Weed Technology. 2006. Volume 20:576–584

Characterization of Spontaneous Crosses between Clearfield Rice (Oryza sativa) and Red Rice (Oryza sativa)

VINOD K. SHIVRAIN, NILDA R. BURGOS, KAREN A. K. MOLDENHAUER, RONALD W. MCNEW, and TOMILEA L. BALDWIN

Abstract: Experiments were conducted to determine the inheritance of resistance in crosses between imazethapyr-resistant rice and red rice. Past experiments on red rice control, using the Clearfield rice technology, resulted in outcrossing between Clearfield rice and Stuttgart strawhull red rice. The F2 generation of these spontaneous crosses were characterized with respect to inheritance of imazethapyr resistance, leaf color and leaf pubescence, and seed shattering, pubescence, color, and size. Agronomic traits of hybrids were also observed in relation to their parents. To determine the segregation of resistance among F2 phenotypes, the response of three- to four-leaf plants to imazethapyr was scored 3 wk after application as resistant (R, no imazethapyr symptoms), susceptible (S, death of plants), or intermediate (I, stunted plants). R, I, and S phenotypes segregated in a 1:2:1 ratio in the F2 generation. Two- or three-gene inheritance was documented for leaf and seed characteristics. A wide range in onset of flowering (70 to 130 d after planting) was observed in F2 families, although 6% of the plants did not flower during the growing season. F2 plants were taller and had more tillers than any of their parents. Resistance to imazethapyr is associated with a single, incompletely dominant allele. 

Weed Science, 53:567–577. 2005

Mutations in the red rice ALS gene associated with resistance to imazethapyr

Satyendra N. Rajguru Crop, Soil, and Environmental Sciences, 1366 West Altheimer Drive, University of Arkansas, Fayetteville, AR 72704
Nilda R. Burgos Corresponding author. Crop, Soil, and Environmental Sciences, 1366 West Altheimer Drive, University of Arkansas, Fayetteville, AR 72704; nburgos@uark.edu
Vinod K. Shivrain Crop, Soil, and Environmental Sciences, 1366 West Altheimer Drive, University of Arkansas, Fayetteville, AR 72704
James McD. Stewart Crop, Soil, and Environmental Sciences, PTSC115, University of Arkansas, Fayetteville, AR 72701

The introduction of Clearfield (CL) rice cultivars resistant to imidazolinone herbicides, acetolactate synthase (ALS) inhibitors, has raised concerns of gene flow to weedy rice genotypes collectively called ‘‘red rice’’ that infest rice-growing areas in the southern United States. This experiment was conducted to study hybridization between CL rice and red rice using simple sequence repeats (SSR) markers, identify mutations in the ALS gene of imazethapyr-resistant red rice, and to detect the introgression of the ALS-resistant gene from CL rice into red rice. Natural outcrossing experiments between CL rice and strawhull (SH) red rice were set up in Stuttgart, AR, in 2002 and 2003. Putative red rice hybrids were detected among volunteer plants in the following year. Hybridization was confirmed using SSR markers, and introgression of the resistant ALS gene from CL rice to red rice was detected by ALS gene sequencing. The ALS gene sequences of U.S. rice cultivars ‘Bengal’ and ‘Cypress’, SH red rice, CL rice (CL161), and imazethapyr-resistant red rice/CL rice hybrids were compared. Nucleotide sequences of the ALS gene from the rice cultivars were identical. Three point mutations were present in the SH red rice ALS gene coding region relative to Bengal/Cypress. One of these resulted in the substitution of Asp630 for Glu630. The ALS gene sequences of confirmed hybrids were identical to that of the herbicide-resistant pollen source, CL161. We identified four ALS gene mutations in the herbicide-resistant red rice hybrids relative to the susceptible rice cultivars. One point mutation, resulting in a substitution of Ser653 with Asn, was linked to ALS resistance in callus tissue derived from a Kinmaze rice line from Japan. The other three mutations (Ser186—Pro, Lys416—Glu, and Leu662—Pro) are novel. This experiment confirmed that gene flow from imidazolinone-resistant rice resulted in herbicide-resistant red rice plants.

Weed Science 2008 56:485–489

Amino Acid Substitutions in the Acetolactate Synthase Gene of Red Rice (Oryza sativa) Confer Resistance to Imazethapyr

 
Marites A. Sales, Vinod K. Shivrain, Nilda R. Burgos, and Yong I. Kuk

Two red rice accessions from Arkansas have been found to be resistant to the labeled rate of imazethapyr, which is used to control red rice in ClearfieldTM rice. Full-length amplification of the acetolactate synthase (ALS) gene in imazethapyrresistant red rice revealed a coding sequence of 1,935 base pairs, which is the same as that of the cultivated rice. Coding sequences were generated from four red rice accessions collected from different geographical regions in Arkansas, consisting of accessions that were either resistant or susceptible to imazethapyr. Nucleotide sequence alignments identified six base polymorphisms, three of which resulted in amino acid substitutions in the ALS gene. One amino acid substitution, Gly654Glu, involves a residue required for imazethapyr binding to the ALS. The other substitution, Val669Met, implies conformational changes in the ALS structure that enhances binding of thiamine diphosphate, an ALS cofactor. These novel amino acid substitutions first reported for ALS-resistant red rice accessions support the hypothesis that ALS-resistant red rice can evolve with sustained herbicide selection pressure. Thus, it behooves growers to integrate the Cleafield rice technology with other tools to achieve a successful, long-term weed management program.

Weed Science 2008 56:1–11

Natural Tolerance to Imazethapyr in Red Rice (Oryza sativa)

Yong I. Kuk, Nilda R. Burgos, and Vinod K. Shivrain

Red rice is a major weed problem in rice production of the southern United States and other rice-producing countries. One hundred thirty red rice accessions from 26 rice-growing counties in Arkansas were tested for tolerance to imazethapyr in seed- and whole-plant response bioassays. The red rice accessions were compared with imazethapyr-resistant (ClearfieldTM) rice cultivars (‘CL121’, ‘CL161’, and ‘CL-XL8’) and conventional rice cultivars (‘Bengal’, ‘Dongjin’, ‘Drew’, and ‘Wells’). Red rice accessions 79, 84, and 118 showed 17-, 48-, and 37-fold more tolerance to imazethapyr, respectively, than the standard susceptible red rice accession (82) in whole-plant bioassays. The imazethapyr-resistant rice cultivars, CL121, CL161, and CL-XL8 were 41-, .177-, and 48-fold more resistant to imazethapyr, respectively than the susceptible standard. The imazethapyr-tolerant red rice and ClearfieldTM cultivars were generally cross tolerant to other acetolactate synthase (ALS; EC 4.1.3.18) inhibiting herbicides such as imazapyr, imazaquin, imazamox, and pyrithiobac. The tolerance level of red rice or rice to imidazolinone herbicides was highest with imazaquin and lowest with imazapyr. The imazethapyr-tolerant red rice accessions and ClearfieldTM rice were susceptible to glufosinate and glyphosate. The ALS enzyme of tolerant red rice accessions was less sensitive to imazethapyr than the susceptible standard, but tolerance at the enzyme level was less than at the whole-plant level. Therefore, tolerance of red rice to imazethapyr may involve other mechanisms besides an insensitive target site. We learned that a few imazethapyr-tolerant red rice populations existed probably before ClearfieldTM rice was introduced, supporting the hypothesis that evolution of herbicide-resistant red rice populations can happen with intensive herbicide selection pressure.