| 质粒名称: | pEcCas |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 73227 |
| 存储实验室: | Sheng Yang |
|
载体骨架基本信息 |
|
| 载体名称: | pWM119 |
| 载体出品公司: | -- |
| 载体抗性: | 卡那霉素 |
| 空载体大小: | -- |
| 完整质粒大小: | -- |
| 载体修饰: | 用Kan替换Amp |
| 载体类型: | 细菌表达，CRISPR；红色重组酶 |
| 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,pCASsac |
| 物种来源: | S. pyogenes MGAS5005 |
| 插入大小: | -- |
| 基因突变: | -- |
| GenBank编号 | -- |
| 启动子 | native promoter |
| 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 | pENTR-R |
| 3' 测序引物 | AraC-F-New |
|
文献引用方法 |
|
| 材料和方法部分: | pEcCas was a gift from Sheng Yang (Addgene plasmid # 73227 ; http://n2t.net/addgene:73227 ; RRID:Addgene_73227) |
| 参考文献部分: | A modified pCas/pTargetF system for CRISPR-Cas9-assisted genome editing in Escherichia coli. Li , Q i, Sun, Bingbing, Chen, Jun, Zhang, Yiwen, Jiang, Y u Jiang, Yang, Sheng. Acta Biochimica et Biophysica Sinica, 2021 10.1093/abbs/gmab036 |
买家导航
pEcCas载体质粒基本信息
pEcCas质粒图谱载体图谱和pEcCas载体序列质粒序列多克隆位点信息
pEcCas质粒载体简介
pEcCas质粒序列载体序列
LOCUS pEcCas 14589 bp DNA circular SYN 14-APR-2025
DEFINITION Constitutive expression of cas9 and inducible expression of lambda
RED and sgRNA.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 14589)
AUTHORS Li Q, Sun B, Chen J, Zhang Y, Jiang Y, Yang S
TITLE A modified pCas/pTargetF system for CRISPR-Cas9-assisted genome
editing in Escherichia coli.
JOURNAL Acta Biochim Biophys Sin (Shanghai). 2021 Apr 15;53(5):620-627. doi:
10.1093/abbs/gmab036.
PUBMED 33764372
REFERENCE 2 (bases 1 to 14589)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Apr 14, 2025 from SnapGene Server 8.0.1
https://www.biofeng.com
COMMENT SGRef: number: 1; type: "Journal Article"; doi: "10"; journalName:
"Acta Biochim Biophys Sin (Shanghai)"; date: "2021-04-15- 15";
volume: "53"; issue: "5"; pages: "620-627"
COMMENT Sequence Label: pEcCas
FEATURES Location/Qualifiers
source 1..14589
/mol_type="other DNA"
/organism="synthetic DNA construct"
rep_origin 245..467
/label=pSC101 ori
/note="low-copy replication origin that requires the Rep101
protein"
CDS complement(956..1771)
/codon_start=1
/product="aminoglycoside phosphotransferase"
/label=KanR
/note="confers resistance to kanamycin in bacteria or G418
(Geneticin(R)) in eukaryotes"
CDS complement(956..1771)
/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"
primer_bind 1679..1698
/label=Kan-R
/note="Kanamycin resistance gene, reverse primer"
primer_bind 1763..1782
/label=pENTR-R
/note="pENTR vectors, reverse primer"
CDS 2119..6225
/codon_start=1
/product="Cas9 (Csn1) endonuclease from the Streptococcus
pyogenes Type II CRISPR/Cas system"
/label=Cas9
CDS 2119..6222
/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"
CDS complement(6355..7233)
/codon_start=1
/gene="araC"
/product="L-arabinose regulatory protein"
/label=araC
/translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
EKVNDVAVKLS"
promoter 7260..7544
/gene="araBAD"
/label=araBAD promoter
