Team:RIS BKK/Description
Project
Research Paper
https://static.igem.org/mediawiki/2019/8/87/T--RIS_BKK--img--research_paper.pdf
Thailand is an agriculture-based country. However, Thai farmers face economic hardships. The aim is to reduce the input cost by developing transgenic bacteria that will help plants fix nitrogen without the application of chemical fertilizers. We aim to synthesize Agrobacterium that is able to infect plants with inserted Nif genes from Azospirillum brasilense ATCC 29711 (A. brasilense). By using Agrobacterium’s ability to infect and transfer its DNA, we hope to allow plants to independently fix nitrogen. Nif genes were extracted from Azospirillum brasilense, transferred into E.coli as part of 2 vectors, pGEM and pCAMBIA, and put into Agrobacterium. Our design allows for the creation of a product such as biofertilizers or a gel-like-substance mixed with Agrobacterium which should increase the yield of crops. Although we were not able to transform our transgenic Agrobacterium into a product due to time constraints and other limitations we still hope that others will continue this in the future.
Inspirations
“…เมืองไทยนี้ต้องพึ่งเกษตรกรเป็นสำคัญ เพราะว่าเกษตรกรเป็นประชาชนส่วนใหญ่ของประเทศและต้องยึดอาชีพนี้มาและไม่ใช่เพราะเหตุนั่นเท่านั้นเอง แต่ว่าประเทศหนึ่งประเทศใดจะอยู่ได้ก็เพราะว่ามีกสิกรรม การประกอบอาชีพ ในด้านผลิตผลที่ได้จากธรรมชาติ
ทั้งในด้านที่จะเป็นการปลูกข้าว ปลูกพืชไร่ ปลูกผลไม้ หรือทำมาหากินในด้านปศุสัตว์หรือประมง…”
ความตอนหนึ่ง ในพระราชดำรัส
ในโอกาสที่คณะกรรมการสหกรณ์การเกษตร สหกรณ์นิคม สหกรณ์ประมง และสมาชิกผู้รับนมสดเข้าเฝ้าฯ
ณ โครงการส่วนพระองค์ สวนจิตรลดา
วันที่ ๘ พฤษภาคม ๒๕๓๐ [1]
“…Thailand relies significantly on agriculture because most of the Thai population are farmers. We have always been involved in Thai agricultural production from generation to generation. Not only that, but for a country to be able to survive, there must be some field of agriculture; jobs that involve products of nature such as growing rice or fruits, or livings involving livestock or fishing…”
One part of the royal speech
...committee of agricultural cooperatives, settlement cooperatives,
fisheries cooperatives and fresh milk recipients
At the Royal Chitralada Royal Project
8 May 1987
His Majesty, the late King Bhumibol Adulyadej, has always and will forever be in the heart of Thai people. He had dedicated his whole life to improving the lives of Thai people and helping to fight poverty. His dedication is nowhere more evident than in the hundreds and thousands of royal projects established in rural areas throughout the country. Especially, the late king had established many projects regarding the improvement of agriculture in the country, since he recognized how reliant his subjects are on the products of agriculture.
Thailand is an agriculture-based country, the majority of exports come from agriculture products like rice and other crops. Rice, not only is the national staple, but also a major cash crop (only second of rubber). It is also Thailand’s most planted crop, taking up to 45% of all farmland in the country, and nearly 1 in 5 of the population are somehow involved in the production of rice [2].
The late king had the goal in mind of improving the lives of all Thai citizens. Since much of the income of the people comes from selling rice and other similar types of crops, team RIS_BKK, would like to continue and pursue the late king’s vision of Thai citizens having a higher quality of life. The team is doing this by finding a way to increase productivity and output at the same time. Through increasing productivity and output, Thailand can become more competitive in the global market, exporting the same amount of rice as other countries. Evidence of this can be seen in Vietnam. Vietnam is currently more competitive than Thailand in terms of exports because they have used GMO plants, thus helping their economy as shown by the increase in GDP [3]. By increasing the output, the team is helping the farmers earn more, leading them to have a higher standard of living. The rice-growing community in Thailand, the northeast region, more commonly known in Thailand as ‘Isan’, continues to be regarded as some of the poorest regions in the country. The National Statistic Office of Thailand suggests that, in 2012, the income of the population of Isan was 67,888 baht per capita, while the overall Thai population has an income of 183,803 baht per capita [4]. There’s even a large enough income gap between farmers and those engaged in other occupations that the farmers are considered as the poorest group in the country [5]. Through increasing the income of these families, they will have more money to spend meaning that more money is circulating in the system. Therefore, making Thailand not only competitive in the agricultural field but also in other fields. RIS_BKK’s aim is to help increase productivity by increasing the output of local crops like rice, in order to improve the lives of Thai farmers. This can be done by increasing the growth rate of rice so that farmers are able to farm and harvest more frequently throughout the year. Thus, they are able to rely on only rice farming and not other freelance work. We noticed that the farming community in Thailand has been using chemicals to increase the output from plants. Currently, consumers are shifting their demands towards organically grown produce, however, the output and quality of organically grown produce are unmatched when compared to chemically grown produce. Because of this, we wanted to find an alternative method that would rival the output and quality of other chemical-based methods, like fertilizers and pesticides.
