Progress 02/28/07 to 03/01/12
Outputs
OUTPUTS: The final report focuses on Year 2011 and earlier portion of 2012 activities. The original main goal of the study was to molecularly profile individual organs of the peanut plant. This goal was lastly adjusted during the Year 2011 to maximize the research benefits by taking advantage of a latest most efficient sequencing technology, "Next Generation Sequencing" - which is about a million times more efficient than the standard "Sanger Sequencing". For broader impact, instead of separate molecular profiling of individual organs, which originally would have produced a limited impact due to limited number of genes that were planned for sequencing due to cost and limited efficiencies of "First Generation Sequencing" that was popular at the inception of the project, we decided to better the outcomes by molecularly profiling the peanut whole plant including all organs. This gave us an unprecedented opportunity to fully sequence all expressed genes we have captured, which we did. During this period, we successfully sequenced 8,308,655,800 nucleotides from the peanut Spanish botanic type whole plant normalized cDNA library. The sequencing quality scores of Q30 (99.9% base call accuracy) were achieved across all bases in forward direction sequencing and from 1 to 84 bases in reverse direction sequencing. De novo bioinformatics assembly of all sequencing data by the SOAP2 program generated 26,048 unigenes of the average sequence length of 550 bp and sequence length range of 201 to 2,431 bp, which represent the total sequence length of 14,305,500 bp of the peanut genome. All assembled unigene sequences were confirmed by sequence similarity comparisons against NCBI NR nucleotide and protein databases. The functional annotation of all unigenes showed that 12,302 and 8,817 unigenes matched the records in SWISS-PROT database (version 20120304) and COG database, respectively. Further proceeding showed that the analysis of COG's classification of all known function unigenes resulted in categorizing them into 23 clusters based on their common functions, while the functions of 209 genes remained unknown. Overall, there still were 13,746 (52.77%) unigenes that had no significant match with current records in any currently applicable databases. These activities were also used for year-round laboratory hands-on research experience of three undergraduate students listed under participants. Thirteen high school graduates were provided with pre-college science laboratory skills and experiences during the University summer camp. A peer-reviewed research manuscript is now under review of a peer-reviewed research journal for results dissemination. PARTICIPANTS: Main participants in this project during year 2011/2012 included Langston University faculty, Dr. Kanyand Matand (PI) and Dr. Ning Wu (Co-PI), and three undergraduate students - Kayla Love, Rajah Singh, and Mele Miro. TARGET AUDIENCES: The communities targeted include peanut and other legumes genomic research scientists, and Langston University science-major students and other underrepresented communities for biotechnology and science education. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.
Impacts
This project has developed the first reference transcriptome and unigene database, with a significant resource of 26,048 individual records, from the peanut Spanish botanical group whole plant. It is the first, fully-sequenced expressed-genome of the whole peanut plant, in general, and Spanish type, in particular with potentially tremendous research implications. The success of the present activities have enhanced the global image of biotechnology research program of Langston University, such that in the month of September 2012 this program was a major highlight during the visit of OSU/A&M Board of Regents Chair and delegates at Langston University. The biotechnology activities also received a special highlight during the Langston University Administrative General Council Meeting of the same month. Except one, all high school graduates, who participated in summer biotechnology training via Langston University summer camp, adopted Langston University for their education and sciences for their major. Increasing number of undergraduate students, more than we can accommodate, are currently requesting for the opportunities to participate in biotechnology research experiences through the present activities. The success of this project has enabled current subsequent similar research on the other three peanut cultivated groups (Virginia, Valencia, and Runner) with CRIS funding. This success has also enabled new collaborative research opportunities with New Mexico State University, with which a brainstorming meeting was held in Texas on potential avenues we should adopt in resolving pathological challenges associated with Peanut Leaf Spot Disease, also in last September.
Publications
- Wu N, Matand K, Wu H, Li B, Li Y, Zhang X, He Z, Qian J, Conley S*, Bailey M*, Acquaah G. 2012. De novo next generation sequencing, assembling and annotation of Arachis hypogaea L. Spanish botanical type whole plant transcriptome. Theoretical & Appl.Genetics (under review).
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Progress 01/01/09 to 12/31/09
Outputs
OUTPUTS: During 2009, the normalized cDNA library, which was constructed in the previous year was used for colony picks. Overall, 20,000 clones were picked. Those clones were further amplified using PCR method, prior to subsequent purification process. This is the collection that will be used to produce microarray chips for screening. These activities were used for year-round laboratory hands-on research experience of three undergraduate students listed under participants. Thirteen 4-H students and 28 high school graduates were also provided with pre-college science laboratory skills and experience during summer camps. The results were disseminated through 4 conference presentations and a peer-reviewed publication of the peanut library subtraction protocol we developed. PARTICIPANTS: The permanent participants in this project during this year included Langston University faculty, Dr. Kanyand Matand (PI) and Dr. Ning Wu (Co-PI), and three undergraduate students - Antwon Watson, Stephan Conley, and Sharhonda Pickett. TARGET AUDIENCES: The communities targeted include peanut and other legumes genomic research scientists, and Langston University science-major students and other underrepresented communities for biotechnology education. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.
Impacts
The results of the studies were shared at the conferences with many other participants of the scientific community. 98% of high school graduates, who participated in summer biotechnology training via Langston University summer camp, adopted Langston University for their education and sciences for their major and other programs, including, the Departments of Biology and Chemistry, and Honors Program have not only been extended but also expanded. Because of our success and improved biotechnology image of Langston University, the biotechnology research collaboration with China has been expanded to include both research and outreach to faculty and students. Increased number of students are currently requesting for the opportunities to participate in biotechnology research experience through the present activities. An extended study on developing 'a more efficient method for providing underrepresented students with research experience' has been derived from the project's activities and is very promising.
