T1-araC-Pbad-D10A/H840A cas9 ABCD-pACYC-BB

110547 Synthetic Bacterial Expression

Plasmid Information

Purpose
CRISPR interference; works in conjunction with an sgRNA expression vector to silence genes in the E. coli chromosome.
Depositor
Ichiro Matsumura
Insert
dCas9
Mutation
D10A, H840A
Promoter
pBAD
Available Since
July 24, 2018
Availability
Available

Sequence Information

Length
7,974 bp
Topology
circular
Molecule Type
other DNA
View on AddGene.org

Features

CDS 4
Label Gene Product Position Strand
araC araC L-arabinose regulatory protein 186..1065 Rev
DNA Sequence
ATGGCTGAAGCGCAAAATGATCCCCTGCTGCCGGGATACTCGTTTAATGCCCATCTGGTGGCGGGTTTAACGCCGATTGAGGCCAACGGTTATCTCGATTTTTTTATCGACCGACCGCTGGGAATGAAAGGTTATATTCTCAATCTCACCATTCGCGGTCAGGGGGTGGTGAAAAATCAGGGACGAGAATTTGTTTGCCGACCGGGTGATATTTTGCTGTTCCCGCCAGGAGAGATTCATCACTACGGTCGTCATCCGGAGGCTCGCGAATGGTATCACCAGTGGGTTTACTTTCGTCCGCGCGCCTACTGGCATGAATGGCTTAACTGGCCGTCAATATTTGCCAATACGGGGTTCTTTCGCCCGGATGAAGCGCACCAGCCGCATTTCAGCGACCTGTTTGGGCAAATCATTAACGCCGGGCAAGGGGAAGGGCGCTATTCGGAGCTGCTGGCGATAAATCTGCTTGAGCAATTGTTACTGCGGCGCATGGAAGCGATTAACGAGTCGCTCCATCCACCGATGGATAATCGGGTACGCGAGGCTTGTCAGTACATCAGCGATCACCTGGCAGACAGCAATTTTGATATCGCCAGCGTCGCACAGCATGTTTGCTTGTCGCCGTCGCGTCTGTCACATCTTTTCCGCCAGCAGTTAGGGATTAGCGTCTTAAGCTGGCGCGAGGACCAACGTATCAGCCAGGCGAAGCTGCTTTTGAGCACCACCCGGATGCCTATCGCCACCGTCGGTCGCAATGTTGGTTTTGACGATCAACTCTATTTCTCGCGGGTATTTAAAAAATGCACCGGGGCCAGCCCGAGCGAGTTCCGTGCCGGTTGTGAAGAAAAAGTGAATGATGTAGCCGTCAAGTTGTCATAA
Protein Sequence
MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGYILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYWHEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRRMEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGISVLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCEEKVNDVAVKLS
dCas9 — catalytically dead mutant of the Cas9 e… 1424..5528 Fwd
DNA Sequence
ATGGACAAAAAATATAGCATCGGCCTCGCGATTGGTACTAACTCCGTCGGTTGGGCGGTGATTACTGATGAATATAAAGTGCCGAGCAAGAAATTTAAGGTGCTCGGCAACACCGACCGCCACAGCATTAAGAAAAATCTGATTGGCGCGTTGCTTTTTGATTCTGGTGAAACAGCTGAAGCGACGCGTCTGAAGCGTACCGCGCGCCGTCGCTATACCCGCCGCAAAAACCGCATTTGCTATTTGCAGGAGATATTTAGCAATGAGATGGCGAAAGTCGATGATTCGTTTTTCCATCGCCTGGAAGAGAGCTTTCTGGTTGAAGAAGACAAAAAACACGAACGCCATCCGATTTTTGGCAATATTGTCGATGAAGTTGCTTACCATGAAAAGTACCCGACCATTTATCACTTACGCAAGAAATTAGTAGACTCCACCGATAAAGCTGATTTACGTCTCATCTATCTCGCCCTGGCGCACATGATTAAATTCCGCGGTCATTTTCTCATTGAAGGCGACCTCAATCCTGATAACAGTGATGTGGACAAGCTGTTCATCCAGCTCGTGCAAACCTATAATCAGCTGTTTGAAGAAAACCCGATCAATGCCAGCGGCGTGGATGCAAAAGCGATCTTAAGCGCCCGTTTGAGTAAATCCCGCCGGCTGGAAAATCTTATTGCTCAGTTACCGGGCGAGAAGAAGAACGGGTTGTTTGGTAACCTGATTGCGCTGTCACTCGGTCTGACGCCAAACTTCAAAAGTAATTTCGACCTGGCAGAAGATGCTAAACTGCAACTTAGCAAAGATACCTACGATGACGACCTGGATAATTTATTGGCACAGATCGGTGACCAGTATGCGGATCTCTTCCTGGCGGCAAAAAATCTCTCAGATGCGATTTTACTCTCTGATATTCTGCGCGTGAATACGGAAATCACAAAAGCGCCGTTGAGCGCAAGCATGATCAAACGTTATGACGAACATCACCAGGATTTGACTCTGTTAAAAGCGCTGGTGCGTCAGCAACTTCCGGAAAAATACAAAGAGATTTTCTTCGATCAGTCGAAAAACGGTTATGCCGGCTATATTGACGGCGGCGCTTCGCAGGAAGAGTTTTACAAGTTTATTAAACCCATTCTGGAGAAAATGGATGGCACCGAAGAGTTACTGGTTAAACTGAACCGTGAGGACTTGCTGCGTAAACAGCGCACCTTCGATAATGGTTCTATCCCGCATCAGATTCATCTGGGGGAACTGCACGCCATCCTGCGGCGTCAGGAAGATTTCTACCCGTTCCTTAAAGACAACCGCGAGAAAATCGAAAAAATCCTCACGTTTCGCATTCCGTATTATGTCGGCCCGCTGGCGCGTGGTAACAGCCGTTTCGCCTGGATGACCCGAAAAAGCGAAGAAACTATTACGCCGTGGAACTTTGAAGAAGTGGTAGATAAGGGTGCCTCTGCGCAAAGTTTTATCGAGCGCATGACGAACTTTGATAAGAATCTGCCGAACGAAAAGGTACTGCCAAAACACTCGCTGCTCTATGAGTATTTCACCGTTTATAACGAGTTGACGAAAGTTAAATACGTCACTGAAGGTATGCGTAAACCTGCCTTTTTAAGTGGCGAACAAAAAAAGGCCATTGTGGATCTGCTGTTCAAAACAAACCGGAAAGTCACCGTGAAGCAACTGAAAGAGGATTACTTCAAGAAGATTGAATGCTTCGACAGCGTAGAAATTTCCGGCGTTGAAGATCGCTTTAACGCCAGTCTGGGCACTTACCACGATCTGCTGAAGATCATCAAGGATAAGGACTTCCTGGACAATGAAGAGAATGAAGATATACTGGAAGACATCGTGCTGACATTAACGCTCTTTGAGGATCGTGAAATGATTGAAGAGCGTCTGAAAACTTATGCACATTTATTTGATGATAAAGTAATGAAGCAGCTAAAAAGGCGTCGTTACACTGGCTGGGGTCGACTATCGCGCAAGTTGATCAACGGCATTCGTGATAAACAGAGCGGTAAAACGATTCTCGATTTTTTGAAGAGCGACGGTTTTGCCAATCGCAATTTTATGCAGCTTATCCATGATGACAGCTTAACCTTTAAAGAAGATATTCAGAAGGCGCAGGTGTCCGGTCAGGGCGATAGCCTGCATGAGCATATCGCGAACCTGGCGGGTTCGCCGGCAATCAAAAAAGGTATTCTTCAGACGGTAAAAGTGGTTGACGAGCTGGTGAAAGTGATGGGCCGTCATAAACCAGAAAATATCGTTATTGAAATGGCACGGGAAAACCAGACCACCCAGAAAGGCCAGAAAAATAGCCGCGAACGTATGAAACGCATCGAGGAAGGGATCAAGGAACTTGGCAGTCAGATCCTGAAAGAGCACCCGGTGGAGAACACGCAATTACAGAACGAAAAACTCTACCTTTATTACCTGCAAAACGGTCGCGATATGTATGTTGATCAAGAATTAGATATTAATCGTCTTTCTGACTACGACGTTGATGCGATTGTACCGCAATCCTTCCTCAAAGACGATTCCATTGATAACAAAGTGCTGACTCGCAGCGATAAAAACCGTGGCAAGTCCGATAACGTGCCTTCTGAAGAAGTCGTGAAAAAAATGAAAAACTACTGGCGCCAGTTGCTGAATGCGAAACTGATCACTCAGCGTAAATTTGACAACCTCACCAAAGCCGAACGCGGCGGGCTGAGTGAACTGGATAAAGCAGGCTTTATTAAACGCCAGCTGGTGGAAACGCGCCAAATTACCAAGCATGTTGCCCAGATTCTGGACTCGCGAATGAATACCAAATACGATGAAAACGATAAGCTGATCCGTGAAGTGAAAGTGATCACCTTGAAATCGAAACTGGTCAGCGATTTCCGTAAAGACTTCCAGTTCTATAAGGTACGTGAAATTAACAACTATCACCATGCGCATGACGCTTATCTTAATGCAGTGGTGGGTACGGCGTTAATAAAAAAATATCCGAAGCTCGAAAGCGAATTTGTCTACGGCGATTATAAAGTCTATGATGTCCGTAAGATGATCGCCAAATCTGAGCAGGAAATTGGGAAAGCCACCGCCAAGTACTTCTTCTACAGCAACATCATGAACTTCTTTAAAACCGAAATCACGCTAGCTAACGGTGAAATCCGCAAACGTCCGTTAATCGAAACCAACGGCGAAACCGGTGAGATCGTCTGGGATAAAGGGCGCGACTTTGCTACGGTTCGTAAGGTTCTGTCGATGCCGCAGGTTAACATTGTTAAGAAAACCGAGGTGCAGACCGGCGGTTTCAGCAAAGAAAGTATTTTGCCGAAACGTAACTCGGACAAATTGATTGCCCGCAAAAAGGACTGGGACCCGAAAAAATATGGCGGATTTGACAGTCCAACCGTCGCCTACAGCGTGCTGGTAGTGGCGAAGGTCGAAAAAGGCAAATCAAAAAAGCTGAAAAGCGTCAAAGAGCTGCTGGGTATCACCATCATGGAACGTAGCAGCTTCGAAAAAAATCCAATCGACTTTCTTGAAGCCAAGGGCTACAAAGAAGTGAAGAAGGACTTGATCATCAAGCTGCCAAAGTATTCCCTGTTTGAACTGGAGAACGGACGCAAACGGATGCTGGCATCGGCGGGCGAATTGCAAAAAGGTAATGAACTGGCACTGCCGTCTAAATATGTGAACTTCCTGTATCTGGCCAGCCACTATGAAAAACTGAAAGGCAGCCCGGAAGATAACGAACAGAAACAACTGTTTGTTGAGCAGCACAAACATTATCTGGATGAGATCATTGAGCAAATCAGCGAGTTCAGTAAGCGCGTTATCCTTGCCGATGCCAACCTGGATAAAGTGTTGTCTGCCTATAATAAACATCGTGACAAACCGATTCGCGAACAGGCGGAAAACATCATCCACCTGTTTACCCTGACCAATTTGGGCGCACCGGCGGCGTTTAAATACTTTGACACCACTATCGACCGTAAACGCTACACCAGCACCAAAGAAGTACTTGATGCCACGCTGATTCATCAAAGCATAACGGGCCTGTACGAAACCCGTATTGATCTGAGCCAGTTAGGTGGTGAT
Protein Sequence
MDKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKNLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESFLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKFRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQIGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLRKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGNSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYFTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDSVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERLKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRHKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYLYYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPSEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHVAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLNAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTEITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGITIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNELALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLDATLIHQSITGLYETRIDLSQLGGD
CmR cat chloramphenicol acetyltransferase 6915..7575 Fwd
DNA Sequence
