LOCUS Exported 2138 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION T7 promoter crRNA entry vector used for in vitro transcription of LbCas12a guides. ACCESSION . VERSION . KEYWORDS pT7-G-LbCas12a crRNA-BsaI cassette (MSP3495) SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 2138) AUTHORS Kleinstiver BP, Sousa AA, Walton RT, Tak YE, Hsu JY, Clement K, Welch MM, Horng JE, Malagon-Lopez J, Scarfo I, Maus MV, Pinello L, Aryee MJ, Joung JK TITLE Engineered CRISPR-Cas12a variants with increased activities and improved targeting ranges for gene, epigenetic and base editing. JOURNAL Nat Biotechnol. 2019 Feb 11. pii: 10.1038/s41587-018-0011-0. doi: 10.1038/s41587-018-0011-0. PUBMED 30742127 REFERENCE 2 (bases 1 to 2138) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..2138 /organism="synthetic DNA construct" /mol_type="other DNA" rep_origin 36..623 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 524..543 /label=pBR322ori-F /note="pBR322 origin, forward primer" terminator 635..662 /label=T7Te terminator /note="phage T7 early transcription terminator" CDS complement(686..1501) /codon_start=1 /gene="aph(3')-Ia" /product="aminoglycoside phosphotransferase" /label=KanR /note="confers resistance to kanamycin in bacteria or G418 (Geneticin(R)) in eukaryotes" /translation="MSHIQRETSCLKPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNGDRVFRLAQAQSRMNNGLVGA SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF" primer_bind 1409..1428 /label=Kan-R /note="Kanamycin resistance gene, reverse primer" promoter complement(1502..1604) /label=cat promoter /note="promoter of the E. coli cat gene encoding chloramphenicol acetyltransferase" terminator 1605..1636 /label=tonB terminator /note="bidirectional E. coli tonB-P14 transcription terminator" primer_bind 1684..1700 /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind 1684..1700 /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" promoter 1829..1846 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" primer_bind complement(2117..2134) /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind complement(2117..2133) /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" ORIGIN 1 tgggcctttc ttcggtagaa aatcaaagga tcttcttgag atcctttttt tctgcgcgta 61 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa 121 gagctaccaa ctctttttcc gaggtaactg gcttcagcag agcgcagata ccaaatactg 181 ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat 241 acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta 301 ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg 361 gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc 421 gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa 481 gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc 541 tttatagtcc tgtcgggttt cgccacctct gacttgagca tcgatttttg tgatgctcgt 601 caggggggcg gagcctatgg aaaaacgcca gcaacgcaga aaggcccacc cgaaggtgag 661 ccaggtgatt acatttaggt cctcattaga aaaactcatc gagcatcaag tgaaactgca 721 atttattcat atcaggatta tcaataccat atttttgaaa aagccgtttc tgtaatgaag 781 gagaaaactc accgaggcag ttccatagga tggcaagatc ctggtatcgg tctgcgattc 841 cgactcgtcc aacatcaata caacctatta atttcccctc gtcaaaaata aggttatcaa 901 gtgagaaatc accatgagtg acgactgaat ccggtgagaa tggcaagagt ttatgcattt 961 ctttccagac ttgttcaaca ggccagccat tacgctcgtc atcaaaatca ctcgcaccaa 1021 ccaaaccgtt attcattcgt gattgcgcct gagcgagacg aaatacgcga tcgccgttaa 1081 aaggacaatt acaaacagga atcgaatgca accggcgcag gaacactgcc agcgcatcaa 1141 caatattttc acctgaatca ggatattctt ctaatacctg gaatgctgtt ttccctggga 1201 tcgcagtggt gagtaaccat gcatcatcag gagtacggat aaaatgcttg atggtcggaa 1261 gaggcataaa ttccgtcagc cagtttagcc tgaccatctc atctgtaaca tcattggcaa 1321 cgctaccttt gccatgtttc agaaacaact ctggcgcatc gggcttccca tacaatcgat 1381 agattgtcgc acctgattgc ccgacattat cgcgagccca tttataccca tataaatcag 1441 catccatgtt ggaatttaat cgcggcttca agcaagacgt ttcccgttga atatggctca 1501 ttttagcttc cttagctcct gaaaatctcg ataactcaaa aaatacgccc ggtagtgatc 1561 ttatttcatt atggtgaaag ttggaacctc ttacgtgccg atcaagtcaa aagcctccgg 1621 tcggaggctt ttgactttct gctatggagg tcaggtatga tttaaatggt cagtattgag 1681 cctcaggaaa cagctatgac atcaagctga ctagataatc tagctgatcg tggaccgatc 1741 atacgtataa tgccgtaaga tcacgggtcg cagcacagct cgcggtccag tagtgatcga 1801 cactgctcga tccgctcgca ccgctagcta atacgactca ctatagaatt tctactaagt 1861 gtagatggag acccatgcca tagcgttgtt cggaatatga atttttgaac agattcacca 1921 acacctagtg gtctccttta aaaagcttgg atcgacgaga gcagcgcgac tggatctgtc 1981 gcccgtctca aacgcaaccc tccggcggtc gcatatcatt caggacgagc ctcagactcc 2041 agcgtaactg gactgcaatc aactcactgg ctcaccttcc ggtccacgat cagctagaat 2101 caagctgact agataaactg gccgtcgttt tacacggg //