Publications
- Wu, Ning, Matand, K, and Williams, S. 2009. A novel mRNA level subtraction method for quick identification of target-oriented uniquely expressed genes between peanut immature pod and leaf. Biological procedure.online.D01:10.1007/S12575-009-9022-Z.
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Progress 01/01/08 to 12/31/08
Outputs
OUTPUTS: Activities during this year focused on re-developing primary experimental biological materials from the Spanish peanut variety, Blanco, because all those that were initially developed during the previous year were lost due to holiday's blackouts. During this period, peanut seeds were cultured in the greenhouse to maturity (three-month cycle); and the whole plant tissues were collected for isolating and purifying total RNA and mRNA, respectively, for subsequent primary cDNA library construction. The resulting high quality primary cDNA library met the prerequisite quality control criteria, which included (1) high recombinant rate (>85%), (2) high library clone capacity (>1 million colony forming unites), and (3) satisfactory average clone size (400 bps). This library is a gene pool for all peanut plant organs and essential for subsequent studies. The library was normalized to streamline huge number of repetitive genes and maximized gene discoveries and cost-efficiency. During the same period, three Langston University students, 15 4-H students, and 35 Oklahoma high school graduates benefited from hands-on research experience. The experimental results were presented at a scientific meeting. PARTICIPANTS: The permanent participants in this project during this year included Langston University faculty, Dr. Kanyand Matand (PI), Dr. Ning Wu (Co-PI), and Dr. George Acquaah (Co-PI) and one undergraduate student, Kariel Ross. TARGET AUDIENCES: The communities targeted include peanut and other legumes genomic research scientists, and Langston University science-major students and other underrepresented communities for biotechnology education. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.
Impacts
The results of the studies were shared at the conference with many other participants of the scientific community. 100% of high school graduates, who participated in summer biotechnology training via Langston University summer camp, adopted Langston University for their education and sciences for their major. Other Langston University programs (Departments of Biology and Chemistry, and Honors Program) recommended students for hands-on research experience in biotechnology field. Both PI and Co-PI received Langston University's outstanding research awards based on their biotechnology activities through this project. Langston University, which is known primarily for its goat program, improved its new biotechnology image. As a result, new biotechnology research collaboration was established with China.
Publications
- No publications reported this period
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Progress 01/01/07 to 12/31/07
Outputs
OUTPUTS: Activities conducted during 2007 period focused on groundwork leading to the development of peanut growth and development specific gene microarray screenings. Peanut were cultured in the greenhouse to maturity (for 3 months) and tissue-sampled at younger stage (seedling with cotyledonary leaves), young plant (stage prior to flowering), and mature stage (including plants with root, stem, leaf, pod - "stages R1, R2, and R3", flower, and peg tissues). All targeted tissues were harvested at their proposed stages and stored for long-term usage for the project. Total RNA and mRNA were purified from the whole peanut mature plant tissues of a Spanish group, Blanco, for subsequent development of the primary cDNA library. This step is foundational to subsequent project activities and critical to the overall success of the project. High quality whole mature primary cDNA library was constructed and had satisfied our prerequisite quality control criteria, that included (1) high
recombinant rate (>85%), (2) high library clone capacity (>106 colony forming unites), and (3) satisfactory average clone size (>400 bps). This library is a gene pool for all peanut plant organs listed earlier; and is essential in subsequent microarray chip development and subsequent high-throughput gene screening. This is also a pioneering library that includes genes of different peanut plant organs and developmental stages within the peanut community.
PARTICIPANTS: All activities during the 2007 period were conducted in the Biotechnology Program of Langston University. Thus, permanent participants included Langston University faculty, including Dr. Kanyand Matand (PI), Dr. Ning Wu (Co-PI), and Dr. George Acquaah (Co-PI). Three undergraduate students, who participated in these activities, include Kariel Ross, Wondwessen Kebede, and Kimberley Jones.
TARGET AUDIENCES: The communities targeted include peanut and other legumes genomic research scientists, and Langston University science-major students.
Impacts
These studies were designed in response to the general appeal of the 'Cross-Legume Advances Through Genomics (CATG)' to individual legume species scientists to focus their research on areas of common interests to legume genomics as reflected in the general guidelines for research platform that were initially developed in 2004 in Santa Fe, NM (CATG, 2004). Related general guidelines, clearly stressed the importance of continuous development of ESTs from peanut, and the importance of better understanding the legume molecular biology and physiology to enable legume researchers to answer long-term key questions such as (1) what makes a legume legume? (2) What are the genes that are involved in the biology of legume seeds? (3) What are the genes that are involved in the biology of nitrogen fixation? (4) What are the genes that are involved in the overall biology of the legume plant?, etc. This cDNA library that was developed represents a pioneering collection of genes from
different organs and developmental stages of the peanut plant that could be further screened for genes that could contribute to research efforts to finding answers to some of those outstanding questions. Further, this collected unique gene pool is foundational to the enhancing of one aspect of the peanut genomic studies that has for so long been overlooked, 'the biology of peanut plant growth and development including its organs, per se'. It could also enable the isolation of targeted full length genes and further related gene interaction and expression studies that could play significant roles in designing new superior varieties for the twenty-first century that are high-yielding, stress tolerant, and of high-nutritional qualities with photosynthetic efficiency, and efficient flowering, fruiting, rooting and nutrients accumulation. This gene collection will be used to generate microarray chips and ESTs for this project.
Publications
- No publications reported this period
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