Figure - PMC

Skip to main content

An official website of the United States government

Here's how you know

Here's how you know

Official websites use .gov
A .gov website belongs to an official government organization in the United States.

Secure .gov websites use HTTPS
A lock ( ) or https:// means you've safely connected to the .gov website. Share sensitive information only on official, secure websites.

View full-text article in PMC

. 2018 Jun 28;69(20):4715–4721. doi: 10.1093/jxb/ery245

Search in PMC
Search in PubMed
View in NLM Catalog
Add to search

© The Author(s) 2018. Published by Oxford University Press on behalf of the Society for Experimental Biology.

This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited.

PMC Copyright notice

Fig. 2. — Generation of stable precisely edited rice lines through SDSA-mediated HDR. (A) Only the left homology arm strategy for CRISPR/LbCpf1-mediated HDR in rice. (A-1) PCR-RE analyses of the different genotypes. PCR products amplified by primers ALStestF/R were digested with EcoRV (GACATC). M, DL2000; WT, wild-type; T1, target 1; T2, target 2; MT1, mutated PAM site and mutated target 1; MT2, mutated PAM site and mutated target 2; ME, mutated EcoRV; WT/+, EcoRV cuts the PCR products of the wild type, resulting in 481 bp and 322 bp fragments. Successful HDR leads to EcoRV-resistant bands. (A-2) Sequence analyses of the HDR events. Line 171-2 has one allele with precise HDR, while the other is wild type. Line 171-9 is a chimeric line with three alleles: a precise HDR, a partial HDR, and a wild type. Line 170-1 has a heterozygous partial HDR. Line 172-1 is a chimeric line with three alleles: one base pair substitution, a partial HDR, and a wild type. The sequences shaded in yellow and blue represent wild-type and the designed donor repair template, respectively. Specifically, the sequences shaded in red indicate the expected targeted substitution. (B) Generation of stable precisely edited rice lines using pCXUN-LbCpf1-OsU3-RCR1-RCR2-armed donor vector through CRISPR/Cpf1-mediated HDR. (B-1) PCR-RE analyses of the different genotypes. PCR products amplified by primers ALStestF/R were digested with EcoRV (GACATC). M, DL2000; WT, wild-type; T1, target 1; T2, target 2; MT1, mutated PAM site and mutated target 1; MT2, mutated PAM site and mutated target 2; ME, mutated EcoRV; WT/+, EcoRV cuts the PCR products of the wild type, resulting in 481 bp and 322 bp fragments. EcoRV failed to digest the PCR products if HDR was successful. (B-2) Representative sequences of the different genotypes. Line 169-3 is a chimeric line with three alleles: one precise HDR, one partial HDR, and a wild type. Line 169-2 is a chimeric line with four alleles: a precise HDR, two partial HDR events, and a wild type. Lines 168-4, 168-5, 169-5, and 169-6 have one partial HDR, except for a synonymous G to T base substitution at position 1911 bp in Line 168-4. The 168-2 is a chimeric line with three alleles. The first and second allele have partial HDR, while the third allele is wild type. Line 169-1 has a 253 bp deletion. The sequences shaded in yellow and blue represent wild-type and the designed donor repair template, respectively. Specifically, the sequences shaded in red indicate the expected targeted substitutions. In d#, the # refers to the number of base pairs deleted from the target sites. Different numbers of lines indicate independent lines developed from resistant calli.