Progress 09/01/08 to 08/31/12
Outputs
Target Audience: For the period from 09/01/2011 to 08/31/2012: 1) Trained the Native America undergraduate student Ashley LaCayo for marker genotyping, data recording and collection from greenhosue plant populations, and germinaiton tests; and 2) Trained six high school students through hosting the 2012 plant molecular biology workshop in June, 2012. Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided?
Nothing Reported
How have the results been disseminated to communities of interest? Data from this project were published in five papers and 11 abstracts, presented in 10 national/international meetings and about 10 invited lectures/seminars, and contributed to the PI's classes "Crop physiology", "Genome mapping and QTL analysis", and "Molecular plant physiology", and to the Plant Molecular Bioogy Workshops for High Shcool Students or Teachers. Some plant materials were shown on campus tours for local farmers and visiotrs and advanced genetics/biotechnology classes. What do you plan to do during the next reporting period to accomplish the goals?
Nothing Reported
Impacts
What was accomplished under these goals?
Rare recombinants for the qSD7-1/qPC7 region were selected, inlcuding one recombinant that contains only one DNA segment of about 2000 base pairs within the Rc gene from the parental line of weedy red rice. This intergenic resombinant provided unambiguous evidence that the Rc transcription factor controls both seed dormancy and red pericarp color in rice. This was the first time to demonstrate that the association between seed dormancy and grain color in cereal crops arises from pleiotropic effects of the gene SD7-1/Rc. A pair of perfect isogenic lines for the dormancy and non-dormancy alleles was developed from the intergenic recombinant. Data collected from the perfect isogenic lines demonstrated that the pleiotropic gene SD7-1/Rc enahnced the biosynthesis of the plant hormone abscisic acid in early developing seeds to induce the primary seed dormancy and also activated the eight genes of the flavonoid biosynthesis pathway in the lower epidermal cell layer of the pericapr tissue to produce red pigments. The SD7-1 seed dormancy gene did not respond to the treatment of the plant hormone gibberrelic acid in germinaiton. The phylogenetic and QTL analysis confirmed that the SD7-1 dormancy gene has differentiated into two groups corresponding to tropical and temperate ecotypes, respectively. The transformation and the following hybridization experiments revealed that the phenotypic effect of the SD7-1 dormancy gene was affected by genetic backgrounds.
Publications
- Type:
Journal Articles
Status:
Awaiting Publication
Year Published:
2013
Citation:
Mispan, M. S., L. Zhang, J. Feng, and X.-Y. Gu. 2013. Quantitative trait locus and haplotype analyses of wild and crop-mimic traits in U.S.
weedy rice. G3 (in press).
- Type:
Journal Articles
Status:
Published
Year Published:
2013
Citation:
Ye, H., D. H. Beighley, J. Feng, and X.-Y. Gu. 2013. Genetic and physiological characterization of two clusters of quantitative trait loci associated with seed dormancy and plant height in rice. G3 3:323-331.
- Type:
Journal Articles
Status:
Published
Year Published:
2011
Citation:
Gu, X.-Y., M. E. Foley, D. P. Horvath, J. V. Anderson, J. Feng, L. Zhang, C. R. Mowry, H. Ye, J. C. Suttle, K. Kadowaki, and Z. Chen. 2011.
Association between seed dormancy and pericarp color is controlled by a pleiotropic gene that regulates ABA and flavonoid synthesis in
weedy red rice. Genetics 189:1515-1524.
- Type:
Journal Articles
Status:
Published
Year Published:
2010
Citation:
Ye, H., M. E. Foley, and X.-Y. Gu. 2010. New seed dormancy loci detected from weedy rice-derived advanced populations with major QTL
alleles removed from the background. Plant Science 179:612-619.
- Type:
Journal Articles
Status:
Published
Year Published:
2008
Citation:
Gu, X.-Y., E.B. Turnipseed, and M.E. Foley. 2008. The qSD12 locus controls offspring tissue-imposed seed dormancy in rice. Genetics
179:2263-2273.
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2012
Citation:
Mispan M. S., and X.-Y. Gu. 2012. QTL and haplotype analysis of wild and crop-mimic traits in US weedy rice (abstract). The 6th International
Weed Science Congress. Hangzhou, China. June, 2012.
- Type:
Journal Articles
Status:
Published
Year Published:
2011
Citation:
Gu X.-Y., J. Zhang, J. Feng, and H. Ye. 2011. Genetic and molecular characterization of genes controlling seed dormancy through the
maternal, embryo, and endosperm tissues in rice (abstract). INFORMATIVO ABSTRACTES 21(1):196.
