| 质粒名称: | pCas9-CR4 |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 62655 |
| 存储实验室: | Kristala Prather |
|
载体骨架基本信息 |
|
| 载体名称: | pdCas9-bacteria |
| 载体出品公司: | -- |
| 载体抗性: | 氯霉素 |
| 空载体大小: | -- |
| 完整质粒大小: | 6770 bp |
| 载体修饰: | -- |
| 载体类型: | 细菌表达 |
| 筛选标记: | -- |
| 高拷贝/低拷贝: | 低拷贝 |
| 生长条件: | 37 ℃ |
|
插入基因信息 |
|
| 插入基因名称: | Cas9 nuclease |
| 物种来源: | -- |
| 插入大小: | -- |
| 基因突变: | -- |
| GenBank编号 | -- |
| 启动子 | -- |
| 标签/融合蛋白: | -- |
| 克隆方法: | -- |
| 5' 克隆位点: | -- |
| 3' 克隆位点: | -- |
| 5' 测序引物 | -- |
| 3' 测序引物 | -- |
|
文献引用方法 |
|
| 材料和方法部分: | pCas9-CR4 was a gift from Kristala Prather (Addgene plasmid # 62655 ; http://n2t.net/addgene:62655 ; RRID:Addgene_62655) |
| 参考文献部分: | The no-SCAR (Scarless Cas9 Assisted Recombineering) system for genome editing in Escherichia coli. Reisch CR, Prather KL. Sci Rep. 2015 Oct 14;5:15096. doi: 10.1038/srep15096. 10.1038/srep15096 PubMed 26463009 |
买家导航
pCas9CR4载体基本信息
pCas9CR4载体图谱质粒图谱和多克隆位点信息
pCas9CR4载体简介
pCas9CR4载体序列
LOCUS pCas9-CR4 6770 bp ds-DNA circular SYN 06-AUG-2018
DEFINITION Cas9 nuclease under control of pTet promoter with ssrA tag and
constitutive tetR.
ACCESSION .
VERSION .
KEYWORDS pCas9-CR4
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 6770)
AUTHORS Reisch CR, Prather KL
TITLE The no-SCAR (Scarless Cas9 Assisted Recombineering) system for
genome editing in Escherichia coli.
JOURNAL Sci Rep. 2015 Oct 14;5:15096. doi: 10.1038/srep15096.
PUBMED 26463009
REFERENCE 2 (bases 1 to 6770)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Aug 6, 2018 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..6770
/organism="synthetic DNA construct"
/mol_type="other DNA"
CDS complement(7..630)
/codon_start=1
/gene="tetR from transposon Tn10"
/product="tetracycline repressor TetR"
/label=TetR
/note="TetR binds to the tetracycline operator tetO to
inhibit transcription. This inhibition can be relieved by
adding tetracycline or doxycycline."
/translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV
KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT
DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
primer_bind 538..557
/label=Tet-R
/note="Tetracycline resistance gene, reverse primer"
promoter 681..736
/gene="tetR"
/label=tetR/tetA promoters
/note="overlapping promoters for bacterial tetR and tetA"
protein_bind 717..735
/gene="tetO"
/label=tet operator
/bound_moiety="tetracycline repressor TetR"
/note="bacterial operator O2 for the tetR and tetA genes"
CDS 772..4875
/codon_start=1
/product="Cas9 (Csn1) endonuclease from the Streptococcus
pyogenes Type II CRISPR/Cas system"
/label=Cas9
/note="generates RNA-guided double strand breaks in DNA"
/translation="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
DATLIHQSITGLYETRIDLSQLGGD"
terminator 4935..5006
/label=rrnB T1 terminator
/note="transcription terminator T1 from the E. coli rrnB
gene"
terminator 5022..5049
/label=T7Te terminator
/note="phage T7 early transcription terminator"
primer_bind complement(5077..5094)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(5211..5756)
/direction=LEFT
/label=p15A ori
/note="Plasmids containing the medium-copy-number p15A
origin of replication can be propagated in E. coli cells
that contain a second plasmid with the ColE1 origin.
