Table of Content

01 February 2023, Volume 56 Issue 3

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CONTENTS

CONTENTS

Scientia Agricultura Sinica. 2023, 56(3):  0.  doi:10.3864/j.issn.0578-1752.2023560300

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CROP GENETICS & BREEDING·GERMPLASM RESOURCES·MOLECULAR GENETICS

Breeding and Application of a New Thermo-Tolerance Rice Germplasm R203

LIU Gang, XIA KuaiFei, WU Yan, ZHANG MingYong, ZHANG ZaiJun, YANG JinSong, QIU DongFeng

Scientia Agricultura Sinica. 2023, 56(3):  405-415.  doi:10.3864/j.issn.0578-1752.2023.03.001

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【Objective】 The global warming has led to the increasingly serious heat damage on the heading and flowering stage of rice. To reduce the impact of heat damage on rice production and to ensure food security in China and even the world, new rice germplasms with thermo-tolerance on heading stage should be identified and new thermo-tolerance varieties need be bred. 【Method】Guanghui 128 (Qiguizao/Ce64//Minghui 63) was used as the heat resistant parent, through hybridization, multiple crossing and pedigree selection, the lines with high seed setting rate and small variation on heading and flowering stage during the high temperature were screened out for several generations’ breeding process. Then the selected higher generation lines were identified to create new thermos-tolerance rice germplasms in artificial climate chamber (The treated plants will be moved into the chamber on the flowering day, high temperature treatment is 9:00-15:00, 38℃, 15:01-8:59 28℃, the relative humidity is 75%, and the treatment lasts for 7 days), with analysis of agronomic trait. 【Result】The new germplasm R203 has stronger thermo-tolerance and higher seed setting rates under both normal and high temperature conditions (94.5% at normal temperature, 81.9% at high temperature, and 86.7% at relative). Its agronomic traits, quality and comprehensive resistance all meet the production standards. Above all, R203 has the potential to breed new thermos-tolerance hybrid rice varieties. The seed setting rates of 7 hybrid combinations with R203 as the male parent and seven three-line male sterile lines as the female parent were between 83.4%-99.4% under natural high temperature conditions. Among them, Taiyou 203, a new three-line medium indica hybrid rice has good qualities, the seed setting rate was 87.9%, the comprehensive relative heat resistance coefficient was 1.11, and the heat resistance reached level 1. In the production test, the yield increased by 5.36% compared with the control, and the yield increase point accounted for 85.71%. It has good high and stable yield, and the rice quality reached the second level of the ministerial standard. Thus Taiyou 203 has good promotion and application value. 【Conclusion】Currently, basic research on heat resistance is not enough to support the breeding of new practical heat resistant varieties, the rice resources in areas prone to high temperature and humidity are preferred as materials for breeding new heat tolerance lines, a new heat-resistant rice variety R203 was created by phenotypic selection, and a practical heat-resistant rice variety Taiyou 203 was developed by using heterosis.

CROP GENETICS & BREEDING·GERMPLASM RESOURCES·MOLECULAR GENETICS

Cloning of MYBL2 Gene from Brassica and Its PCR Identification in Genomes A, B and C

ZOU Ting, LIU LiLi, XIANG JianHua, ZHOU DingGang, WU JinFeng, LI Mei, LI Bao, ZHANG DaWei, YAN MingLi

Scientia Agricultura Sinica. 2023, 56(3):  416-429.  doi:10.3864/j.issn.0578-1752.2023.03.002

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【Objective】In Arabidopsis, MYBL2 negatively regulates the biosynthesis of anthocyanins and proanthocyanidins. The MYBL2 genes from six Brassica species with different leaf colors were cloned. By analyzing the sequence and expression pattern of MYBL2, the function of MYBL2 in the biosynthesis of anthocyanins in Brassica species was explored. 【Method】The sequences of MYBL2 from the six Brassica species with different leaf colors were obtained using homology-based cloning method and multi sequence alignment and phylogenetic tree analysis were performed. The purple leaf materials of B. rapa, B. napus, B. juncea and B. carinata were treated with shading, and the expression level of MYBL2 gene was analyzed by transcriptome sequencing and qRT-PCR. The qRT-PCR in B. oleracea, B. napus, B. juncea and B. carinata with purple and green leaves were also performed to evaluate the expression level of MYBL2. Based on the nucleotide variation sites of the cloned MYBL2-1 and MYBL2-2 sequences, PCR markers which could distinguish the genomic origin of alleles were developed. 【Result】A total of 56 copies of 9 homologs of MYBL2-1 and MYBL2-2 were cloned from 19 samples of six species of Brassica. The BcaMYBL2-1 gene was obtained for the first time. The total length of BcaMYBL2-1 sequence was 867 bp, including two introns of 168 bp and 102 bp respectively, encoding 198 amino acids, with a molecular weight of 22.69 kD and an isoelectric point (pI) of 8.72. Sequence alignment and evolutionary analysis showed that BcaMYBL2-1 was derived from B genome. Among the MYBL2-1 and MYBL2-2 copies of six species in Brassica, only BraA07.MYBL2-1, BolC06.MYBL2-1 and BcaMYBL2-1 exhibited sequence differences in different leaf color materials. After shading treatment, the leaf color of purple leaf material becomes lighter than that of the unshaded part. In Chinese cabbage Zibao 5, the expression of BraA07.MYBL2-1 and BraA02.MYBL2-2 in the shaded part were 0.7 and 0.4 times of that in the unshaded part, respectively. In Brassica napus with purple leaves and white flowers, the expression of BnaA07.MYBL2-1, BnaC06.MYBL2-1, BnaA02.MYBL2-2 and BnaC02.MYBL2-2 in the shaded part were 0.4, 0.5, 0.4 and 0.4 times of that in the unshared part, respectively. In purple leaf mustard, the expression of BjuA07.MYBL2-1, BjuB03.MYBL2-1, BjuA02.MYBL2-2 and BjuB05.MYBL2-2 in the shaded part were 0.4, 0.3, 0.4 and 0.2 times of that in the unshared part, respectively. In B. carinata with purple leaf, the expression of BcaMYBL2-1, BcaB03.MYBL2-1 and BcaC03.MYBL2-2 in the shaded part were 0.3, 0.4 and 0.5 times of that in the unshared part, respectively. However, the expression of BcaB05.MYBL2-2 in the shaded part was 2.4 times of that in the unshared part. Comparing the expression of MYBL2 gene in different leaf color materials of Brassica, the results showed that the expression of most of the homologous genes of MYBL2 gene in purple leaf materials was higher than that in green leaf materials except kale. In kale, the expression of BolC06.MYBL2-1 and BolC02.MYBL2-2 in green kale were 2.5 and 3.5 times that in purple kale, respectively. In Brassica napus, the expression of BnaA07.MYBL2-1, BnaC06.MYBL2-1 and BnaC02.MYBL2-2 in purple leaf and white flower were 7.5, 8.6 and 26.0 times of that in green leaf and white flower, while the expression of BnaA02.MYBL2-2 in green leaf and white flower was 13.0 times of that in purple leaf and white flower. The expression levels of BjuA07.MYBL2-1, BjuB03.MYBL2-1, BjuA02.MYBL2-2 and BjuB05.MYBL2-2 in Brassica juncea were 8.3 times, 11.8 times, 23.2 times and 14.6 times of those in Sichuan yellow respectively. In B. carinata with purple leaf, BcaMYBL2-1 and BcaB03.MYBL2-1 were 7.1 and 27.6 times as much as W-BCDH76, respectively. However, BcaB05.MYBL2-2 and BcaC03.MYBL2-2 genes of W-BCDH76 were 2.8 and 5.0 times as much as those of B. carinata with purple leaf, respectively. Five pairs of primers were obtained, which can effectively identify the MYBL2 from A, B and C genomes of Brassica. 【Conclusion】After shading treatment, MYBL2 gene expression was closely related to light. The regulation mechanism of MYBL2 gene in Brassica plants involved in anthocyanin biosynthesis was different from that of Arabidopsis plants in which MYBL2 gene negatively regulated anthocyanin biosynthesis.