/note="promoter of the L-arabinose operon of E. coli; the
araC regulatory gene is transcribed in the opposite
direction (Guzman et al., 1995)"
primer_bind 7467..7486
/label=pBAD Forward
/note="For vectors with E. coli araBAD promoter, forward
primer"
RBS 7571..7579
/label=Shine-Dalgarno sequence
/note="full consensus sequence for ribosome-binding sites
upstream of start codons in E. coli; complementary to a
region in the 3' end of the 16S rRNA (Chen et al., 1994)"
CDS 7588..8004
/codon_start=1
/gene="gam"
/product="inhibitor of the host RecBCD nuclease in the
lambda Red system"
/label=Gam
/note="protects the double-stranded DNA substrate for
recombination"
/translation="MDINTETEIKQKHSLTPFPVFLISPAFRGRYFHSYFRSSAMNAYY
IQDRLEAQSWARHYQQLAREEKEAELADDMEKGLPQHLFESLCIDHLQRHGASKKSITR
AFDDDVEFQERMAEHIRYMVETIAHHQVDIDSEV"
CDS 8010..8795
/codon_start=1
/gene="bet"
/product="single-stranded DNA binding recombinase in the
lambda Red system"
/label=Beta
/note="mediates pairing of a single-stranded DNA with a
complementary target"
/translation="MSTALATLAGKLAERVGMDSVDPQELITTLRQTAFKGDASDAQFI
ALLIVANQYGLNPWTKEIYAFPDKQNGIVPVVGVDGWSRIINENQQFDGMDFEQDNESC
TCRIYRKDRNHPICVTEWMDECRREPFKTREGREITGPWQSHPKRMLRHKAMIQCARLA
FGFAGIYDKDEAERIVENTAYTAERQPERDITPVNDETMQEINTLLIALDKTWDDDLLP
LCSQIFRRDIRASSELTQAEAVKALGFLKQKAAEQKVAA"
CDS 8792..9472
/codon_start=1
/gene="exo"
/product="5' to 3' double-stranded DNA exonuclease in the
lambda Red system"
/label=Exo
/note="creates 3' single-stranded DNA tails"
/translation="MTPDIILQRTGIDVRAVEQGDDAWHKLRLGVITASEVHNVIAKPR
SGKKWPDMKMSYFHTLLAEVCTGVAPEVNAKALAWGKQYENDARTLFEFTSGVNVTESP
IIYRDESMRTACSPDGLCSDGNGLELKCPFTSRDFMKFRLGGFEAIKSAYMAQVQYSMW
VTRKNAWYFANYDPRMKREGLHYVVIERDEKYMASFDEIVPEFIEKMDEALAEIGFVFG
EQWR"
terminator 9473..9717
/label=lambda tL3 terminator
/note="transcription terminator tL3 from phage lambda"
CDS join(10054..10140,10141..11475)
/codon_start=1
/gene="Bacillus subtilis sacB"
/product="secreted levansucrase that renders bacterial
growth sensitive to sucrose"
/label=SacB
/note="negative selection marker"
misc_RNA complement(11598..11673)
/label=gRNA scaffold
/note="guide RNA scaffold for the Streptococcus pyogenes
CRISPR/Cas9 system"
promoter complement(11695..11813)
/label=rhaB promoter
/note="promoter of the E. coli rhaBAD operon, conferring
tight induction with L-rhamnose and repression with
D-glucose in the presence of RhaR and RhaS (Giacalone et
al., 2006)"
CDS 11959..12795
/codon_start=1
/gene="E. coli rhaS"
/product="positive regulator of the rhaB promoter
"
/label=rhaS
/note="activated by L-rhamnose"
CDS 12869..13717
/codon_start=1
/gene="E. coli rhaR"
/product="transcriptional activator of rhaR and rhaS"
/label=rhaR
/note="needed for rhamnose-dependent activation of the rhaB
promoter"
CDS complement(join(13836..14589,1..197))
/codon_start=1
/gene="rep101"
/product="RepA protein needed for replication with the
pSC101 origin"
/label=Rep101
