LOCUS Exported 3217 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION tRNA and scaffold for the assembly of GBoligomers for position [2-3] of a polycistronic tRNA-gRNA. ACCESSION . VERSION . KEYWORDS pVD1 tRNA-gRNA position E2 (GB2239) SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 3217) TITLE Orzaez lab plasmids REFERENCE 2 (bases 1 to 3217) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..3217 /organism="synthetic DNA construct" /mol_type="other DNA" rep_origin complement(121..709) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(201..220) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(880..1740) /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 1503..1522 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(1741..1845) /gene="bla" /label=AmpR promoter rep_origin complement(1923..2378) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(2060..2081) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 2272..2291 /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 2504..2526 /label=M13/pUC Forward /note="In lacZ gene" primer_bind 2518..2535 /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind 2519..2535 /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind 2542..2561 /label=T7 /note="T7 promoter, forward primer" promoter 2542..2560 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" regulatory 2591..2600 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" regulatory 2595..2604 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" misc_RNA complement(2631..2705) /label=gRNA scaffold /note="guide RNA scaffold for the Streptococcus pyogenes CRISPR/Cas9 system" promoter complement(2900..2918) /label=SP6 promoter /note="promoter for bacteriophage SP6 RNA polymerase" primer_bind complement(2901..2918) /label=SP6 /note="SP6 promoter, forward primer" primer_bind complement(2936..2952) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(2936..2952) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(2949..2971) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 2960..2976 /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(2984..3014) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 3029..3050 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(3167..3184) /label=L4440 /note="L4440 vector, forward primer" ORIGIN 1 agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa 61 catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt 121 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg 181 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg 241 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag 301 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc 361 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa 421 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg 481 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc 541 taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac 601 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg 661 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt 721 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt 781 catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa 841 atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga 901 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt 961 gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg 1021 agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga 1081 gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga 1141 agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg 1201 catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc 1261 aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc 1321 gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca 1381 taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac 1441 caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg 1501 ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc 1561 ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg 1621 tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac 1681 aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat 1741 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata 1801 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa 1861 agtgccacct gatgcggtgt gaaataccgc acagatgcgt aaggagaaaa taccgcatca 1921 ggaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc 1981 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga 2041 gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc 2101 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc 2161 ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag 2221 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa 2281 agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac 2341 cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcc attcgccatt caggctgcgc 2401 aactgttggg aagggcgatc ggtgcgggcc tcttcgctat tacgccagct ggcgaaaggg 2461 ggatgtgctg caaggcgatt aagttgggta acgccagggt tttcccagtc acgacgttgt 2521 aaaacgacgg ccagtgaatt gtaatacgac tcactatagg gcgaattggg cccgacgtcg 2581 catgctcccg gccgccatgg cggccgcggg aattcgattc ccgtctcgct caccgactcg 2641 gtgccacttt ttcaagttga taacggacta gccttatttt aacttgctat ttctagctct 2701 aaaactgaga cggcgcgcgc cgtctctgca ccagccggga atcgaacccg ggtctgtacc 2761 gtggcagggt actattctac cactagacca ctggtgcttt gttgcaccga gcgagacggg 2821 atcactagtg aattcgcggc cgcctgcagg tcgaccatat gggagagctc ccaacgcgtt 2881 ggatgcatag cttgagtatt ctatagtgtc acctaaatag cttggcgtaa tcatggtcat 2941 agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa 3001 gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc 3061 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc 3121 aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact 3181 cgctgcgctc ggtcgttcgg ctgcggcgag cggtatc //