TILLAGE & CULTIVATION·PHYSIOLOGY & BIOCHEMISTRY·AGRICULTURE INFORMATION TECHNOLOGY

Effect of Post-Anthesis Shading Stress on Eating Quality of Indica Rice in Chengdu Plain

ZHU YouYun, ZENG YuLing, LI Bo, YUAN YuJie, ZHOU Xing, LI QiuPing, HE ChenYan, CHEN Yong, WANG Li, CHENG Hong, ZHOU Wei, TAO YouFeng, LEI XiaoLong, REN WanJun, DENG Fei

Scientia Agricultura Sinica. 2023, 56(3):  430-440.  doi:10.3864/j.issn.0578-1752.2023.03.003

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【Objective】 The aim of this study was to clarify the effect of post-anthesis shading stress on the eating quality of indica rice in Chengdu Plain, so as to provide the theoretical and practical basis for the selecting of shade-tolerant rice varieties in low light rice region. 【Method】 Field light control experiments were conducted in 2019 and 2020 in Wenjiang, Sichuan by using indica rice varieties with different amylose contents as materials. The effect of shading stress on eating quality of rice was studied by measuring the components and RVA spectrum values of rice, as well as the texture properties and eating quality of cooked rice. 【Result】(1)Shading stress significantly decreased amylose, amylopectin, and total starch contents of rice, but significantly increased the contents of protein and fat. (2)Shading stress significantly decreased the peak viscosity and breakdown of rice, while increased the pasting temperature and setback. (3)Shading stress also significantly increased the hardness of cooked rice, but decreased the stickiness and elasticity, which contributed to a significant reduction in the comprehensive score of cooked rice by decreasing both appearance and taste. (4)The results of principal component analysis showed that the rice components, RVA spectrum values, and texture properties could explain 81.2% of the total variation of eating quality of cooked rice. The comprehensive score of rice was significantly and positively correlated with the amylose, starch content, peak viscosity, breakdown, elasticity and stickiness, but negatively related to the protein and fat contents. 【Conclusion】 The protein and fat contents possessed greater influence on the eating quality of cooked rice than that of amylose and starch contents under shading stress. The shading stress leaded to significant variations in starch, protein, and fat components in rice, which contributed to a decrease in peak viscosity and breakdown, but an increase in pasting temperature and setback. And then, those variations contributed to the significantly decrease in eating quality of cooked rice by increasing the hardness, but decreasing both stickiness and elasticity of cooked rice.

Effects of Different Nitrogen Gradients on Yield and Nitrogen Uptake of Hybrid Seed Maize in Northwest China

LIU Dan, AN YuLi, TAO XiaoXiao, WANG XiaoZhong, LÜ DianQiu, GUO YanJun, CHEN XinPing, ZHANG WuShuai

Scientia Agricultura Sinica. 2023, 56(3):  441-452.  doi:10.3864/j.issn.0578-1752.2023.03.004