ORIGIN
1 gcattttccc tactgatgtt catcatctga gcgtattggt tataagtgaa cgataccgtc
61 cgttctttcc ttgtagggtt ttcaatcgtg gggttgagta gtgccacaca gcataaaatt
121 agcttggttt catgctccgt taagtcatag cgactaatcg ctagttcatt tgctttgaaa
181 acaactaatt cagacataca tctcaattgg tctaggtgat tttaatcact ataccaattg
241 agatgggcta gtcaatgata attactagtc cttttccttt gagttgtggg tatctgtaaa
301 ttctgctaga cctttgctgg aaaacttgta aattctgcta gaccctctgt aaattccgct
361 agacctttgt gtgttttttt tgtttatatt caagtggtta taatttatag aataaagaaa
421 gaataaaaaa agataaaaag aatagatccc agccctgtgt ataactcact actttagtca
481 gttccgcagt attacaaaag gatgtcgcaa acgctgtttg ctcctctaca aaacagacct
541 taaaacccta aaggcttaag tagcaccctc gcaagctcgg gcaaatcgct gaatattcct
601 tttgtctccg accatcaggc acctgagtcg ctgtcttttt cgtgacattc agttcgctgc
661 gctcacggct ctggcagtga atgggggtaa atggcactac aggcgccttt tatggattca
721 tgcaaggaaa ctacccataa tacaagaaaa gcccgtcacg ggcttctcag ggcgttttat
781 ggcgggtctg ctatgtggtg ctatctgact ttttgctgtt cagcagttcc tgccctctga
841 ttttccagtc tgaccacttc ggattatccc gtgacaggtc attcagactg gctaatgcac
901 ccagtaaggc agcggtatca tcaacccact gcagcaacca attaaccaat tctgattata
961 aaaactcatc gagcatcaaa tgaaactgca atttattcat atcaggatta tcaataccat
1021 atttttgaaa aagccgtttc tgtaatgaag gagaaaactc accgaggcag ttccatagga
1081 tggcaagatc ctggtatcgg tctgcgattc cgactcgtcc aacatcaata caacctatta
1141 atttcccctc gtcaaaaata aggttatcaa gtgagaaatc accatgagtg acgactgaat
1201 ccggtgagaa tggcaaaagc ttatgcattt ctttccagac ttgttcaaca ggccagccat
1261 tacgctcgtc atcaaaatca ctcgcatcaa ccaaaccgtt attcattcgt gattgcgcct
1321 gagcgagacg aaatacgcga tcgctgttaa aaggacaatt acaaacagga atcgaatgca
1381 accggcgcag gaacactgcc agcgcatcaa caatattttc acctgaatca ggatattctt
1441 ctaatacctg gaatgctgtt ttcccgggga tcgcagtggt gagtaaccat gcatcatcag
1501 gagtacggat aaaatgcttg atggtcggaa gaggcataaa ttccgtcagc cagtttagtc
1561 tgaccatctc atctgtaaca tcattggcaa cgctaccttt gccatgtttc agaaacaact
1621 ctggcgcatc gggcttccca tacaatcgat agattgtcgc acctgattgc ccgacattat
1681 cgcgagccca tttataccca tataaatcag catccatgtt ggaatttaat cgcggcctcg
1741 agcaagacgt ttcccgttga atatggctca taacacccct tgtattactg tttatgtaag
1801 cagacagttt tattgttcat gatgatatat ttttatcttg tgcaatgtaa catcagagat
1861 tttgagacac aacgtggctt tccctgcagc tgaacaatac ttaatactat agaatgataa
1921 caaaataaac tactttttaa aagaattttg tgttataatc tatttattat taagtattgg
1981 gtaatatttt ttgaagagat attttgaaaa agaaaaatta aagcatatta aactaatttc
2041 ggaggtcatt aaaactatta ttgaaatcat caaactcatt atggatttaa tttaaacttt
2101 ttattttagg aggcaaaaat ggataagaaa tactcaatag gcttagatat cggcacaaat
2161 agcgtcggat gggcggtgat cactgatgat tataaggttc cgtctaaaaa gttcaaggtt
2221 ctgggaaata cagaccgcca cagtatcaaa aaaaatctta taggggctct tttatttgac
2281 agtggagaga cagcggaagc gactcgtctc aaacggacag ctcgtagaag gtatacacgt
2341 cggaagaatc gtatttgtta tctacaggag attttttcaa atgagatggc gaaagtagat
2401 gatagtttct ttcatcgact tgaagagtct tttttggtgg aagaagacaa gaagcatgaa