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2010
Citation:
Gu, X.-Y., L. Zhang, J. Feng, and M. E. Foley. Comparative genetics of seed dormancy between tropical and temperate weedy rice(abstract).
Seed Ecology III. Salt Lake City, UT. June, 2010.
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2010
Citation:
Gu, X.-Y., J. Feng, and M. E. Foley. 2010. Seed dormancy genes cloned from weedy rice and their applications (abstract). The 33rd Rice
Technology Group Meeting, Biloxi, MS. Feb., 2010.
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2009
Citation:
Gu, X.-Y., H. Ye, T. Liu., J.V. Anderson, D.P. Horvath, J.C. Suttle, and M.E. Foley. 2009. Seed dormancy of rice: from natural variation to
genes (abstract). The 4th International Plant Dormancy Symposium, Fargo, ND. June, 2009.
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2009
Citation:
Ye, H., and X.-Y. Gu. 2009. Seed dormancy QTLs detected from weedy rice-derived advanced backcross populations (abstract). The 4th
International Plant Dormancy Symposium, Fargo, ND. June, 2009.
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2009
Citation:
Gu, X.-Y., M. E. Foley, J. V. Anderson, and D. P. Horvath. 2009. Positional cloning and characterization of the qSD7-1 underlying gene in
rice(abstract). 2009 USDA-NRI Project Directors meeting. Bethesda, MD. May, 2009.
- Type:
Journal Articles
Status:
Published
Year Published:
2008
Citation:
Gu, X.-Y., M. E. Foley, J. V. Anderson, and D. P. Horvath. 2008. qSD7-1 is the first dormancy QTL cloned from weedy rice (abstract). Polish J.
Natural Sci. suppl. 5:51.
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2008
Citation:
Gu, X.-Y., J.-Q. Lou, and M. E. Foley. 2008. Genetics of weedy rice traits: seed dormancy and red pericarp color (abstract). The 5th
International Weed Science Congress, Vancouver, Canada. Jun, 2008. Anderson, J. V., M. E. Foley, D. P. Horvath, and X.-Y. Gu. 2008.
Dormancy regulation in reproductive structures of weedy plants; a comparison between seeds and vegetative buds(abstract). Sleeping
Beauties - Dormancy and resistance in harsh environments - Molecular, proteomic and metabolomic aspects, Berlin. May, 2008.
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2008
Citation:
Foley, M. E., X.-Y. Gu, J. V. Anderson, and D. P. Horvath. 2008. Positional cloning of a seed dormancy QTL from weedy rice
(Oryza sativa)(abstract). Plant & Aminal Genome XVI. San Diego, CA. Jan., 2008.
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Progress 09/01/11 to 08/31/12
Outputs
Target Audience:
Nothing Reported
Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided? Advanced transgene and bioinformaticanalyses and molecular biological experiments wereused to train 2 postgraduate researchers and 2 graduate students with the project, How have the results been disseminated to communities of interest? Publication in peer-reviewed journals and presented in national and internaitonal conferences and seminars. What do you plan to do during the next reporting period to accomplish the goals?
Nothing Reported
Impacts
What was accomplished under these goals?
Completed map-based cloning and molecualr characterization of the qSD7-1 seed dormancy QTL with a high quality, and received anew research grant to extendthe cloned seed dormancy gene to a practical applicaiton bydeveloping a novel strategy to reduce the risk oftransgene flow from cultivatedinto weedy rice.
Publications
- Type:
Journal Articles
Status:
Published
Year Published:
2013
Citation:
Gu, X.-Y., M. E. Foley, D. P. Horvath, J. V. Anderson, J. Feng, L. Zhang, C. R. Mowry, H. Ye, J. C. Suttle, K. Kadowaki, Z. Chen. 2011. Association between seed dormancy and pericarp color is controlled by a pleiotropic gene that regulates ABA and flavonoid synthesis in weedy red rice. Genetics 189:1515-1524
Mispan, M. S., L. Zhang, J. Feng, X.-Y. Gu. 2013. Quantitative trait locus and haplotype analyses of wild and crop-mimic traits in U.S. weedy rice. G3: Genes, Genomes, Genetics 3:1049-1059.
Ye, H., D. H. Beighley, J. Feng, X.-Y. Gu. 2013. Genetic and physiological characterization of two clusters of quantitative trait loci associated with seed dormancy and plant height in rice. G3: Genes, Genomes, Genetics 3:323-331.