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【Objective】The law of nitrogen uptake and accumulation of maize in seed production in China is unclear. In view of this, the responses of biomass accumulation, yield formation, and the nitrogen uptake to nitrogen supply were studied in order to provide the theoretical basis for green and efficient hybrid maize seed production. 【Method】 Taking the parent varieties combination of large area seed production as the experimental material, the on-site experiment was carried out from 2019 to 2020. A completely random block design was applied to study the effects of different nitrogen gradients on parental biomass, grain yield and nitrogen uptake and accumulation of maize seed production. Four nitrogen gradients were set up as basal fertilizer control (CK), 168, 240 and 320 kg N·hm^-2, respectively. 【Result】The accumulation of parental biomass of hybrid maize seed production increased with the increase of nitrogen gradients. The yield of maize seed production increased at first and then remained stable with the increase of nitrogen gradients. N240 treatment achieved higher yield, nitrogen use efficiency and grain nitrogen concentration at the same time, which were consistent in the two years. The yield under N168 treatment was higher than that under N240 treatment in the second year, but the nitrogen concentration was lower than that under N240 treatment. The nitrogen concentration of the whole plant of female parent straw and male parent was higher in high nitrogen gradient treatment than that in low nitrogen gradient treatment. The critical nitrogen concentration of maximum biomass of female parent at filling stage was 15.08 g·kg^-1, and there was a linear correlation between female parent biomass and nitrogen concentration at harvest stage. The post-silking biomass of each topdressing nitrogen fertilizer treatment was higher than that of pre-silking stages in two years, and increased with the increase of nitrogen gradients. The change rule of post-silking nitrogen uptake ratio with nitrogen gradients was consistent with the biomass. There was no significant difference in yield level, biomass accumulation, and nitrogen uptake between N320 treatment and N240 treatment. Under the premise of comprehensive consideration of yield and hybrid seed quality, N240 could be regarded as the recommended nitrogen application rate for hybrid maize seed production in this region. 【Conclusion】 The optimal nitrogen application rate could increase yield and nitrogen use efficiency by regulating the ratio of nitrogen uptake for maize parents in pre- and post-silking in seed maize production. This study revealed that optimizing nitrogen application rate to stabilize nitrogen uptake in the pre-silking stage and ensure nitrogen supply after anthesis was the key to achieve high yield and nitrogen use efficiency of seed maize, and provided a theoretical basis for sustainable production of hybrid seed maize.

Effect of Phosphorus Application on Physiological Parameters and Antioxidant Capacity in Alfalfa Leaves

ZHAO JianTao, YANG KaiXin, WANG XuZhe, MA ChunHui, ZHANG QianBing

Scientia Agricultura Sinica. 2023, 56(3):  453-465.  doi:10.3864/j.issn.0578-1752.2023.03.005

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【Objective】The aim of this study was to investigate the effects on nutrient uptake, photosynthetic pigment content, stomatal aperture and antioxidant system of alfalfa leaves after three consecutive years of phosphorus fertilizer application, so as to provide scientific fertilization methods for efficient production of alfalfa.【Method】Field trials were conducted at the forage experiment station of Shihezi University in Xinjiang from 2019 to 2021. ‘WL366HQ’ alfalfa was used as the test material, and four fertilizer treatments in the experiment included 0 (CK), 50 (Low P, LP), 100 (Middle P, MP), and 150 (High P, HP) kg·hm^-2. Samples were taken at the first flowering of alfalfa to determine the hay yield, nitrogen and phosphorus content, pigment content, stomatal opening, antioxidant enzyme activity, and oxidizing substances of leaves.【Result】The nitrogen and phosphorus content, photosynthetic pigment content and stomatal aperture of leaves of alfalfa increased significantly (P<0.05) after 3 years of continuous phosphorus fertilization, with the highest nitrogen and phosphorus content under MP treatment (54.74 and 2.99 g·kg^-1, respectively), the highest chlorophyll a and chlorophyll b content under MP, and the highest carotenoid content under LP or MP, and the lowest in CK. The stomatal aperture was the highest under MP and significantly higher than that under CK (P<0.05). Therefore, phosphorus fertilizer had different effects on the morphological and physiological characteristics of alfalfa leaves, and MP treatment significantly affected the photosynthetic and physiological characteristics of alfalfa leaves (P<0.05). Superoxide dismutase (SOD), peroxidase (POD) and catalase (CAT) activities were the highest under MP, with the value of 162.55, 406.40 and 147.13 U·g^-1, respectively, and were significantly higher than those under CK (P<0.05). The malondialdehyde (MDA), hydrogen peroxide (H₂O₂) and proline (Pro) content were the lowest under MP, at 2.38 and 1.04 μmol·g^-1 and 56.85 μg·g^-1, respectively. According to the Pearson correlation analysis, the total N and P contents of alfalfa leaves showed significant positive correlation (P<0.05) with chlorophyll content, carotenoid content, stomatal aperture, SOD activity and POD activity, and significant negative correlation (P<0.05) with MDA and H₂O₂ content. The overall evaluation showed that the principal component scores of phosphorus application treatments were ranked as MP>HP>LP>CK. 【Conclusion】 The reasonable application of phosphorus fertilizer increased the nutritional characteristics and photosynthetic physiological characteristics of alfalfa leaves, thus avoiding the stressful effects brought by other environmental factors, enhancing the adaptive capacity of alfalfa, and making it better adapted to external environmental changes. Considering the leaf morphology, photosynthetic physiological characteristics and antioxidant enzymes and oxidizing substances of alfalfa, the phosphorus application rate of 100 kg·hm^-2 was suitable under this experiment.

PLANT PROTECTION

Distribution and Variation Analysis of AVR-Pita Family in Magnaporthe oryzae from Heilongjiang Province and Hainan Province

LIU RUI, ZHAO YuHan, GU XinYi, WANG YanXia, JIN XueHui, WU WeiHuai, ZHANG YaLing

Scientia Agricultura Sinica. 2023, 56(3):  466-480.  doi:10.3864/j.issn.0578-1752.2023.03.006