"
primer_bind complement(5291..5310)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
terminator 5870..5964
/label=lambda t0 terminator
/note="transcription terminator from phage lambda"
CDS complement(5985..6644)
/codon_start=1
/gene="cat"
/product="chloramphenicol acetyltransferase"
/label=CmR
/note="confers resistance to chloramphenicol"
/translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
primer_bind 6559..6578
/label=CAT-R
/note="Chloramphenicol resistance gene, reverse primer"
promoter complement(6645..6747)
/label=cat promoter
/note="promoter of the E. coli cat gene encoding
chloramphenicol acetyltransferase"
ORIGIN
1 gacgtcttaa gacccacttt cacatttaag ttgtttttct aatccgcata tgatcaattc
61 aaggccgaat aagaaggctg gctctgcacc ttggtgatca aataattcga tagcttgtcg
121 taataatggc ggcatactat cagtagtagg tgtttccctt tcttctttag cgacttgatg
181 ctcttgatct tccaatacgc aacctaaagt aaaatgcccc acagcgctga gtgcatataa
241 tgcattctct agtgaaaaac cttgttggca taaaaaggct aattgatttt cgagagtttc
301 atactgtttt tctgtaggcc gtgtacctaa atgtactttt gctccatcgc gatgacttag
361 taaagcacat ctaaaacttt tagcgttatt acgtaaaaaa tcttgccagc tttccccttc
421 taaagggcaa aagtgagtat ggtgcctatc taacatctca atggctaagg cgtcgagcaa
481 agcccgctta ttttttacat gccaatacaa tgtaggctgc tctacaccta gcttctgggc
541 gagtttacgg gttgttaaac cttcgattcc gacctcatta agcagctcta atgcgctgtt
601 aatcacttta cttttatcta atctagacat cattaattcc taattgctag cattgtacct
661 aggactgagc tagccataaa gttgacactc tatcgttgat agagttattt taccactccc
721 tatcagtgat agagaaaaga attcaaaaga tctaaagagg acttcggatc tatggataag
781 aaatactcaa taggcttaga tatcggcaca aatagcgtcg gatgggcggt gatcactgat
841 gaatataagg ttccgtctaa aaagttcaag gttctgggaa atacagaccg ccacagtatc
901 aaaaaaaatc ttataggggc tcttttattt gacagtggag agacagcgga agcgactcgt
961 ctcaaacgga cagctcgtag aaggtataca cgtcggaaga atcgtatttg ttatctacag
1021 gagatttttt caaatgagat ggcgaaagta gatgatagtt tctttcatcg acttgaagag
1081 tcttttttgg tggaagaaga caagaagcat gaacgtcatc ctatttttgg aaatatagta
1141 gatgaagttg cttatcatga gaaatatcca actatctatc atctgcgaaa aaaattggta
1201 gattctactg ataaagcgga tttgcgctta atctatttgg ccttagcgca tatgattaag
1261 tttcgtggtc attttttgat tgagggagat ttaaatcctg ataatagtga tgtggacaaa
1321 ctatttatcc agttggtaca aacctacaat caattatttg aagaaaaccc tattaacgca
1381 agtggagtag atgctaaagc gattctttct gcacgattga gtaaatcaag acgattagaa
1441 aatctcattg ctcagctccc cggtgagaag aaaaatggct tatttgggaa tctcattgct
1501 ttgtcattgg gtttgacccc taattttaaa tcaaattttg atttggcaga agatgctaaa
1561 ttacagcttt caaaagatac ttacgatgat gatttagata atttattggc gcaaattgga
1621 gatcaatatg ctgatttgtt tttggcagct aagaatttat cagatgctat tttactttca
1681 gatatcctaa gagtaaatac tgaaataact aaggctcccc tatcagcttc aatgattaaa
1741 cgctacgatg aacatcatca agacttgact cttttaaaag ctttagttcg acaacaactt
1801 ccagaaaagt ataaagaaat cttttttgat caatcaaaaa acggatatgc aggttatatt
1861 gatgggggag ctagccaaga agaattttat aaatttatca aaccaatttt agaaaaaatg