ATGGAGAAAAAAATCACTGGATATACCACCGTTGATATATCCCAATGGCATCGTAAAGAACATTTTGAGGCATTTCAGTCAGTTGCTCAATGTACCTATAACCAGACCGTTCAGCTGGATATTACGGCCTTTTTAAAGACCGTAAAGAAAAATAAGCACAAGTTTTATCCGGCCTTTATTCACATTCTTGCCCGCCTGATGAATGCTCATCCGGAGTTCCGTATGGCAATGAAAGACGGTGAGCTGGTGATATGGGATAGTGTTCACCCTTGTTACACCGTTTTCCATGAGCAAACTGAAACGTTTTCATCGCTCTGGAGTGAATACCACGACGATTTCCGGCAGTTTCTACACATATATTCGCAAGATGTGGCGTGTTACGGTGAAAACCTGGCCTATTTCCCTAAAGGGTTTATTGAGAATATGTTTTTCGTCTCAGCCAATCCCTGGGTGAGTTTCACCAGTTTTGATTTAAACGTGGCCAATATGGACAACTTCTTCGCCCCCGTTTTCACTATGGGCAAATATTATACGCAAGGCGACAAGGTGCTGATGCCGCTGGCGATTCAGGTTCATCATGCCGTTTGTGATGGCTTCCATGTCGGCAGAATGCTTAATGAATTACAACAGTACTGCGATGAGTGGCAGGGCGGGGCGTAA
Protein Sequence
MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFLKTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSSLWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANMDNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA
S-Tag — affinity and epitope tag derived from p… 7894..7939 Rev
DNA Sequence
AAAGAAACCGCTGCTGCGAAATTTGAACGCCAGCACATGGACTCG
Protein Sequence
KETAAAKFERQHMDS
promoter 2
Label Gene Product Position Strand
araBAD promoter araBAD — 1091..1376 Fwd
DNA Sequence
AAGAAACCAATTGTCCATATTGCATCAGACATTGCCGTCACTGCGTCTTTTACTGGCTCTTCTCGCTAACCAAACCGGTAACCCCGCTTATTAAAAGCATTCTGTAACAAAGCGGGACCAAAGCCATGACAAAAACGCGTAACAAAAGTGTCTATAATCACGGCAGAAAAGTCCACATTGATTATTTGCACGGCGTCACACTTTGCTATGCCATAGCATTTTTATCCATAAGATTAGCGGATCCTACCTGACGCTTTTTATCGCAACTCTCTACTGTTTCTCCAT
cat promoter — — 6812..6915 Fwd
DNA Sequence
TGATCGGCACGTAAGAGGTTCCAACTTTCACCATAATGAAATAAGATCACTACCGGGCGTATTTTTTGAGTTATCGAGATTTTCAGGAGCTAAGGAAGCTAAA
rep_origin 1
Label Gene Product Position Strand
p15A ori — — 5741..6287 Fwd
DNA Sequence
TTGAGATCGTTTTGGTCTGCGCGTAATCTCTTGCTCTGAAAACGAAAAAACCGCCTTGCAGGGCGGTTTTTCGAAGGTTCTCTGAGCTACCAACTCTTTGAACCGAGGTAACTGGCTTGGAGGAGCGCAGTCACCAAAACTTGTCCTTTCAGTTTAGCCTTAACCGGCGCATGACTTCAAGACTAACTCCTCTAAATCAATTACCAGTGGCTGCTGCCAGTGGTGCTTTTGCATGTCTTTCCGGGTTGGACTCAAGACGATAGTTACCGGATAAGGCGCAGCGGTCGGACTGAACGGGGGGTTCGTGCATACAGTCCAGCTTGGAGCGAACTGCCTACCCGGAACTGAGTGTCAGGCGTGGAATGAGACAAACGCGGCCATAACAGCGGAATGACACCGGTAAACCGAAAGGCAGGAACAGGAGAGCGCACGAGGGAGCCGCCAGGGGGAAACGCCTGGTATCTTTATAGTCCTGTCGGGTTTCGCCACCACTGATTTGAGCGTCAGATTTCGTGATGCTTGTCAGGGGGGCGGAGCCTATGGAAA
terminator 2
Label Gene Product Position Strand
lambda t0 terminator — — 5605..5700 Fwd
DNA Sequence
GACTCCTGTTGATAGATCCAGTAATGACCTCAGAACTCCATCTGGATTTGTTCAGAACGCTCGGTTGCCGCCGGGCGTTTTTTATTGGTGAGAAT
T7 terminator — — 7795..7843 Rev
DNA Sequence
CTAGCATAACCCCTTGGGGCCTCTAAACGGGTCTTGAGGGGTTTTTTG