2461 cgtcatccta tttttggaaa tatagtagat gaagttgctt atcatgagaa atatccaact
2521 atctatcatc tgcgaaaaaa attggtagat tctactgata aagcggattt gcgcttaatc
2581 tatttggcct tagcgcatat gattaagttt cgtggtcatt ttttgattga gggagattta
2641 aatcctgata atagtgatgt ggacaaacta tttatccagt tggtacaaac ctacaatcaa
2701 ttatttgaag aaaaccctat taacgcaagt ggagtagatg ctaaagcgat tctttctgca
2761 cgattgagta aatcaagacg attagaaaat ctcattgctc agctccccgg tgagaagaaa
2821 aatggcttat ttgggaatct cattgctttg tcattgggtt tgacccctaa ttttaaatca
2881 aattttgatt tggcagaaga tgctaaatta cagctttcaa aagatactta cgatgatgat
2941 ttagataatt tattggcgca aattggagat caatatgctg atttgttttt ggcagctaag
3001 aatttatcag atgctatttt actttcagat atcctaagag taaatactga aataactaag
3061 gctcccctat cagcttcaat gattaaacgc tacgatgaac atcatcaaga cttgactctt
3121 ttaaaagctt tagttcgaca acaacttcca gaaaagtata aagaaatctt ttttgatcaa
3181 tcaaaaaacg gatatgcagg ttatattgat gggggagcta gccaagaaga attttataaa
3241 tttatcaaac caattttaga aaaaatggat ggtactgagg aattattggt gaaactaaat
3301 cgtgaagatt tgctgcgcaa gcaacggacc tttgacaacg gctctattcc ccatcaaatt
3361 cacttgggtg agctgcatgc tattttgaga agacaagaag acttttatcc atttttaaaa
3421 gacaatcgtg agaagattga aaaaatcttg acttttcgaa ttccttatta tgttggtcca
3481 ttggcgcgtg gcaatagtcg ttttgcatgg atgactcgga agtctgaaga aacaattacc
3541 ccatggaatt ttgaagaagt tgtcgataaa ggtgcttcag ctcaatcatt tattgaacgc
3601 atgacaaact ttgataaaaa tcttccaaat gaaaaagtac taccaaaaca tagtttgctt
3661 tatgagtatt ttacggttta taacgaattg acaaaggtca aatatgttac tgaaggaatg
3721 cgaaaaccag catttctttc aggtgaacag aagaaagcca ttgttgattt actcttcaaa
3781 acaaatcgaa aagtaaccgt taagcaatta aaagaagatt atttcaaaaa aatagaatgt
3841 tttgatagtg ttgaaatttc aggagttgaa gatagattta atgcttcatt aggtacctac
3901 catgatttgc taaaaattat taaagataaa gattttttgg ataatgaaga aaatgaagat
3961 atcttagagg atattgtttt aacattgacc ttatttgaag atagggagat gattgaggaa
4021 agacttaaaa catatgctca cctctttgat gataaggtga tgaaacagct taaacgtcgc
4081 cgttatactg gttggggacg tttgtctcga aaattgatta atggtattag ggataagcaa
4141 tctggcaaaa caatattaga ttttttgaaa tcagatggtt ttgccaatcg caattttatg
4201 cagctgatcc atgatgatag tttgacattt aaagaagaca ttcaaaaagc acaagtgtct
4261 ggacaaggcg atagtttaca tgaacatatt gcaaatttag ctggtagccc tgctattaaa
4321 aaaggtattt tacagactgt aaaagttgtt gatgaattgg tcaaagtaat ggggcggcat
4381 aagccagaaa atatcgttat tgaaatggca cgtgaaaatc agacaactca aaagggccag
4441 aaaaattcgc gagagcgtat gaaacgaatc gaagaaggta tcaaagaatt aggaagtcag
4501 attcttaaag agcatcctgt tgaaaatact caattgcaaa atgaaaagct ctatctctat
4561 tatctccaaa atggaagaga catgtatgtg gaccaagaat tagatattaa tcgtttaagt
4621 gattatgatg tcgatcacat tgttccacaa agtttcctta aagacgattc aatagacaat
4681 aaggtcttaa cgcgttctga taaaaatcgt ggtaaatcgg ataacgttcc aagtgaagaa
4741 gtagtcaaaa agatgaaaaa ctattggaga caacttctaa acgccaagtt aatcactcaa
4801 cgtaagtttg ataatttaac gaaagctgaa cgtggaggtt tgagtgaact tgataaagct
4861 ggttttatca aacgccaatt ggttgaaact cgccaaatca ctaagcatgt ggcacaaatt