1921 gatggtactg aggaattatt ggtgaaacta aatcgtgaag atttgctgcg caagcaacgg
1981 acctttgaca acggctctat tccccatcaa attcacttgg gtgagctgca tgctattttg
2041 agaagacaag aagactttta tccattttta aaagacaatc gtgagaagat tgaaaaaatc
2101 ttgacttttc gaattcctta ttatgttggt ccattggcgc gtggcaatag tcgttttgca
2161 tggatgactc ggaagtctga agaaacaatt accccatgga attttgaaga agttgtcgat
2221 aaaggtgctt cagctcaatc atttattgaa cgcatgacaa actttgataa aaatcttcca
2281 aatgaaaaag tactaccaaa acatagtttg ctttatgagt attttacggt ttataacgaa
2341 ttgacaaagg tcaaatatgt tactgaagga atgcgaaaac cagcatttct ttcaggtgaa
2401 cagaagaaag ccattgttga tttactcttc aaaacaaatc gaaaagtaac cgttaagcaa
2461 ttaaaagaag attatttcaa aaaaatagaa tgttttgata gtgttgaaat ttcaggagtt
2521 gaagatagat ttaatgcttc attaggtacc taccatgatt tgctaaaaat tattaaagat
2581 aaagattttt tggataatga agaaaatgaa gatatcttag aggatattgt tttaacattg
2641 accttatttg aagataggga gatgattgag gaaagactta aaacatatgc tcacctcttt
2701 gatgataagg tgatgaaaca gcttaaacgt cgccgttata ctggttgggg acgtttgtct
2761 cgaaaattga ttaatggtat tagggataag caatctggca aaacaatatt agattttttg
2821 aaatcagatg gttttgccaa tcgcaatttt atgcagctga tccatgatga tagtttgaca
2881 tttaaagaag acattcaaaa agcacaagtg tctggacaag gcgatagttt acatgaacat
2941 attgcaaatt tagctggtag ccctgctatt aaaaaaggta ttttacagac tgtaaaagtt
3001 gttgatgaat tggtcaaagt aatggggcgg cataagccag aaaatatcgt tattgaaatg
3061 gcacgtgaaa atcagacaac tcaaaagggc cagaaaaatt cgcgagagcg tatgaaacga
3121 atcgaagaag gtatcaaaga attaggaagt cagattctta aagagcatcc tgttgaaaat
3181 actcaattgc aaaatgaaaa gctctatctc tattatctcc aaaatggaag agacatgtat
3241 gtggaccaag aattagatat taatcgttta agtgattatg atgtcgatca cattgttcca
3301 caaagtttcc ttaaagacga ttcaatagac aataaggtct taacgcgttc tgataaaaat
3361 cgtggtaaat cggataacgt tccaagtgaa gaagtagtca aaaagatgaa aaactattgg
3421 agacaacttc taaacgccaa gttaatcact caacgtaagt ttgataattt aacgaaagct
3481 gaacgtggag gtttgagtga acttgataaa gctggtttta tcaaacgcca attggttgaa
3541 actcgccaaa tcactaagca tgtggcacaa attttggata gtcgcatgaa tactaaatac
3601 gatgaaaatg ataaacttat tcgagaggtt aaagtgatta ccttaaaatc taaattagtt
3661 tctgacttcc gaaaagattt ccaattctat aaagtacgtg agattaacaa ttaccatcat
3721 gcccatgatg cgtatctaaa tgccgtcgtt ggaactgctt tgattaagaa atatccaaaa
3781 cttgaatcgg agtttgtcta tggtgattat aaagtttatg atgttcgtaa aatgattgct
3841 aagtctgagc aagaaatagg caaagcaacc gcaaaatatt tcttttactc taatatcatg
3901 aacttcttca aaacagaaat tacacttgca aatggagaga ttcgcaaacg ccctctaatc
3961 gaaactaatg gggaaactgg agaaattgtc tgggataaag ggcgagattt tgccacagtg
4021 cgcaaagtat tgtccatgcc ccaagtcaat attgtcaaga aaacagaagt acagacaggc
4081 ggattctcca aggagtcaat tttaccaaaa agaaattcgg acaagcttat tgctcgtaaa
4141 aaagactggg atccaaaaaa atatggtggt tttgatagtc caacggtagc ttattcagtc
4201 ctagtggttg ctaaggtgga aaaagggaaa tcgaagaagt taaaatccgt taaagagtta
4261 ctagggatca caattatgga aagaagttcc tttgaaaaaa atccgattga ctttttagaa
4321 gctaaaggat ataaggaagt taaaaaagac ttaatcatta aactacctaa atatagtctt