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【Objective】By detecting the distribution and variation characteristics of the AVR-Pita family in different Magnaporthe oryzae strains from Heilongjiang Province and Hainan Province, the pathogenic phenotypes of the variation types were understood, which provided a reference for the screening and breeding of disease-resistant germplasm in the region.【Method】Six pairs of specific primers were designed for the promoter region and CDS region by referring to the AVR-Pita family sequence published in NCBI. DNA was extracted from 397 single spore strains of M. oryzae collected from different regions of Heilongjiang Province and Hainan Province in 2020, and PCR amplification was performed. By electrophoresis test results, representative strains from different regions were selected to sequence the amplified fragments. The sequencing results were compared with the corresponding base and amino acid sequences in NCBI, and the avirulence function of different variation types of M. oryzae strains was verified by using rice resistant single gene lines.【Result】In the PCR electrophoresis results, the M. oryzae strains in Heilongjiang Province carried all the genes to be tested, which were widely distributed and had a high detection frequency. However, only AVR-Pita1 and AVR-Pita2 were carried by M. oryzae strains in Hainan Province, and concentrated in low frequency. The results of avirulence gene composition analysis showed that the strains of M. oryzae in Heilongjiang were complex and diverse, and the genotypes were more abundant than those in Hainan. The sequencing results of PCR products showed that the AVR-Pita family was divided into 19 types with point mutation, insertion and deletion as the main variation types, and the variation types of strains from different M. oryzae populations were specific. Ten variation types were detected in AVR-Pita1, among which AVR-Pita1- (1-5) was a unique variation type of M. oryzae strains in Heilongjiang Province, and AVR-Pita1- (6-10) was a unique variation type of M. oryzae strains in Hainan Province. After functional verification of these ten variation types, it was found that the avirulence functions were lost. Eight variation types were detected in AVR-Pita2, among which AVR-Pita2- (1-4) was a unique variation type of M. oryzae strains in Heilongjiang Province, and AVR-Pita2- (5-8) was a unique variation type of M. oryzae strains in Hainan Province. After functional verification, it was found that the avirulence functions were lost. Only one variation type (AVR-Pita3-1) was detected in AVR-Pita3 in Heilongjiang Province.【Conclusion】The AVR-Pita family in Heilongjiang and Hainan populations were mutated alleles. After pathogenicity identification, all mutation types could not be identified by the corresponding resistance genes Pi-ta and Pi-ta². Therefore, the resistance genes Pi-ta and Pi-ta² can be used to polymerize other resistance genes to ensure the disease resistance of varieties in the process of disease resistance breeding and utilization of rice blast in Heilongjiang Province and Hainan Province. At the same time, the distribution and variation types of AVR-Pita family in M. oryzae strains from different geographical sources are specific.

SOIL & FERTILIZER·WATER-SAVING IRRIGATION·AGROECOLOGY & ENVIRONMENT

Response of Biomass and Nutrient Competition Between Oilseed Rape and Weed to the Rate of N, P and K Fertilizer

SHENG QianNan, YU XiaoHong, ZHOU Xiong, TIAN GuiSheng, WU HaiYa, GENG GuoTao, YAN JinYao, LI Jing, REN Tao, LU JianWei

Scientia Agricultura Sinica. 2023, 56(3):  481-489.  doi:10.3864/j.issn.0578-1752.2023.03.007

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【Objective】 Weed is one of the important factors limiting the high yield of oilseed rape. The reasonable management of nutrients plays a key role in weed control. This study explored the effects of N, P and K fertilizer application rate on the biomass and nutrient competition between oilseed rape and weed under field conditions, so as to provide a basis for ecological grass control and sustainable agricultural development.【Method】 The field experiment was carried out in Wuxue City, Hubei Province, and a single factor experimental design was adopted. Four different rate gradients of N (0, 90, 180 and 270 kg N·hm^-2, expressed by N0, N1, N2 and N3 respectively), P (0, 45, 90 and 135 kg P₂O₅·hm^-2, expressed by P0, P1, P2 and P3 respectively), and K (0, 60, 120 and 180 kg K₂O·hm^-2, expressed by K0, K1, K2 and K3 respectively) were set. Oilseed rape yield, the biomass of oilseed rape and weed were investigated at the mature stage. The corresponding nutrient content was measured, the nutrient accumulation was calculated, and the nutrient competition relationship between oilseed rape and weed and its response to the rate of fertilizer were analyzed. 【Result】 Fertilization significantly increased oilseed rape yield, total shoot biomass and corresponding nutrient accumulation, among the three nutrients of N, P and K, the growth and nutrient accumulation of oilseed rape were the most sensitive to phosphorus deficiency. The yield of oilseed rape treated with N0, P0 and K0 were 560, 227 and 1 490 kg·hm^-2 respectively, accounting for 18.2%, 7.5% and 50.1% of the highest yield of corresponding nutrient treatments (N3, P3 and K3), respectively. The variation trend of total shoot biomass with nutrient input was consistent with yield. The oilseed rape corresponding nutrient accumulation of N0, P0 and K0 treatments were 19.96, 0.88 and 26.21 kg·hm^-2, respectively, which accounted for 12.24%, 3.72% and 22.26% of the highest nutrient accumulation of corresponding nutrient treatments, respectively. With the increasing of the application rate of three kinds of nutrient, the weed biomass and corresponding nutrient accumulation decreased (except the highest treatment P2 in the P fertilizer rate test), the weed biomass of N0, P0 and K0 treatments were 1 365, 3 060 and 1 535 kg·hm^-2, respectively, which were the corresponding minimum weed biomass (N3, P2 and K3 treatment) 7.59 times, 5.19 times and 3.61 times, respectively; the corresponding nutrient accumulation of weed in N0, P0 and K0 treatments were 17.60, 1.91 and 9.38 kg·hm^-2, respectively, which were 3.78 times, 1.54 times and 1.52 times higher than that in the corresponding minimum weed nutrient accumulation (N3, P2 and K3 treatment), respectively. Compared with P and K, the weed and oilseed rape had the greatest nutrient competitiveness to N, and the all N fertilizer treatment, the N content of weed was higher than that of oilseed rape. Fertilization increased the ratio of biomass and corresponding nutrient accumulation between oilseed rape and weed, except for no P application, the ratio of biomass and corresponding nutrient accumulation between oilseed rape and weed in other treatments was greater than 1, and this ratio increased with the increase of fertilization (except the highest treatment P2 in the P fertilizer rate test), which showed adequate nutrient supply can significantly improve the growth and nutrient absorption capacity, meanwhile it played a role in weed suppression.【Conclusion】 The harm degree of weed in winter oilseed rape field was controlled by nutrient supply, fertilization effectively made up for the deficiency that herbicide in the early stage could not completely control weed in the whole growth period. The sufficient application of N, P and K fertilizer could significantly reduce weed biomass and corresponding nutrient accumulation, under the experimental conditions, the response degree of three nutrients to the competitiveness between oilseed rape and weed was P>N>K.