4921 ttggatagtc gcatgaatac taaatacgat gaaaatgata aacttattcg agaggttaaa
4981 gtgattacct taaaatctaa attagtttct gacttccgaa aagatttcca attctataaa
5041 gtacgtgaga ttaacaatta ccatcatgcc catgatgcgt atctaaatgc cgtcgttgga
5101 actgctttga ttaagaaata tccaaaactt gaatcggagt ttgtctatgg tgattataaa
5161 gtttatgatg ttcgtaaaat gattgctaag tctgagcaag aaataggcaa agcaaccgca
5221 aaatatttct tttactctaa tatcatgaac ttcttcaaaa cagaaattac acttgcaaat
5281 ggagagattc gcaaacgccc tctaatcgaa actaatgggg aaactggaga aattgtctgg
5341 gataaagggc gagattttgc cacagtgcgc aaagtattgt ccatgcccca agtcaatatt
5401 gtcaagaaaa cagaagtaca gacaggcgga ttctccaagg agtcaatttt accaaaaaga
5461 aattcggaca agcttattgc tcgtaaaaaa gactgggatc caaaaaaata tggtggtttt
5521 gatagtccaa cggtagctta ttcagtccta gtggttgcta aggtggaaaa agggaaatcg
5581 aagaagttaa aatccgttaa agagttacta gggatcacaa ttatggaaag aagttccttt
5641 gaaaaaaatc cgattgactt tttagaagct aaaggatata aggaagttaa aaaagactta
5701 atcattaaac tacctaaata tagtcttttt gagttagaaa acggtcgtaa acggatgctg
5761 gctagtgccg gagaattaca aaaaggaaat gagctggctc tgccaagcaa atatgtgaat
5821 tttttatatt tagctagtca ttatgaaaag ttgaagggta gtccagaaga taacgaacaa
5881 aaacaattgt ttgtggagca gcataagcat tatttagatg agattattga gcaaatcagt
5941 gaattttcta agcgtgttat tttagcagat gccaatttag ataaagttct tagtgcatat
6001 aacaaacata gagacaaacc aatacgtgaa caagcagaaa atattattca tttatttacg
6061 ttgacgaatc ttggagctcc cgctgctttt aaatattttg atacaacaat tgatcgtaaa
6121 cgatatacgt ctacaaaaga agttttagat gccactctta tccatcaatc catcactggt
6181 ctttatgaaa cacgcattga tttgagtcag ctaggaggtg actgaagtat attttagatg
6241 aagattattt cttaatctag acatgagcgg atacatattt gaatgtattt agaaaaataa
6301 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgcatcga tttattatga
6361 caacttgacg gctacatcat tcactttttc ttcacaaccg gcacggaact cgctcgggct
6421 ggccccggtg cattttttaa atacccgcga gaagtagagt tgatcgtcaa aaccaacatt
6481 gcgaccgacg gtggcgatag gcatccgggt ggtgctcaaa agcagcttcg cctggctgat
6541 acgttggtcc tcgcgccagc ttaagacgct aatccctaac tgctggcgga aaagatgtga
6601 cagacgcgac ggcgacaagc aaacatgctg tgcgacgctg gcgatatcaa aattgctgtc
6661 tgccaggtga tcgctgatgt actgacaagc ctcgcgtacc cgattatcca tcggtggatg
6721 gagcgactcg ttaatcgctt ccatgcgccg cagtaacaat tgctcaagca gatttatcgc
6781 cagcagctcc gaatagcgcc cttccccttg cccggcgtta atgatttgcc caaacaggtc
6841 gctgaaatgc ggctggtgcg cttcatccgg gcgaaagaac cccgtattgg caaatattga
6901 cggccagtta agccattcat gccagtaggc gcgcggacga aagtaaaccc actggtgata
6961 ccattcgcga gcctccggat gacgaccgta gtgatgaatc tctcctggcg ggaacagcaa
7021 aatatcaccc ggtcggcaaa caaattctcg tccctgattt ttcaccaccc cctgaccgcg
7081 aatggtgaga ttgagaatat aacctttcat tcccagcggt cggtcgataa aaaaatcgag
7141 ataaccgttg gcctcaatcg gcgttaaacc cgccaccaga tgggcattaa acgagtatcc
7201 cggcagcagg ggatcatttt gcgcttcagc catacttttc atactcccgc cattcagaga
7261 agaaaccaat tgtccatatt gcatcagaca ttgccgtcac tgcgtctttt actggctctt
7321 ctcgctaacc aaaccggtaa ccccgcttat taaaagcatt ctgtaacaaa gcgggaccaa