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Progress 09/01/10 to 08/31/11
Outputs
OUTPUTS: 1) Seeds harvsted from about 1500 plants from the greenhouse and field experiments in summer 2010 were evaluated for seed dormancy by germinaiton testes; analysis was conducted to correlate the germination data with the marker genotypifn data. 2) Forteen new lines of weedy and wild rice lines were sequenced for the SD7-1/Rc alleles to understand the relation betweeen the allelic differentitation and the degree of seed dormancy. 3) A library of cDNAs for genes expressed in developing seeds from the isogenic line for the SD7-1 dormancy allele was developed for Y2H experiments; the library was screened using three probes developed from difference sections of the SD7-1 full-length cDNA sequence; and posstive clones were sequenced and sequences analyzed. 4) Recurrent backcross with marker-assisted selection was conducted to advance the transgene SD7-1/Rc-derived BC4F2 popluaitons; selected lines were grown in greenhouse; and seeds harvested for dormancy assay. And 5) Two sets of segregating populations were advanced from F3 to F4 generations, which combined the SD7-1 dormancy alleles from the tropic weedy rice SS18-2, the temperate weedy rice line LD, and wild rice O. rufipogn, to synchronized the genetic background. About 400 plants from two of these advanced segregation populations were grown in field plots in summer 2011. PARTICIPANTS: 1). Dr. Xinli Sun, a postdoctoral research associate worked full time on the project and developed expertise of gene expression analyses at both mRNA and protein (Y2H)levels. 2) Mr. Chase Mowry, a gradaute research assitant, worked on the project with 49.5% of the time effort. The GRA is preparing his thesis with data collected from some of the above-described experiments. 3) Dr. Jiuhuan Feng, research associate II, worked on the project and was reponsible for sequencing additonal SD7-1/Rc alleles. 4) The GRAs Shakirin Mispan and Lihua Zhang worked part-time on this project in the experiments such as field and greenhosue planting, harvesting, and germination tests. 5) Alexentra Chase, a undergradaute research assistant, worked partly on this project in assisting with marker genotyping and germination test. TARGET AUDIENCES: 1) Trained 3 undergradaue research assistants at SDSU for molecualar and seed biology. 2) Provided a lab tour for 12 students in the Advanced Genetics and Biotechnology class to introduce techniques from marker genotyping,map-based cloning, to trnasgenic analysis. 3) Provided job-shadowing for high school students to get experince in molecular biotechnologies.and, 4) Provided experimental data for real-data practices and lab tour for a new graduate class "QTL analysis and Applications". PROJECT MODIFICATIONS: Not relevant to this project.
Impacts
1) The SD7-1/Rc transgene was retained in all segregation populations planted from the 2010 experiments, as indicated by marker genotyping data and phenotypic data for pericarp color; these populations were also segregating for other traits such as flowering time and plant height, which likely affect detection of the dormancy gene by germination. One of the population displayed significant correlation between pericarp color and seed dormancy, suggested the transgene had an effect on seed dormancy or delaying germination. This results will be confirmed using the advanced populaitons grownn in 2011. 2) A phylogenetic analysis revealed that the SD7-1/Rc alleles from weedy rice differentiated into two clades, which corresponds to the tropic and temperate ecotypes. 3)There was no significant correlation between SD7-1-based markers and see dormancy in the 2010 populations segregating for the SD7-1/Rc alleles of different geographic origins. This suggests that the point mutations outside the 14-by deletion, which causes a functinal muation from red to white pericarp color, may not cause a detectable effect on germinaiton. This result will also be confirmed with the advanced populations harvested in 2011. 4) The cDNA library contains ~1.3 million clones. One of SD7-1 probes was detected to be autoactiviation by a bait quality check. Two-hybrid screening using yeast mating detected several hundreds of postive clones. Bioinformationtics analysis for sequences of selected clones detected candiate genes for on-going experiments. 5) The most advanced generation of populations segregating for the transgene or the SD7-1/Rc alleles from weedy and wild rice displayed synchronized background and are subject to germination test.