Before the start of our summer holiday, we all had the opportunity to attend the workshop hosted by iGEM at Mahidol University, Bangkok. There, we were introduced officially to the iGEM competition. The workshop was extremely inspirational, and there, we decided that we would enter the iGEM competition.
Description
Nitrogen-fixing is a process in which nitrogen gas (N2) from the atmosphere is incorporated into the tissue of certain plants. Plants are able to use nitrogen in the form of ammonium ions or nitrate ions in order to form proteins from carbohydrates. Nitrogen is essential to plants because it is a major component of chlorophyll, essential for photosynthesis, and a major component of amino acids, which some proteins act as structural units in plant cells, etc. They also help regulate plant growth and development as well as plant structure [6][7][8][9].
This process could be performed by certain soil microorganisms,
called diazotrophs (Nitrogen-fixing bacteria and archaea) [7]. 90%
of all nitrogen fixed in the soil in agriculture land is carried
out by nitrogen-fixing bacteria and cyanobacteria [6]. The
mechanism of nitrogen fixation could be represented by the
following equation [7]:
N2 + 8H+ + 8e- + 16ATP -> 2NH3 + H2 + 16ADP + 16Pi
Nitrogen-fixing bacteria use the enzyme complex termed nitrogenase to convert gaseous nitrogen from the atmosphere into ammonia [9].
Azospirillum is the best-characterized genus (α-subclass of proteobacteria) of plant growth-promoting rhizobacteria. Presently, five species have been discovered: Azospirillum lipoferum, Azospirillum brasilense, Azospirillum amazonense, Azospirillum halopraeferens, and Azospirillum irakense. Other types of free-living diazotrophs are detected in association with plant roots include Gluconacetobacter diazotrophicus (A. diazotrophicus), Herbaspirillum seropedicae, Azoarcus spp. and Azotobacter. Provided certain environmental and soil condition, Azospirillum can positively impact plant growth, crop yields, and nitrogen content of the plant. This effect on plants created by Azospirillum has been assigned to several mechanisms including biological nitrogen fixation and auxin production. It converts atmospheric nitrogen into ammonium under microaerobic conditions at low nitrogen levels, through using the enzyme nitrogenase. Azospirillum has been isolated from the rhizosphere of many types of grasses and cereals and has been shown to have beneficial effects on plant growth and crop yields [10].
Azospirillum is beneficial and is best suited for the project. Azospirillum is a non-symbiotic and microaerophilic nitrogen-fixer. It has the special ability to fix nitrogen in an environment of low oxygen and nitrogen. By having a natural association with roots of cereal, millet, and grasses, Azospirillum is effective for the development of low land rice. Thailand is a tropical country and has a higher average temperature compared so many parts of the world, making Azospirillum is the perfect bacteria as it can tolerate soil temperature of 30-40 degrees celsius [10].
In Thailand, there is a very short period of time between harvesting and the cultivation of rice. As a result, many Thai farmers adopt the process of rice residue burning to manage common residue. Countless Thai farmers believe that rice residue burning is beneficial to the yields and adopt the process to control the number of weeds, remove waste residue, and release nutrients for the next crop cycle [11]. However, this has an effect most farmers don’t realise. Burning rice fields, not only burns weeds and waste but also beneficial microbes in the soil. Thus, rice do not have anything to help fix nitrogen.
Our solution to help increase productivity by increasing the output of crops is to make plants fix nitrogen by themselves so that they are less dependent on the environment around them. Nif genes, genes responsible for the mechanism of nitrogen-fixation, are extracted from Azospirillum and transferred into competent cells of E. coli DH5-α as a part of 2 different vectors, pGEM and pCAMBIA. We, then, transformed the inserted gene in pCAMBIA into Azospirillum. In the future, we hope that we can allow Azospirillum to infect plants and transfer its nitrogen-fixing genes into crops.
There are many different ways to consider and look at this problem. Similarly, there are many different solutions to this problem. Synthetic fertilizers perform the same process of increasing the nitrogen content in the soil. But, when synthetic fertilizers are overused, there is an overabundance of nitrogen in the soil. If plants cannot use all of it, the excess nitrate, which is converted from the nitrogen, can run off into streams or groundwater. This could potentially cause sources of water to become undrinkable in communities. Not only that but as nitrate is also used by aquatic plants, this could increase the population of aquatic plants in polluted streams. This has a negative impact on the fish and other aquatic animals because when those plants decompose, oxygen is stripped from the water. As a result, many aquatic animals die from this [7]. As a consequence, synthetic biology is perfect for the solution to our problem. Synthetic biology will not damage and ruin the environment and other living organisms and provide a more permanent solution.
References
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