7381 agccatgaca aaaacgcgta acaaaagtgt ctataatcac ggcagaaaag tccacattga
7441 ttatttgcac ggcgtcacac tttgctatgc catagcattt ttatccataa gattagcgga
7501 tcctacctga cgctttttat cgcaactctc tactgtttct ccatacccgt ttttttggga
7561 attcgagctc taaggaggtt ataaaaaatg gatattaata ctgaaactga gatcaagcaa
7621 aagcattcac taaccccctt tcctgttttc ctaatcagcc cggcatttcg cgggcgatat
7681 tttcacagct atttcaggag ttcagccatg aacgcttatt acattcagga tcgtcttgag
7741 gctcagagct gggcgcgtca ctaccagcag ctcgcccgtg aagagaaaga ggcagaactg
7801 gcagacgaca tggaaaaagg cctgccccag cacctgtttg aatcgctatg catcgatcat
7861 ttgcaacgcc acggggccag caaaaaatcc attacccgtg cgtttgatga cgatgttgag
7921 tttcaggagc gcatggcaga acacatccgg tacatggttg aaaccattgc tcaccaccag
7981 gttgatattg attcagaggt ataaaacgaa tgagtactgc actcgcaacg ctggctggga
8041 agctggctga acgtgtcggc atggattctg tcgacccaca ggaactgatc accactcttc
8101 gccagacggc atttaaaggt gatgccagcg atgcgcagtt catcgcatta ctgatcgttg
8161 ccaaccagta cggccttaat ccgtggacga aagaaattta cgcctttcct gataagcaga
8221 atggcatcgt tccggtggtg ggcgttgatg gctggtcccg catcatcaat gaaaaccagc
8281 agtttgatgg catggacttt gagcaggaca atgaatcctg tacatgccgg atttaccgca
8341 aggaccgtaa tcatccgatc tgcgttaccg aatggatgga tgaatgccgc cgcgaaccat
8401 tcaaaactcg cgaaggcaga gaaatcacgg ggccgtggca gtcgcatccc aaacggatgt
8461 tacgtcataa agccatgatt cagtgtgccc gtctggcctt cggatttgct ggtatctatg
8521 acaaggatga agccgagcgc attgtcgaaa atactgcata cactgcagaa cgtcagccgg
8581 aacgcgacat cactccggtt aacgatgaaa ccatgcagga gattaacact ctgctgatcg
8641 ccctggataa aacatgggat gacgacttat tgccgctctg ttcccagata tttcgccgcg
8701 acattcgtgc atcgtcagaa ctgacacagg ccgaagcagt aaaagctctt ggattcctga
8761 aacagaaagc cgcagagcag aaggtggcag catgacaccg gacattatcc tgcagcgtac
8821 cgggatcgat gtgagagctg tcgaacaggg ggatgatgcg tggcacaaat tacggctcgg
8881 cgtcatcacc gcttcagaag ttcacaacgt gatagcaaaa ccccgctccg gaaagaagtg
8941 gcctgacatg aaaatgtcct acttccacac cctgcttgct gaggtttgca ccggtgtggc
9001 tccggaagtt aacgctaaag cactggcctg gggaaaacag tacgagaacg acgccagaac
9061 cctgtttgaa ttcacttccg gcgtgaatgt tactgaatcc ccgatcatct atcgcgacga
9121 aagtatgcgt accgcctgct ctcccgatgg tttatgcagt gacggcaacg gccttgaact
9181 gaaatgcccg tttacctccc gggatttcat gaagttccgg ctcggtggtt tcgaggccat
9241 aaagtcagct tacatggccc aggtgcagta cagcatgtgg gtgacgcgaa aaaatgcctg
9301 gtactttgcc aactatgacc cgcgtatgaa gcgtgaaggc ctgcattatg tcgtgattga
9361 gcgggatgaa aagtacatgg cgagttttga cgagatcgtg ccggagttca tcgaaaaaat
9421 ggacgaggca ctggctgaaa ttggttttgt atttggggag caatggcgat gacgcatcct
9481 cacgataata tccgggtagg cgcaatcact ttcgtctact ccgttacaaa gcgaggctgg
9541 gtatttcccg gcctttctgt tatccgaaat ccactgaaag cacagcggct ggctgaggag
9601 ataaataata aacgaggggc tgtatgcaca aagcatcttc tgttgagtta agaacgagta
9661 tcgagatggc acatagcctt gctcaaattg gaatcaggtt tgtgccaata ccagtagaaa
9721 cagacgaaga atccatgggt atggacagat cctctttagg cccgtagtct gcaaatcctt
9781 ttatgatttt ctatcaaaca aaagaggaaa atagaccagt tgcaatccaa acgagagtct
