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CRISPR/Cas9 gene editing technology has been very effective in editing genes in many plant species including rice. Here we further improve the current CRISPR/Cas9 gene editing technology in both efficiency and time needed for isolation of transgene-free and target gene-edited plants. We coupled the CRISPR/Cas9 cassette with a unit that activates anthocyanin biosynthesis, providing a visible marker for detecting the presence of transgenes. The anthocyanin-marker assisted CRISPR (AAC) technology enables us to identify transgenic events even at calli stage, to select transformants with elevated Cas9 expression, and to identify transgene-free plants in the field. We used the AAC technology to edit LAZY1 and G1 and successfully generated many transgene-free and target gene-edited plants at T1 generation. The AAC technology greatly reduced the labor, time, and costs needed for editing target genes in rice.

Leaf mustard (Brassica juncea L. Czern & Coss), an important vegetable crop, experiences pronounced adversity due to seasonal drought stress, particularly at the seed germination stage. Although there is partial comprehension of drought-responsive genes, the role of long non-coding RNAs (lncRNAs) in adjusting mustard's drought stress response is largely unexplored. In this study, we showed that the drought-tolerant cultivar 'Weiliang' manifested a markedly lower base water potential (−1.073 MPa vs −0.437 MPa) and higher germination percentage (41.2% vs 0%) than the drought-susceptible cultivar 'Shuidong' under drought conditions. High throughput RNA sequencing techniques revealed a significant repertoire of lncRNAs from both cultivars during germination under drought stress, resulting in the identification of 2,087 differentially expressed lncRNAs (DELs) and their correspondingly linked 12,433 target genes. It was noted that 84 genes targeted by DEL exhibited enrichment in the photosynthesis pathway. Gene network construction showed that MSTRG.150397, a regulatory lncRNA, was inferred to potentially modulate key photosynthetic genes (Psb27, PetC, PetH, and PsbW), whilst MSTRG.107159 was indicated as an inhibitory regulator of six drought-responsive PIP genes. Further, weighted gene co-expression network analysis (WGCNA) corroborated the involvement of light intensity and stress response genes targeted by the identified DELs. The precision and regulatory impact of lncRNA were verified through qPCR. This study extends our knowledge of the regulatory mechanisms governing drought stress responses in mustard, which will help strategies to augment drought tolerance in this crop. [ABSTRACT FROM AUTHOR]

Introgression is an important driver of new genetic variation that increases species and genetic diversity. However, the relationship between introgression and geographic distribution of upland cotton is still unclear. Herein, we explored geographically comprehensive genomic data based on 890 upland cotton accessions to decipher the degree of introgression between different geographic distributions and its effect on selection and fiber quality. We found introgression intervals to exist across different geographic distributions. Introgression is affected by the similarity of the environment in which they live, and those with similar ecological environments tend to share the same introgression area. Introgression is affected by artificial selection. A genome-wide association study (GWAS) meta-analysis was performed with 6 fiber traits and identified 261 quantitative trait loci (QTLs). We found that 67 QTLs had introgression signals, and the genome interval size was 118.81 Mb, while 123 QTLs had selection signals, and the genomic interval was 28.38 Mb. These results provide insights into the population-scaled introgression landscape, suggesting that introgression contributed to the cotton genetic improvement, which provides a useful reference for studying intraspecific introgressions from different geographical distributions in other species. [ABSTRACT FROM AUTHOR]

The knock-out vector of the three miR156 family miR156A, miR156B, miR156C was constructed by the CRISPR system, which establishes a foundation for future studies of gene function of Eucalyptus grandis miR156 family genes. pCBC plasmid was selected as the template to amplify the target genes miR156A-gRNA, miR156B-gRNA, miR156C-gRNA by PCR, and connect them to the linear pHDE vector with digestion using Bsal, that the recombinant had been transferred into DH5α. PCR identification proved that the insertion of the recombinant cloning, the quality identification of the positive plasmid by PCR and sequencing analysis. The sequence analysis showed that the CRISPR vector sequence constructed by homologous recombination method had no mutation. The positive rate of recombinants was 100%. [ABSTRACT FROM AUTHOR]