Publications
- Gu, X.-Y., M. E. Foley, D. P. Horvath, J. V. Anderson, J. Feng, L. Zhang, C. R. Mowry, H. Ye, J. C. Suttle, K. Kadowaki, Z. Chen. 2011. Association between seed dormancy and pericarp color is controlled by a pleiotropic gene that regulates ABA and flavonoid synthesis in weedy red rice. Genetics (in press)
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Progress 09/01/09 to 08/31/10
Outputs
OUTPUTS: New populations of plants were developed for: 1) the T6 generation of transgenic SD7-1/Rc lines with the japonica cultivar Nipponbare as the background; 2)BC2F2s with the indica line EM93-1 as the recurrent background; 3)F3 generation of populations derived from the F2 plants heterozygous for SD7-1/Rc alleles from the tropic weedy line SS18-2 and the temperate weedy line LD, and SS18-2 and the wild rice Oryza rufipogon; and 4) BC3F1 populations with EM93-1 as the recurrent parent. Seed dormancy was evaluated for the above-stated three categories of populations grown in greenhouse or field conditions. Microarray data were reanalyzed and some candidate genes from the analysis were confirmed using qRT-PCR with newly extracted mRNA from developing seeds from isogenic lines. QTL analysis of seed dormancy was conducted for two populations derived from two weedy rice lines from China and USA, respectively. The SD7-1/Rc allele was sequenced from 10 lines of weedy red rice different in ecotypes. A histological experiment was conducted to determine the expression of the Rc gene in seed component tissues. An yeast-two-hybridization experiment was initiated to identify the protein complex involving the SD7-1 bHLH. Information from this project was presented in two national meetings, and one international meeting. PARTICIPANTS: Xing-You Gu: PD, designed hybridization, greenhouse/field experiments, and germination experiments; gene expression data analysis; preparing manuscripts developed based on the results from this project. Lihua Zhang: Graduate Research Assistant (GRA) (PhD level), developed advanced transgenic lines and backcross populations; marker genotyping and dormancy evaluation; microarray data analysis; QTL analysis of seed dormancy. Chase R. Mowry: GRA (master level): mapped the transgene insertion locus; field experiment in Southeast Missouri; population advancement and marker genotyping; dormancy evaluation. Heng Ye: GRA (PhD level); qRT-PCR analysis to confirm candidate genes selected from the microarray analysis. Muhamad Shakirin Mispan (PhD level): QTL analysis of seed dormancy and its related traits based on a segregation population derived from a US weedy red rice. Jiuhuan Feng (lab manager), coordinated lab, greenhouse, and field experiments; marker genotyping and dormancy evaluation; screened a BAC library using the SD7-1 genome sequence as probes; histological analysis of expression of the Rc gene in seed component tissues. Xinli Sun: postdoctoral research associate: development of the SD7-1 RNAi construct; development of a yeast library to initiate an Y2H experiment. TARGET AUDIENCES: Provided internship for two undergraduate students in summer 2010; provided work study opportunities for 4 students at SDSU. PROJECT MODIFICATIONS: Reccurent backcross was used to purify the genetic background to detect the effect of transgene on seed dormancy.
Impacts
A significant correlation between transgenic SD7-1/Rc and seed dormancy was detected in the transgenic T3 line-derived BC2F2 populations, suggesting that the Rc locus is one of seed dormancy gene in weedy rice. However, there was no genetic difference in seed dormancy between the red- and white-pericarp colored T6 transgenic lines, strongly suggesting that the effect of SD7-1/Rc on seed dormancy is affected by the genetic background. The insertion locus of the transgenic SD7-1/Rc was mapped onto the long arm of chromosome 1 and is about 5 centiMorgan away from the qSD7-2 seed dormancy QTL detected in the PI's lab. Correlation between the SD7-1 alleles and seed dormancy was also not significant in the F3 populations. A total of 14 "wild alleles" at SD7-1/Rc from different geographic areas were clustered into two distinct clades. The phylogenetic and linkage analysis indicate the Rc alleles from the weedy red rice lines in East Asia (East China and Korea) are greatly different from the Rc alleles in wild rice and tropic red rice lines at the DNA level; however, the differentiation between the red-pericarp-colored alleles did not result in a large phenotypic effect on seed dormancy. Both the microarray and qRT-PCR analyses revealed the SD7-1/Rc transcription factor up-regulate seven genes on the flavonoid biosynthetic pathway to control the pigment in the lower epidermal cell layer of the pericarp tissue.
Publications
- Ye, H., M. E. Foley, X.-Y. Gu. 2010. New seed dormancy loci detected from weedy rice-derived advanced populations with major QTL alleles removed from the background. Plant Science 179:612-619.
- Gu, X.-Y., L. Zhang, J. Feng, M. E. Foley. Comparative genetics of seed dormancy between tropical and temperate weedy rice. Seed Ecology III. Salt Lake City, UT, June 20-24, 2010.
- Gu, X.-Y., J. Feng, M. E. Foley. 2010. Seed dormancy genes cloned from weedy rice and their applications. The 33rd Rice Technology Group Meeting, Biloxi, MS, Feb. 21-25, 2010.
