ndCRIPRCastargetedgenomeediting教程分析.pptVIP

Screening for essential genes for PA/LFnDTA and diphtheria toxin (DT) toxicity. Screening for essential genes for PA/LFnDTA and diphtheria toxin (DT) toxicity. Genetic validation of candidate genes 3.3 CRISPR/Cas9在植物中的应用 Targeted genes： Bread wheat (Triticum aestivumL., 2n= 42, AABBDD) TaMLO-A1, TaMLO-B1and TaMLOD1 CRISPR/Cas9TALEN in Bread whea (tresistance to powdery mildew) PAM The coding sequences of the two nuclease monomers are expressed from the maize Ubiquitin 1 promoter and separated by a T2A translational skipping sequence PCR-RE assay TALEN-induced mutant TaMLOalleles identified by sequencing 15 representative transgenic wheat plants Molecular and genetic analysis of TALEN-induced mutations in TaMLOhomoeologs and their transmission to the T1 generation Sequences of T-MLO-induced mutations in the three MLOhomoeoalleles in the protoplasts Loss of TaMLO function confers resistance of bread wheat to powdery mildew disease Percentage of microcolonies formed from the total number of germinated spores of Blumeria graminisf. sp. tritici(Bgt) inoculated on the leaves Micrographs of microcolony formation of Bgton the surfaces of leaves Loss of TaMLO function confers resistance of bread wheat to powdery mildew disease Conclusions TALENs and the CRISPR/Cas9 system can be used to generate beneficial genetic traits in hexaploid bread wheat。 The NHEJ pathway enables targeted DNA insertion。 Provide a methodological framework to improve polyploid crops。 Other Conclusions The variation in mutagenesis efficiency（诱变效率） among different genes may stem from distinct sgRNA binding strength to individual target sequences or distinct chromatin structure and epigenetic（表观遗传） state . The different genome mutagenesis frequencies and patterns in plants are due to distinct plant genotypes or physiological states requires future investigation. the sgRNA:pcoCas9 system could facilitate（促进） multiplex genome editing in plants. Targeted gene with multiple sgRNAs co