Environmental Safety Risks in Agricultural Application of Effluents from Sugar Molasses-Based Fermentation Industries

WANG XiaoBin, YAN Xiang, LI XiuYing, TU Cheng, SUN ZhaoKai

Scientia Agricultura Sinica. 2023, 56(3):  490-507.  doi:10.3864/j.issn.0578-1752.2023.03.008

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Sugar molasses is a by-product from sugar industries. The sugar molasses-based fermentation industries mainly refer to the fermentation industries using molasses from sugar mills as raw materials for alcohol or yeast fermentation. A large volume of effluents can be produced in the process of sugar molasses-based alcohol or yeast fermentation. Considering the possibility of resource utilization with such effluents, many sugar-producing countries (such as Brazil, India, and China) use the effluents for crop irrigation and fertilization or soil remediation directly into the farmlands by waste disposal methods. Because the effluents from sugar molasses-based fermentation industries are both high concentration organic wastewater, and heavy metal-polluted wastewater, which are difficult to be treated. With the long-term disposal of such effluents into the farmlands in some sugar-producing countries, the problems about ecological environment pollution in soil-crop-water systems are increasingly exposed. At present, some fertilizer production enterprises in China use such effluents as raw materials to produce organic water-soluble fertilizers (accounting for 32%), but the long-term research and monitoring data about environmental safety risks for agricultural application of the effluents from sugar molasses-based fermentation industries are still lack. This paper collected the relevant scientific research literatures since 1980 on the pollution characteristics of the effluents from sugar molasses-based fermentation industries, and their environmental impacts on agricultural application. Through the investigation and review on the relevant research data, this paper evaluated the environmental safety risks for agricultural application of the effluents from sugar molasses-based fermentation industries: (1) Such effluents were at a risk of seriously exceeding the limits for water quality standards, and a risk of ecotoxicity to plants. For example, such effluents had strong acidity, and high salinity, and contained not only high load organic pollutants, but also several heavy metals including 5 heavy metals (As, Hg, Cd, Pb and Cr), as well as other pollutants (such as Mn, Cu, Zn, Ni and Se, etc.). The concentrations of these pollutants mostly exceeded the limits of the Standards for Irrigation Water Quality (GB 5084—2021). (2) Such effluents for agricultural application were at a risk of farmland pollution. The concentrations of pollutants (such as Cu, Cd, Cr, Zn, Ni, Mn, Pb and Cl) detected from the soil samples irrigated with such effluents were about 10-641 times higher than those in the control soil. (3) Such effluents for agricultural application were at a safety risk of agricultural products. The concentrations of pollutants (such as Cu, Cd, Cr, Zn, Ni, Mn and Pb) detected in the grains of crops (wheat and mustard) irrigated with such effluents were about 3-12 times higher than those in the control crops, in which all the pollutants detected in the crops irrigated with such effluents exceeded both the allowable limits specified by FAO/WHO, but also the Maximum Levels of Contaminants in Food (GB 2762—2017) specified by China. In view of the issue of environmental safety risks for agricultural application of such effluents, therefore, it is necessary to strengthen the quality detection and risk control on the organic water-soluble fertilizer products with such effluents as raw material, to enable the safety of effluent utilization in agriculture.

HORTICULTURE

Cloning and Analysis of Key Genes for Vitamin C Synthesis in Cucumber Fruit

WANG ZhuangZhuang, DONG ShaoYun, ZHOU Qi, MIAO Han, LIU XiaoPing, XU KuiPeng, GU XingFang, ZHANG ShengPing

Scientia Agricultura Sinica. 2023, 56(3):  508-518.  doi:10.3864/j.issn.0578-1752.2023.03.009

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【Objective】The aim of this study was to identify the location, quantity and expression characteristics of genes involved in regulating the synthesis of vitamin C (Vc) by L-galactose pathway in cucumber fruits, and to clone the key genes, so as to lay a foundation for the regulation of Vc synthesis in cucumber.【Method】According to the reported Vc-related genes within the L-galactose pathway in Arabidopsis, the encoded amino acid sequence was used for BLAST in Cucumber 9930_V2 reference genome database. TBtools software was used to map the gene position on cucumber chromosomes. The expression of these genes in two cucumber accessions with significant differences in fruit Vc content was analyzed by qRT-PCR. The homologous genes encoding rate limiting enzymes GDP-L-galactose phosphorylase (GGP) and GDP mannose-3'5'- epimerase (GME) were cloned by PCR amplification, and the sequence differences of these genes in cucumber with high Vc and low Vc were analyzed by sequencing. Phylogenetic tree was constructed to analyze the relatedness cucumber GME, GGP and homologs in other species.【Result】Twenty one homologous genes involved in the synthesis of Vc related enzymes, including PMI, PMM, GMPase, GME, GGP, GPP, GalLDH, and GalLDH in L-galactose pathway, were compared in cucumber and were obtained by BLAST, which were distributed on seven chromosomes, with the most numbers on chromosome 5 and chromosome 1. By analyzing the expression of these genes in R48 (with low Vc) and CG45 (with high Vc), it was found that the genes regulating PMI, PMM, GMPase, GME and GalLDH were significantly different between the two materials. The sequence analysis of Vc synthesis rate-limiting enzyme GGP and GME related genes showed that the full length of CsGME2 gene was 3 537 bp in R48 and 3 541 bp in CG45. There were multiple SNP sites and Indel difference between the two materials, among which one mutation site was located in the CDS region, and resulted in the amino acids changes. Through the analysis of the protein properties of rate limiting enzymes GME and GGP regulating vitamin C synthesis, it was found that the protein properties of GME and GGP in different species were not significantly different, which were hydrophilic proteins and their functions were relatively conservative. Evolutionary tree analysis found that the clusters with close genetic relationship among different species were highly conservative during evolution.【Conclusion】Twenty one L-galactose pathway related genes of cucumber Vc synthesis were identified, which were distributed on seven chromosomes. It was speculated that the key enzymes including PMI, PMM, GMPase, GME, GalLDH and GGP might affect the Vc content in cucumber fruits. The functions of key enzymes GME and GGP regulating the rate limiting step of Vc synthesis were relatively conservative. The SNP site on CsGME2 gene in the two materials of high Vc and low Vc resulted in changes in amino acid sequence.