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Progress 09/01/08 to 08/31/09
Outputs
OUTPUTS: SD7-1/Rc transgenic lines were advanced from T3 to T5 generations. Six lines from the T5 plants with a relatively high seed set rate are used to develop the T6 generation to confirm the effect of the transgene on seed dormancy. Selected T3 plants were crossed with isogenic lines for SD7-1 dormancy and non-dormancy alleles, and backcross BC1F1 populations are used to examine positional and dosage effects of the SD7-1 seed dormancy gene on germination. Crosses were made between lines with the SD7-1/Rc alleles for the tropic and temperate weedy rice lines and the introgression line derived from the wild rice Oryza rufipogon; the F2 populations are grown in greenhouse to detect Rc alleleic and dosage effects on seed dormancy. Gene expression data from Affy. 57K microarray analysis were analyzed with multiple sets of software, including Pathway Studio, to identify physiological pathways regulated by the seed dormancy gene; about 50 selected candidates were confirmed for expression levels. The SD7-1 seed dormancy gene was detected for germination response to gibberellic acid. The SD7-1/Rc transgenic T5 lines were detected for accumulation patern of the hormone abscisic acid in developing seeds and degree of seed dormancy. Available information from this project was presented in three international and one national meetings and in the seminars at three universities. PARTICIPANTS: South Dakota State University: 1) Gu (PI): experimental designs, data analysis and presentations of research results in international and national meetings; and research associates/assistants identification and mentoring. 2) Munshi (postdoctoral research associate): expression analysis of the seed dormancy gene in seed tissues; high generations of transgenic line development; and preliminary research on effect of the dormancy gene (transcription factor) on saline stress. 3) & 4) Zhang & Lou (two graduate research assistants): hybridizations, segregation population development; marker genotyping and germination assessment, and data analysis. 5) & 6) Carsrud & Li (two technicians): management of greenhouse populations and germination assessment. 7) Ge (collaborator): bioinformatics analysis of microarray data. USDA -ARS, Fargo, ND: Suttle (collaborator): seed ABA content detection. Horvath (collaborator): microarray data annotation. Cornell University: McCouch (collaborator): development of introgression lines for the Rc allele from wild rice. TARGET AUDIENCES: Seed scientists and workers, researchers in plant dormancy areas, students from the departments of plant science and biology, plant breeders and farmers. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.
Impacts
T5 transgenic lines with seed set rate significantly improved were obtained and the resulting BC1F1 population segregating for the transgene were developed. The lines and population are important materials to confirm the function of the transgene on seed dormancy and germination. The F2 populations segregating for different SD7-1/Rc alleles were obtained. The qSD7-1 seed dormancy QTL was detected from the tropic weedy rice line SS18-2, and was not detected in three reported wild rice lines. Thus, the obtained populations should be valuable to determine if the aforementioned difference arose from allelic differentiation or genetic background effects. Some physiological pathways downstream to the SD7-1/Rc genes were developed or suggested. For example, the SD7-1/Rc gene up-regulates five genes encoding enzymes on the proanthocyanidin biosynthesis pathway, which explains the effect of Rc gene on red pericarp color. The SD7-1 gene promotes accumulation of abscisic acid (ABA) in developing seeds from our weedy rice-derived isogenic line, but no in the transgenic lines in one preliminary experiment. This suggests that genetic background or allelic differentiation may affect the function of the dormancy gene on seed dormancy by modifying ABA metabolism.
Publications
- Gu, X.-Y., M. E. Foley, J. V. Anderson, D. P. Horvath. 2009. Positional cloning and characterization of the qSD7-1 underlying gene in rice. 2009 USDA-NRI Project Directors meeting, Bethesda, MD, May,2009.
- Gu, X.-Y., M. E. Foley, J. V. Anderson, D. P. Horvath. 2008. qSD7-1 is the first dormancy QTL cloned from weedy rice. Polish J. Natural Sci. suppl (5:51).
- Gu, X.-Y., J.-Q. Lou, M. E. Foley. 2008. Genetics of weedy rice traits: seed dormancy and red pericarp color. The 5th International Weed Science Congress, Vancouver, Canada, June, 2008.
- Gu, X.-Y., H. Ye, T. Liu., J.V. Anderson, D.P. Horvath, J.C. Suttle, M.E. Foley. 2009. Seed dormancy of rice: from natural variation to genes. The 4th International Plant Dormancy Symposium, Fargo, ND, June, 2009.
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