4381 tttgagttag aaaacggtcg taaacggatg ctggctagtg ccggagaatt acaaaaagga
4441 aatgagctgg ctctgccaag caaatatgtg aattttttat atttagctag tcattatgaa
4501 aagttgaagg gtagtccaga agataacgaa caaaaacaat tgtttgtgga gcagcataag
4561 cattatttag atgagattat tgagcaaatc agtgaatttt ctaagcgtgt tattttagca
4621 gatgccaatt tagataaagt tcttagtgca tataacaaac atagagacaa accaatacgt
4681 gaacaagcag aaaatattat tcatttattt acgttgacga atcttggagc tcccgctgct
4741 tttaaatatt ttgatacaac aattgatcgt aaacgatata cgtctacaaa agaagtttta
4801 gatgccactc ttatccatca atccatcact ggtctttatg aaacacgcat tgatttgagt
4861 cagctaggag gtgacgctgc taacgacgaa aactacgctc tggctgctta actcgagtaa
4921 ggatctccag gcatcaaata aaacgaaagg ctcagtcgaa agactgggcc tttcgtttta
4981 tctgttgttt gtcggtgaac gctctctact agagtcacac tggctcacct tcgggtgggc
5041 ctttctgcgt ttatacctag ggatatattc cgcttcctcg ctcactgact cgctacgctc
5101 ggtcgttcga ctgcggcgag cggaaatggc ttacgaacgg ggcggagatt tcctggaaga
5161 tgccaggaag atacttaaca gggaagtgag agggccgcgg caaagccgtt tttccatagg
5221 ctccgccccc ctgacaagca tcacgaaatc tgacgctcaa atcagtggtg gcgaaacccg
5281 acaggactat aaagatacca ggcgtttccc cctggcggct ccctcgtgcg ctctcctgtt
5341 cctgcctttc ggtttaccgg tgtcattccg ctgttatggc cgcgtttgtc tcattccacg
5401 cctgacactc agttccgggt aggcagttcg ctccaagctg gactgtatgc acgaaccccc
5461 cgttcagtcc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggaaag
5521 acatgcaaaa gcaccactgg cagcagccac tggtaattga tttagaggag ttagtcttga
5581 agtcatgcgc cggttaaggc taaactgaaa ggacaagttt tggtgactgc gctcctccaa
5641 gccagttacc tcggttcaaa gagttggtag ctcagagaac cttcgaaaaa ccgccctgca
5701 aggcggtttt ttcgttttca gagcaagaga ttacgcgcag accaaaacga tctcaagaag
5761 atcatcttat taatcagata aaatatttct agatttcagt gcaatttatc tcttcaaatg
5821 tagcacctga agtcagcccc atacgatata agttgttact agtgcttgga ttctcaccaa
5881 taaaaaacgc ccggcggcaa ccgagcgttc tgaacaaatc cagatggagt tctgaggtca
5941 ttactggatc tatcaacagg agtccaagcg agctcgatat caaattacgc cccgccctgc
6001 cactcatcgc agtactgttg taattcatta agcattctgc cgacatggaa gccatcacaa
6061 acggcatgat gaacctgaat cgccagcggc atcagcacct tgtcgccttg cgtataatat
6121 ttgcccatgg tgaaaacggg ggcgaagaag ttgtccatat tggccacgtt taaatcaaaa
6181 ctggtgaaac tcacccaggg attggctgag acgaaaaaca tattctcaat aaacccttta
6241 gggaaatagg ccaggttttc accgtaacac gccacatctt gcgaatatat gtgtagaaac
6301 tgccggaaat cgtcgtggta ttcactccag agcgatgaaa acgtttcagt ttgctcatgg
6361 aaaacggtgt aacaagggtg aacactatcc catatcacca gctcaccgtc tttcattgcc
6421 atacgaaatt ccggatgagc attcatcagg cgggcaagaa tgtgaataaa ggccggataa
6481 aacttgtgct tatttttctt tacggtcttt aaaaaggccg taatatccag ctgaacggtc
6541 tggttatagg tacattgagc aactgactga aatgcctcaa aatgttcttt acgatgccat
6601 tgggatatat caacggtggt atatccagtg atttttttct ccattttagc ttccttagct
6661 cctgaaaatc tcgataactc aaaaaatacg cccggtagtg atcttatttc attatggtga
6721 aagttggaac ctcttacgtg ccgatcaacg tctcattttc gccagatatc
//
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