Effects of Urea Slow-Release Functional Fertilizer Containing Melatonin on Growth, Yield and Phosphorus Use Efficiency of Tomato Under Reduced Phosphorus Application Conditions

LIU MingHui, TIAN HongYu, LIU ZhiGuang, GONG Biao

Scientia Agricultura Sinica. 2023, 56(3):  519-528.  doi:10.3864/j.issn.0578-1752.2023.03.010

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【Objective】Melatonin has multiple beneficial effects on plants. However, its active chemical properties limit its application in agricultural production. In this study, urea slow-release functional fertilizer containing melatonin (hereinafter referred to as functional fertilizer) was prepared by coating technology, and its effects on tomato growth, yield, and quality as well as phosphorus use efficiency were studied, so as to provide the theoretical basis for melatonin application with high use efficiency and reducing fertilizer rate on tomato.【Method】 The release rate of melatonin in functional fertilizer was studied by water bubble method. Then the effects of melatonin-functional fertilizer on the growth of tomato seedlings were studied by plug seedling. Finally, a pot experiment was conducted to study the effects of functional fertilizer on plant growth and dry matter distribution, root growth, phosphorus absorption and distribution, phosphorus utilization rate and fertilizer yield contribution rate, root phosphatase activity, and fruit yield and quality. Four treatments were set, namely, phosphorus application (+P), non-phosphorus application (-P), urea slow-release fertilizer and functional fertilizer (-P+M and +P+M).【Result】The real melatonin content after coating was 35% of the total melatonin concentration when coating. When water bubbled to 60 d, the amount of melatonin residue in functional fertilizer was 6.61%. The application of functional fertilizer in conventional tomato seedlings could significantly promote seedling growth, and the seedling index was increased by 70.2% compared with the control. The pot experiment showed that the biomass of tomato roots, stems, leaves and fruits under -P treatment decreased by 19.64%, 18.51%, 28.99% and 28.73% compared with that under +P treatment, respectively. The dry matter distribution ratios of roots and stems increased by 10.03% and 11.63%, respectively, while the dry matter distribution ratios of leaves and fruits decreased by 2.74% and 2.39%, respectively. The application of functional fertilizer could significantly increase the biomass accumulation of all tissues under the two phosphorus treatments. The dry matter distribution ratio of root under the condition of increasing +P was 12.14%, the dry matter distribution ratio of stem and leaf under the condition of reducing -P was 6.00% and 5.90%, and the dry matter distribution ratio of fruit increased by 9.06%. -P treatment reduced the total length, total surface area and total volume of roots, and increased the number of root tips. The application of functional fertilizer could increase the total length, total surface area, total volume and number of root tips under two phosphorus levels. -P treatment significantly reduced the phosphorus content in roots, stems, leaves and fruits of tomato plants, increased the phosphorus distribution in roots, stems and leaves, and reduced the phosphorus distribution ratio in fruits. The application of functional fertilizer could significantly increase the phosphorus content of all tissues under the two P treatments, increase the phosphorus distribution ratio of roots under +P treatments, decrease the phosphorus distribution ratio of stems and leaves under -P treatment, and increase the phosphorus distribution ratio of fruits. The application of functional fertilizer could significantly improve phosphatase activity, whole plant phosphorus uptake, fertilizer utilization rate and fertilizer yield contribution rate. -P treatment reduced tomato yield, but application of melatonin functional fertilizer significantly increased yield under -P treatment reduced the tomato yield by 17.57%, but the application of functional fertilizer under -P treatment increased the yield by 21.32%, and had no significant effect on the yield under +P treatment. In addition, the application of functional fertilizer could comprehensively improve the quality of tomato fruits under -P or partial +P conditions.【Conclusion】The application of melatonin coated in slow-release fertilizer could significantly reduce the amount of melatonin, improve the growth quality of tomato seedlings and plants in the whole growth period, increase the utilization efficiency of phosphorus fertilizer, and finally improve the yield and fruit quality.

FOOD SCIENCE AND ENGINEERING

Research Progress on the Application of Non-Saccharomyces During Wine Fermentation

WANG ChunXiao, YU JunZhu, ZHOU WenYa, XU YinHu

Scientia Agricultura Sinica. 2023, 56(3):  529-548.  doi:10.3864/j.issn.0578-1752.2023.03.011

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Non-Saccharomyces yeast are naturally present on grape skin, which mainly play their roles in the maceration and initial stage of fermentation during winemaking process. More and more concerns have been recently focused on the application of non-Saccharomyces yeast in wine fermentation. Non-Saccharomyces yeast often have weaker fermentation ability than Saccharomyces cerevisiae and can convert the reducing sugar into ethanol and other fermentation by-products. Therefore, non-Saccharomyces are the potential excellent yeast for producing wine with complex flavor traits and lower ethanol degree. Different non-Saccharomyces yeast species have different metabolic characteristics in wine fermentation applications. The selection of non-Saccharomyces yeasts with certain characteristics for application in fermentation can improve the specialized quality of wine. This review was based on the initial summary on the species, winemaking traits, and application ways of commercial non-Saccharomyces yeast, and emphasized on the positive role of non-Saccharomyces in wine color, aroma, taste, safety and health, their metabolic mechanisms and corresponding research hotspot. The positive role of non-Saccharomyces in wine color included the promotion of high acid production, high polysaccharide production, high extracellular pyruvate production and low sorption properties. Different non-Saccharomyces yeast could promote the fruity aroma enhancement in wine by low production of ethanol, acetaldehyde, and volatile phenols, and high production of ethyl acetate, acetic acid ester, ethyl ester, higher alcohols, and enzymes related with the release of terpene and mercaptan, and therefore improve the aroma complexity of wine. Non-Saccharomyces yeast could regulate wine mouthfeel traits by high production of glycerol, polysaccharide, and lactic acid, and degradation of malic acid. In addition, non-Saccharomyces yeast could improve the safety quality of wine by decreasing the SO₂ dosage as biological control agent, and reducing the content of toxic compounds through metabolic degradation during alcoholic fermentation. This review further analyzed the current research status on genome and microsatellite loci analysis. The main inoculation strategies were discussed on application of non-Saccharomyces yeast during wine fermentation, and six aspects were posed for further research concern. This review provided theoretical references for the research on application of the non-Saccharomyces yeasts in wine alcoholic fermentation.