9841 aatagaatga ggtcgaaaag taaatcgcgc gggtttgtta ctgataaagc aggcaagacc
9901 taaaatgtgt aaagggcaaa gtgtatactt tggcgtcacc ccttacatat tttaggtctt
9961 tttttattgt gcgtaactaa cttgccatct tcaaacagga gggctggaag aagcagaccg
10021 ctaacacagt acataaaaaa ggagacatga acgatgaaca tcaaaaagtt tgcaaaacaa
10081 gcaacagtat taacctttac taccgcactg ctggcaggag gcgcaactca agcgtttgcg
10141 aaagaaacga accaaaagcc atataaggaa acatacggca tttcccatat tacacgccat
10201 gatatgctgc aaatccctga acagcaaaaa aatgaaaaat atcaagttcc tgaattcgat
10261 tcgtccacaa ttaaaaatat ctcttctgca aaaggcctgg acgtttggga cagctggcca
10321 ttacaaaacg ctgacggcac tgtcgcaaac tatcacggct accacatcgt ctttgcatta
10381 gccggagatc ctaaaaatgc ggatgacaca tcgatttaca tgttctatca aaaagtcggc
10441 gaaacttcta ttgacagctg gaaaaacgct ggccgcgtct ttaaagacag cgacaaattc
10501 gatgcaaatg attctatcct aaaagaccaa acacaagaat ggtcaggttc agccacattt
10561 acatctgacg gaaaaatccg tttattctac actgatttct ccggtaaaca ttacggcaaa
10621 caaacactga caactgcaca agttaacgta tcagcatcag acagctcttt gaacatcaac
10681 ggtgtagagg attataaatc aatctttgac ggtgacggaa aaacgtatca aaatgtacag
10741 cagttcatcg atgaaggcaa ctacagctca ggcgacaacc atacgctgag agatcctcac
10801 tacgtagaag ataaaggcca caaatactta gtatttgaag caaacactgg aactgaagat
10861 ggctaccaag gcgaagaatc tttatttaac aaagcatact atggcaaaag cacatcattc
10921 ttccgtcaag aaagtcaaaa acttctgcaa agcgataaaa aacgcacggc tgagttagca
10981 aacggcgctc tcggtatgat tgagctaaac gatgattaca cactgaaaaa agtgatgaaa
11041 ccgctgattg catctaacac agtaacagat gaaattgaac gcgcgaacgt ctttaaaatg
11101 aacggcaaat ggtacctgtt cactgactcc cgcggatcaa aaatgacgat tgacggcatt
11161 acgtctaacg atatttacat gcttggttat gtttctaatt ctttaactgg cccatacaag
11221 ccgctgaaca aaactggcct tgtgttaaaa atggatcttg atcctaacga tgtaaccttt
11281 acttactcac acttcgctgt acctcaagcg aaaggaaaca atgtcgtgat tacaagctat
11341 atgacaaaca gaggattcta cgcagacaaa caatcaacgt ttgcgccaag cttcctgctg
11401 aacatcaaag gcaagaaaac atctgttgtc aaagacagca tccttgaaca aggacaatta
11461 acagttaaca aataaaaacg caaaagaaaa tgccgatatc ctattggcat tttcttttat
11521 ttcttatcaa cataaaggtg aatcccatat gaactatata aaagcaggca aatggctaac
11581 cgtattcctc aaaaaaagca ccgactcggt gccacttttt caagttgata acggactagc
11641 cttattttaa cttgctattt ctagctctaa aacggttgga ctcaagacga tagttacgac
11701 cagtctaaaa agcgcctgaa ttcgcgacct tctcgttact gacaggaaaa tgggccattg
11761 gcaaccaggg aaagatgaac gtgatgatgt tcacaatttg ctgaattgtg gtgatgtgat
11821 gctcaccgca tttcctgaaa attcacgctg tatcttgaaa aatcgacgtt ttttacgtgg
11881 ttttccgtcg aaaatttaag gtaagaacct gacctcgtga ttactatttc gccgtgttga
11941 cgacatcagg aggccagtat gaccgtatta catagtgtgg atttttttcc gtctggtaac
12001 gcgtccgtgg cgatagaacc ccggctcccg caggcggatt ttcctgaaca tcatcatgat
12061 tttcatgaaa ttgtgattgt cgaacatggc acgggtattc atgtgtttaa tgggcagccc
12121 tataccatca ccggtggcac ggtctgtttc gtacgcgatc atgatcggca tctgtatgaa
12181 cataccgata atctgtgtct gaccaatgtg ctgtatcgct cgccggatcg atttcagttt
12241 ctcgccgggc tgaatcagtt gctgccacaa gagctggatg ggcagtatcc gtctcactgg
