CRISPR/Cas9-Mediated Disruption of Xylanase inhibitor protein (XIP) Gene Improved the Dough Quality of Common Wheat

Abstract

The wheat dough quality is of great significance for the end-use of flour. Some genes have been cloned for controlling the protein fractions, grain protein content, starch synthase, grain hardness, etc. Using a unigene map of the recombinant inbred lines (RILs) for “TN 18 × LM 6,” we mapped a quantitative trait locus (QTL) for dough stability time (ST) and SDS-sedimentation values (SV) on chromosome 6A (QSt/Sv-6A-2851). The peak position of the QTL covered two candidate unigenes, and we speculated that TraesCS6A02G077000 (a xylanase inhibitor protein) was the primary candidate gene (named the TaXip gene). The target loci containing the three homologous genes TaXip-6A, TaXip-6B, and TaXip-6D were edited in the variety “Fielder” by clustered regularly interspaced short palindromic repeats–associated protein 9 (CRISPR/Cas9). Two mutant types in the T2:3 generation were obtained (aaBBDD and AAbbdd) with about 120 plants per type. The SVs of aaBBDD, AAbbdd, and WT were 31.77, 27.30, and 20.08 ml, respectively. The SVs of the aaBBDD and AAbbdd were all significantly higher than those of the wild type (WT), and the aaBBDD was significantly higher than the AAbbdd. The STs of aaBBDD, AAbbdd, and WT were 2.60, 2.24, and 2.25 min, respectively. The ST for the aaBBDD was significantly higher than that for WT and was not significantly different between WT and AAbbdd. The above results indicated that XIP in vivo can significantly affect wheat dough quality. The selection of TaXip gene should be a new strategy for developing high-quality varieties in wheat breeding programs.

Xylanase Inhibitors: Defense Players in Plant Immunity with Implications in Agro-Industrial Processing

“…More than 100 complete or partial XIP genes from monocot plants have been deposited in nucleotide databases ( , accessed on 1 September 2022) based on BLASTp search using wheat XIP-I (ADB81849.1) as a query. In wheat, among XIP type XIs, XIP-I , XIP-II , XIP-III , XIP-R1 , XIP-R2 , xip-9023 , xip-366 , and the three homologous XIP-6A , XIP-6B, and XIP-6D [ 33 , 41 , 42 ] have been successfully cloned. Brachypodium, sorghum, maize, and rice genomic data were used to define XIP genomic organization [ 43 ].…”

“…A promising strategy to reduce the negative effect of XIs on dough quality could be the targeted disruption of specific XIs in elite wheat cultivars ( Table 2 ). As an example, the disruption of the three homologous genes, XIP-6A , XIP-6B , and XIP-6D, in the variety “Fielder” by CRISPR/Cas9 significantly increased dough stability time and SDS-sedimentation values of common wheat [ 42 ].…”

Efficient cloning of genes for wheat yield component traits from QTLs via sequencing of the RIL population

“…In addition, many genes could be cloned for other traits using RIL populations. For example, we have cloned a quality gene XIP (Xylanase Inhibitor Protein) (Sun et al, 2022) and a Rht gene TaDHL ( ATP-dependent DNA helicase ) (Guo et al, 2022) from QTLs of the TL-RILs. Seven other Rht genes were also confirmed via CRISPR/Cas9 technology from QTLs of the TL-RILs and “Shannong0431 × Lumai21” RILs (unpublished data).…”

Cloning a novel reduced-height (Rht) gene TaOSCA1.4 from a QTL in wheat

“…This provides a method for gene cloning. Based on this idea, we constructed a map of unigenes using an RIL population and cloned seven yield component trait genes, TaIFABPL, TaDdRp, TaRLK, TaTD, TaTFC3, TaKMT and TaSPL17 ( Zhang et al., 2024 ), a plant height gene TaDHL ( Guo et al., 2022 ), and a quality gene XIP ( Sun et al., 2022 ) from QTLs. This approach could be an effective method for isolating genes originally from wheat.…”