ANIMAL SCIENCE·VETERINARY SCIENCE

Inhibitory Effect of N-acetylcysteine on Bisphenol A-Induced Apoptosis and Inflammatory Response in Porcine Kidney Cells

TAO WenJing, ZHANG ZiTing, LIU Yuan, SONG Dan, LI XiangChen

Scientia Agricultura Sinica. 2023, 56(3):  549-558.  doi:10.3864/j.issn.0578-1752.2023.03.012

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【Background】 Bisphenol A (BPA) is widely used in the industrial manufacturing of plastic materials, which seeps out from plastic products and exposes to various environmental media such as food, water, soil, and air, resulting in long-term exposure of animals. It is passed to offspring through the placenta and breast milk, interfering with animal growth and development and adversely affecting animal growth performance and production efficiency. N-acetylcysteine (NAC), as a recognized potent antioxidant, can regulate various pathophysiological processes, such as oxidative stress, apoptosis, and inflammation. However, the regulatory role of NAC on BPA-induced porcine kidney cell injury remains unclear. 【Objective】This study aimed to explore the potential role of the antioxidant NAC on BPA-induced apoptosis and inflammatory responses in PK15 cells.【Method】 PK15 cells were selected as experimental materials, and the activities of catalase (CAT), total superoxide dismutase (T-SOD) and glutathione peroxidase (GSH-Px) were measured by the corresponding antioxidant detection kits. PK15 cells were treated with different concentrations of NAC (0, 2, 5, 10 mmol·L^-1) and then co-treated with BPA, and then the cell viability was detected by CCK-8 to select the optimal concentration of NAC. The expression of apoptosis-related genes (BAX, BCL-2 and Caspase3) and inflammatory genes (IL-8, IL-6, IL-1β and TNF-α) as well as protein expression were detected by real-time quantitative PCR (qRT-PCR) and western blotting. The number of apoptotic cells and nuclear factor kappa B (NF-κB) nuclear translocation were detected by immunofluorescence staining. 【Result】 The results showed that BPA significantly reduced the activities of CAT, T-SOD and GSH-Px in PK15 cells, compared with the control group (P<0.05). CCK-8 results showed that BPA significantly decreased PK15 cell viability in contrast to control group, whereas the different concentrations of NAC significantly promoted cell viability, and 5, 10 mmol·L^-1 NAC pretreatment significantly promoted cell viability when compared with BPA alone. qRT-PCR and western blotting showed that BPA treatment significantly increased BAX and Caspase3 mRNA and protein expression, and decreased the BCL-2 mRNA and protein expression, whereas NAC pretreatment could reduce the increased BAX and Caspase3 expression and increase the decreased BCL-2 expression induced by BPA. Hoechst33258 fluorescence staining indicated that the cells treated with BPA showed strong blue fluorescence staining and obvious nuclear shrinkage, whereas NAC pretreated cell showed weak blue fluorescence. BPA treatment significantly increased the relative mRNA expression of inflammation-related factors (IL-8 and IL-6) (P<0.05), whereas NAC pretreatment inhibited BPA-induced increased inflammation-related factors (IL-8, IL-6 and IL-1β mRNA) relative expression. Immunofluorescence analysis of NF-κB nuclear translocation showed that NF-κB was mainly distributed in cytoplasm in the control group, whereas NF-κB was mainly distributed in the nucleus after BPA treatment and NAC pretreatment reduced nuclear NF-κB expression. 【Conclusion】 NAC significantly increased PK15 cell viability and inhibited PK15 cell apoptosis and inflammatory response induced by BPA.

Analysis of Growth Performance as well as Carcass and Meat Quality Traits in Pingliang Red Cattle

WANG XiuJuan, GAO Han, LI HaiPeng, GAO Xue, SUN BaoZhong, CHENG Qiang, XU Lei, ZHANG YaPeng, LEI YuanHua, WEI Meng, LI SanLu, HU JunWei, ZHANG ChangQing, GAO HuiJiang, LI JunYa, ZHANG LuPei, CHEN Yan

Scientia Agricultura Sinica. 2023, 56(3):  559-571.  doi:10.3864/j.issn.0578-1752.2023.03.013