12301 cgcgttaacc acagcgtatt gcagcaggtg cgacagctgg ttgcacagat ggaacagcag
12361 gaaggggaaa atgatttacc ctcgaccgcc agtcgcgaga tcttgtttat gcaattactg
12421 ctcttgctgc gtaaaagcag tttgcaggag aacctggaaa acagcgcatc acgtctcaac
12481 ttgcttctgg cctggctgga ggaccatttt gccgatgagg tgaattggga tgccgtggcg
12541 gatcaatttt ctctttcact gcgtacgcta catcggcagc ttaagcagca aacgggactg
12601 acgcctcagc gatacctgaa ccgcctgcga ctgatgaaag cccgacatct gctacgccac
12661 agcgaggcca gcgttactga catcgcctat cgctgtggat tcagcgacag taaccacttt
12721 tcgacgcttt ttcgccgaga gtttaactgg tcaccgcgtg atattcgcca gggacgggat
12781 ggctttctgc aataacgcga atcttctcaa cgtatttgta cgccatattg cgaataatca
12841 acttcgttct ctggccgagg tagccacggt ggcgcatcag ttaaaacttc tcaaagatga
12901 tttttttgcc agcgaccagc aggcagtcgc tgtggctgac cgttatccgc aagatgtctt
12961 tgctgaacat acacatgatt tttgtgagct ggtgattgtc tggcgcggta atggcctgca
13021 tgtactcaac gatcgccctt atcgcattac ccgtggcgat ctcttttaca ttcatgctga
13081 cgataaacac tcctacgctt ccgttaacga tctggttttg cagaatatta tttattgccc
13141 ggagcgtctg aagctgaatc ttgactggca gggggcgatt ccgggattta acgccagcgc
13201 agggcaacca cactggcgct taggtagcat ggggatggcg caggcgcggc aggttatcgg
13261 tcagcttgag catgaaagta gtcagcatgt gccgtttgct aacgaaatgg ctgagttgct
13321 gttcgggcag ttggtgatgt tgctgaatcg ccatcgttac accagtgatt cgttgccgcc
13381 aacatccagc gaaacgttgc tggataagct gattacccgg ctggcggcta gcctgaaaag
13441 tccctttgcg ctggataaat tttgtgatga ggcatcgtgc agtgagcgcg ttttgcgtca
13501 gcaatttcgc cagcagactg gaatgaccat caatcaatat ctgcgacagg tcagagtgtg
13561 tcatgcgcaa tatcttctcc agcatagccg cctgttaatc agtgatattt cgaccgaatg
13621 tggctttgaa gatagtaact atttttcggt ggtgtttacc cgggaaaccg ggatgacgcc
13681 cagccagtgg cgtcatctca attcgcagaa agattaattc gccatgcccc atgggtatgg
13741 acagttttcc ctttgatatg taacggtgaa cagttgttct acttttgttt gttagtcttg
13801 atgcttcact gatagataca agagccataa gaacctcaga tccttccgta tttagccagt
13861 atgttctcta gtgtggttcg ttgtttttgc gtgagccatg agaacgaacc attgagatca
13921 tgcttacttt gcatgtcact caaaaatttt gcctcaaaac tggtgagctg aatttttgca
13981 gttaaagcat cgtgtagtgt ttttcttagt ccgttatgta ggtaggaatc tgatgtaatg
14041 gttgttggta ttttgtcacc attcattttt atctggttgt tctcaagttc ggttacgaga
14101 tccatttgtc tatctagttc aacttggaaa atcaacgtat cagtcgggcg gcctcgctta
14161 tcaaccacca atttcatatt gctgtaagtg tttaaatctt tacttattgg tttcaaaacc
14221 cattggttaa gccttttaaa ctcatggtag ttattttcaa gcattaacat gaacttaaat
14281 tcatcaaggc taatctctat atttgccttg tgagttttct tttgtgttag ttcttttaat
14341 aaccactcat aaatcctcat agagtatttg ttttcaaaag acttaacatg ttccagatta
14401 tattttatga atttttttaa ctggaaaaga taaggcaata tctcttcact aaaaactaat
14461 tctaattttt cgcttgagaa cttggcatag tttgtccact ggaaaatctc aaagccttta
14521 accaaaggat tcctgatttc cacagttctc gtcatcagct ctctggttgc tttagctaat
14581 acaccataa
//
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pEcCas质粒载体应用举例
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