Paper · 2019 · Plant Biotechnology Journal

Development of an Agrobacterium‐delivered CRISPR/Cas9 system for wheat genome editing

Zhengzhi Zhang; Lei Hua; Ajay Gupta; et al.

5
claims
11
verbatim quotes
201
citations
3
research areas
✓
Open access · cc-by

Abstract

Summary CRISPR /Cas9 has been widely used for genome editing in many organisms, including important crops like wheat. Despite the tractability in designing CRISPR /Cas9, efficacy in the application of this powerful genome editing tool also depends on DNA delivery methods. In wheat, the biolistics based transformation is the most used method for delivery of the CRISPR /Cas9 complex. Due to the high frequency of gene silencing associated with co‐transferred plasmid backbone and low edit rate in wheat, a large T 0 transgenic plant population are required for recovery of desired mutations, which poses a bottleneck for many genome editing projects. Here, we report an Agrobacterium ‐delivered CRISPR /Cas9 system in wheat, which includes a wheat codon optimized Cas9 driven by a maize ubiquitin gene promoter and a guide RNA cassette driven by wheat U6 promoters in a single binary vector. Using this CRISPR /Cas9 system, we have developed 68 edit mutants for four grain‐regulatory genes, Ta CKX 2‐1 , Ta GLW 7 , Ta GW 2, and Ta GW 8 , in T 0 , T 1 , and T 2 generation plants at an average edit rate of 10% without detecting off‐target mutations in the most Cas9‐active plants. Homozygous mutations can be recovered from a large population in a single generation. Different from most plant species, deletions over 10 bp are the dominant mutation types in wheat. Plants homozygous of 1160‐bp deletion in Ta CKX 2‐D1 significantly increased grain number per spikelet. In conclusion, our Agrobacterium ‐delivered CRISPR /Cas9 system provides an alternative option for wheat genome editing, which requires a small number of transformation events because CRISPR /Cas9 remains active for novel mutations through generations.

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Claims 5

Method / resourceobservedoriginal resultpositivemeasured

Claim · model-written summary

The Agrobacterium-delivered CRISPR/Cas9 system gave an average edit rate of 10% in wheat.

Outcome
edit ratee2
Value
10%e2

Organisms pictured · Representative pictures of the matched species, not figures from the paper.

Triticum aestivum
Triticum aestivum · TaxonRasbak · CC BY-SA 3.0 · Wikimedia Commons ↗

Verbatim

t0

“for wheat genome editing”

✓ located verbatim in the source · title, characters 61–85 · show in text ↑
e2

“at an average edit rate of 10%”

✓ located verbatim in the source · abstract, characters 1080–1110 · show in text ↑
Method / resourceobservedoriginal resultpositivemeasured

Claim · model-written summary

The system yielded 68 edit mutants in wheat.

Outcome
edit mutantse1
Value
68e1

Organisms pictured · Representative pictures of the matched species, not figures from the paper.

Triticum aestivum
Triticum aestivum · TaxonRasbak · CC BY-SA 3.0 · Wikimedia Commons ↗

Verbatim

t0

“for wheat genome editing”

✓ located verbatim in the source · title, characters 61–85 · show in text ↑
e1

“Using this CRISPR /Cas9 system, we have developed 68 edit mutants”

✓ located verbatim in the source · abstract, characters 893–958 · show in text ↑
Observationobservedoriginal resultpositivemeasured

Claim · model-written summary

Homozygous mutations can be recovered from a large wheat population in a single generation.

Outcome
Homozygous mutationse3

Organisms pictured · Representative pictures of the matched species, not figures from the paper.

Triticum aestivum
Triticum aestivum · TaxonRasbak · CC BY-SA 3.0 · Wikimedia Commons ↗

Verbatim

t0

“for wheat genome editing”

✓ located verbatim in the source · title, characters 61–85 · show in text ↑
e3

“Homozygous mutations can be recovered from a large population in a single generation”

✓ located verbatim in the source · abstract, characters 1182–1266 · show in text ↑
Observationobservedoriginal resultmeasured

Claim · model-written summary

In wheat, deletions over 10 bp are the dominant mutation types.

Outcome
dominant mutation typese4
Qualifier
deletions over 10 bpe4

Organisms pictured · Representative pictures of the matched species, not figures from the paper.

Triticum aestivum
Triticum aestivum · TaxonRasbak · CC BY-SA 3.0 · Wikimedia Commons ↗

Verbatim

t0

“for wheat genome editing”

✓ located verbatim in the source · title, characters 61–85 · show in text ↑
e4

“Different from most plant species, deletions over 10 bp are the dominant mutation types in wheat”

✓ located verbatim in the source · abstract, characters 1268–1364 · show in text ↑
Intervention effectobservedoriginal resultpositivesignificant (as stated)measured

Claim · model-written summary

Homozygous 1160‐bp deletion in Ta CKX 2‐D1 significantly increased grain number per spikelet in wheat.

Treatment
1160‐bp deletion in Ta CKX 2‐D1e5
Gene
Ta CKX 2‐D1e5
Outcome
grain number per spikelete5
Direction
increasede5

Not reportedComparator

Organisms pictured · Representative pictures of the matched species, not figures from the paper.

Triticum aestivum
Triticum aestivum · TaxonRasbak · CC BY-SA 3.0 · Wikimedia Commons ↗

Verbatim

t0

“for wheat genome editing”

✓ located verbatim in the source · title, characters 61–85 · show in text ↑
e5

“Plants homozygous of 1160‐bp deletion in Ta CKX 2‐D1 significantly increased grain number per spikelet”

✓ located verbatim in the source · abstract, characters 1366–1468 · show in text ↑
e6

“important crops like wheat”

✓ located verbatim in the source · abstract, characters 90–116 · show in text ↑

loss of function · yield · Triticum aestivum (wheat)Positive and negative results

43 papers: 18 positive, 25 negative or no effect

17 papers in the same direction as this paper · 25 in the opposite direction

  1. positive
  2. positive
  3. no effect

39 papers more in this group

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Claims are grouped by organism, treatment and what was measured or targeted. A paper counts as positive when any of its claims in the group is positive.