataa 2401 aattagcttg gtttcatgct ccgttaagtc atagcgacta atcgctagtt catttgcttt 2461 gaaaacaact aattcagaca tacatctcaa ttggtctagg tgattttaat cactatacca 2521 attgagatgg gctagtcaat gataattact agtccttttc ctttgagttg tgggtatctg 2581 taaattctgc tagacctttg ctggaaaact tgtaaattct gctagaccct ctgtaaattc 2641 cgctagacct ttgtgtgttt tttttgttta tattcaagtg gttataattt atagaataaa 2701 gaaagaataa aaaaagataa aaagaataga tcccagccct gtgtataact cactacttta 2761 gtcagttccg cagtattaca aaaggatgtc gcaaacgctg tttgctcctc tacaaaacag 2821 accttaaaac cctaaaggct taagtagcac cctcgcaagc tcgggcaaat cgctgaatat 2881 tccttttgtc tccgaccatc aggcacctga gtcgctgtct ttttcgtgac attcagttcg 2941 ctgcgctcac ggctctggca gtgaatgggg gtaaatggca ctacaggcgc cttttatgga 3001 ttcatgcaag gaaactaccc ataatacaag aaaagcccgt cacgggcttc tcagggcgtt 3061 ttatggcggg tctgctatgt ggtgctatct gactttttgc tgttcagcag ttcctgccct 3121 ctgattttcc agtctgacca cttcggatta tcccgtgaca ggtcattcag actggctaat 3181 gcacccagta aggcagcggt atcatcaaca ggcttacccg tcttactgtc cctagtgctt 3241 ggattctcac caataaaaaa cgcccggcgg caaccgagcg ttctgaacaa atccagatgg 3301 agttctgagg tcattactgg atctatcaac aggagtccaa gcgagctctc gaaccccaga 3361 gtcccgctca gaagaactcg tcaagaaggc gatagaaggc gatgcgctgc gaatcgggag 3421 cggcgatacc gtaaagcacg aggaagcggt cagcccattc gccgccaagc tcttcagcaa 3481 tatcacgggt agccaacgct atgtcctgat agcggtccgc cacacccagc cggccacagt 3541 cgatgaatcc agaaaagcgg ccattttcca ccatgatatt cggcaagcag gcatcgccat 3601 gggtcacgac gagatcctcg ccgtcgggca tgcgcgcctt gagcctggcg aacagttcgg 3661 ctggcgcgag cccctgatgc tcttcgtcca gatcatcctg atcgacaaga ccggcttcca 3721 tccgagtacg tgctcgctcg atgcgatgtt tcgcttggtg gtcgaatggg caggtagccg 3781 gatcaagcgt atgcagccgc cgcattgcat cagccatgat ggatactttc tcggcaggag 3841 caaggtgaga tgacaggaga tcctgccccg gcacttcgcc caatagcagc cagtcccttc 3901 ccgcttcagt gacaacgtcg agcacagctg cgcaaggaac gcccgtcgtg gccagccacg 3961 atagccgcgc tgcctcgtcc tgcagttcat tcagggcacc ggacaggtcg gtcttgacaa 4021 aaagaaccgg gcgcccctgc gctgacagcc ggaacacggc ggcatcagag cagccgattg 4081 tctgttgtgc ccagtcatag ccgaatagcc tctccaccca agcggccgga gaacctgcgt 4141 gcaatccatc ttgttcaatc atgcgaaacg atcctcatcc tgtctcttga tcagatcttg 4201 atcccctgcg ccatcagatc cttggcggca agaaagccat ccagtttact ttgcagggct 4261 tcccaacctt accagagggc gccccagctg gcaattccga cgtctaagaa accattatta 4321 tcatgacatt aacctataaa aataggcgta tcacgaggcc ctttcgtctt cacctcgagg 4381 g //