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【Objective】By studying the growth performance, carcass traits and meat quality traits of Pingliang Red Cattle, the meat characteristics and genetic characteristics of Pingliang Red Cattle were deeply explored, so as to provide a reference for breeding of this cattle.【Method】24 Pingliang Red Cattle steers were slaughtered after intensive felling for 180 days, with the ages from 24 to 36 months. Body size, slaughter performance and carcass index were measured, and the meat quality were analyzed on the Striploin and Outside Flat. 【Result】After fattening, the body weight, body height, hip height, heart girth and cannon bone size of adult Pingliang Red Cattle increased by 6.96%, 4.61%, 4.43%, 4.28% and 7.90%, respectively, compared with 22-28 month old Pingliang Red Cattle tested in 2019, while the body length decreased by 3.52%. In addition, the carcass weight was (417.01±56.94) kg, the lean meat weight was (372.48±54.60) kg, the dressing percentage was 61.56%±1.86% and the lean meat percentage was 54.92%±2.60%, compared with the data of Pingliang Red Cattle at the age of 24 months measured in 2010, they increased by 12.61%, 19.16%, -1.91% and 3.82%, respectively; the ratio of meat to bone was 8.65±2.00, compared with the data of 22-27 months in 2012, with the increase of 27.96%. In addition, the weight of high-grade meat pieces (tenderloin, striploin, ribeye and high rib) was (51.59±7.92) kg, which of high-quality meat pieces (rump, knuckle, topside, outside flat, eyeround (ribeye), chuck tender and shoulder chops) was (62.62±5.56) kg, accounting for 13.91%±1.45% and 17.07±2.24% of the net meat weight, respectively. Pingliang Red Cattle had good growth and meat production performance. In terms of meat quality, the shear force of Striploin and Outside Flat was 2.72 kg and 4.43 kg, respectively, it was 19.58% and 64.82% lower than that of A3 grade snowflake beef, but the difference was not significant (P>0.05); the crude protein content of Striploin and Outside Flat both reached more than 20%, and which of Pingliang Red Cattle was significantly higher than that of snowflake beef (P<0.05). The EAA/NEAA ratios of Striploin and Outside Flat were 71% and 68%, respectively, which were higher than the FAO/WHO standard value (60%). EAA/TAA ratios of Striploin and Outside Flat were 42% and 41%, respectively, higher than the FAO/WHO standard value (40%). Compared with A3 grade snowflake beef purchased in the market, Pingliang Red Cattle had rich contents of essential amino acids and delicious amino acids. The results showed that the lysine content of Pingliang Red Cattle Striploin and Outside Flat was higher than that of FAO/WHO amino acid scoring standard model; in addition, the selenium element in Outside Flat of Pingliang Red Cattle was extremely significantly higher (P<0.01), the contents of Fe, Zn, PUFA, n-6/n-3 and PUFA/SFA ratio were significantly lower than those of the control group (P<0.05); its EPA content in Striploin was significantly higher(P<0.05).【Conclusion】After cross improvement and breeding, the growth and meat production performance of Pingliang Red Cattle has been significantly improved, with a larger body, wider chest, and compact body size. More prominently, the meat quality of Pingliang Red Cattle had the characteristics of tender and delicious, rich meat taste and high nutritional value. Pingliang Red Cattle had great development and utilization prospect to produce high-quality beef cattle population.

Genome-Wide Association Study of Egg Quality Traits in Longyan Shan-Ma Duck

SUN YanFa, WU Qiong, LIN RuLong, CHEN HongPing, GAN QiuYun, SHEN Yue, WANG YaRu, XUE PengFei, CHEN FeiFan, LIU JianTao, ZHOU ChenXin, LAN ShiShi, PAN HaoZhe, DENG Fan, YUE Wen, JIANG XiaoBing, LI Yan

Scientia Agricultura Sinica. 2023, 56(3):  572-586.  doi:10.3864/j.issn.0578-1752.2023.03.014

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【Objective】Single nucleotide polymorphisms (SNPs) and candidate genes for egg quality traits in duck were identified through genome-wide association study (GWAS) technology, so as to provide reference for molecular breeding to improve egg quality traits of Longyan Shan-Ma Duck.【Method】Egg quality traits of 235 female Longyan Shan-Ma Duck were measured, including egg weight (EW), egg shaped index (ESI), eggshell thickness (EST), eggshell strength (ESS), eggshell colour L* (ESCL), a* (ESCA), b* (ESCB), albumin height (AH), Haugh unit (HU), egg yolk colour (EYC), egg yolk weight (EYW), and egg yolk percentage relative to egg weight (EYP). Genetic parameters of these traits were estimated using multi-trait animal model by ASReml-R 4.1 software. Blood genomic DNAs of these ducks were genotyped using genotyping-by-sequencing (GBS) technology. The GWAS between egg quality traits of the late laying period and SNPs were performed. 【Result】 The heritability of EW, ESI, EST, ESL, ESA and AU was higher among the egg quality traits of Longyan Shan-Ma Duck, and ranged from 0.21 to 0.70. There was a strong positive genetic correlation (r_g = 0.91±0.37) between EW and AU, a strong negative genetic correlation (r_g = -0.98±1.03) between ESI and EYC. EST had a positive phenotypic correlation (r_p = 0.41±0.06) with ESS, negative genetic and phenotypic correlations with ESA (r_g = -0.86±0.25 and r_p = -0.15±0.07), and positive genetic and phenotypic correlations with ESB (r_g= 0.96±0.37 and 0.18±0.07). There were negative genetic and phenotypic correlations between ESA and ESB (r_g = -0.64±0.28 and r_p = -0.31±0.06). Results from the GWAS showed that seven SNPs were significantly associated with ESI, EST and yolk color (YC) at 5% Bonferroni-corrected genome-wide significance level (P<4.74×10^-6), involving six candidate genes. One SNP, chr20:11135563: G:C, was associated with ESI, which was in leucine rich repeat containing 75A gene, located on chromosome 20. Two SNPs, chr13:5766560:A:G and chrZ:968819:C:T, were associated with EST, which were located on chromosome 13, downstream 6.86 Kb of LOC106014427 and in transcription factor 4 gene, respectively. Four SNPs were associated with EYC, one SNP chr2: 38155965:G:A in potassium voltage-gated channel subfamily H member 8 gene located on chromosome 2; three SNPs located on chromosome 9, two SNPs, chr9:22623156:G:A and chr9:22623155:T:C, in insulin receptor substrate 1 gene, and one SNP, chr9:22490158:A:T, in LOC106018641 gene. Eighty-one SNPs associated with egg quality traits reached at suggestive genome-wide significance level (P<9.48×10^-5) were also found. Thirteen SNPs associated with YC were distributed in the 0.84 Mb (22.16-23.00 Mb) region of chromosome 9.【Conclusion】In this study, genetic parameters of egg quality traits of Longyan Shan-Ma Duck were estimated. Seven significant SNPs, six candidate genes, and one candidate region affecting ESI, EST and EYC traits were identified through GWAS. The findings from the present study provided a reference for the molecular breeding of egg quality traits in Longyan Shan-Ma Duck.