Increase In Lysine Content Research Articles

Genetic analysis of QTLs for lysine content in four maize DH populations

BackgroundLow levels of the essential amino acid lysine in maize endosperm is considered to be a major problem regarding the nutritional quality of food and feed. Increasing the lysine content of maize is important to improve the quality of food and feed nutrition. Although the genetic basis of quality protein maize (QPM) has been studied, the further exploration of the quantitative trait loci (QTL) underlying lysine content variation still needs more attention.ResultsEight maize inbred lines with increased lysine content were used to construct four double haploid (DH) populations for identification of QTLs related to lysine content. The lysine content in the four DH populations exhibited continuous and normal distribution. A total of 12 QTLs were identified in a range of 4.42–12.66% in term of individual phenotypic variation explained (PVE) which suggested the quantitative control of lysine content in maize. Five main genes involved in maize lysine biosynthesis pathways in the QTL regions were identified in this study.ConclusionsThe information presented will allow the exploration of candidate genes regulating lysine biosynthesis pathways and be useful for marker-assisted selection and gene pyramiding in high-lysine maize breeding programs.

Characterization of Plant-Based Meat Treated with Hot Air and Microwave Heating.

Plant-based meat is growing globally due to health, environmental, and animal welfare concerns, though there is a need for quality improvements. This study assessed how different ratios of wheat gluten (WG) to soy protein isolate (SPI) and various baking methods-hot air (HA), microwave (MW), and a combination of both (HA-MW)-affect the physicochemical properties of plant-based meat. Increasing the SPI from 0% to 40% significantly enhanced lightness, hardness, chewiness, water-holding capacity, moisture content, and lysine (an essential amino acid) (p ≤ 0.05). Hardness and chewiness ranged from 4.23 ± 1.19 N to 25.90 ± 2.90 N and 3.44 ± 0.94 N to 18.71 ± 1.85 N, respectively. Baking methods did not affect amino acid profiles. Compared to HA baking, MW and HA-MW baking increased lysine content (561.58-1132.50 mg/100 g and 544.85-1088.50 mg/100 g, respectively) while reducing fat and carbohydrates. These findings suggest that a 40% SPI and 60% WG ratio with microwave baking (360 W for 1 min) optimizes plant-based meat, offering benefits to both consumers and the food industry in terms of health and sustainability.

Effects of electrolyte balance on intestinal barrier, amino acid metabolism, and mTORC1 signaling pathway in piglets fed low-protein diets

A proper dietary electrolyte balance (dEB) is essential to ensure optimal growth performance of piglets. In the low-protein diet, this balance may be affected by the reduction of soybean meal and the inclusion of high levels of synthetic amino acids. The objective of this experiment was to evaluate the optimal dEB of low-protein diets and its impact on the growth performance of piglets. A total of 108 piglets (initial age of 35 d) were randomly divided into 3 groups with 6 replicates of 6 pigs each as follows: low electrolyte diet (LE group; dEB = 150 milliequivalents [mEq]/kg); medium electrolyte diet (ME group; dEB = 250 mEq/kg); high electrolyte diet (HE group; dEB = 350 mEq/kg). Results indicated that the LE and HE diet significantly decreased the average daily gain, average daily feed intake, and crude protein digestibility (P < 0.05) in piglets. Meanwhile, LE diets disrupted the structural integrity of the piglets' intestines and decreased jejunal tight junction protein (occludin and claudin-1) expression (P < 0.05). Additionally, the pH and HCO3− in the arterial blood of piglets in the LE group were lower than those in the ME and HE groups (P < 0.05). Interestingly, the LE diet significantly increased lysine content in piglet serum (P < 0.05), decreased the levels of arginine, leucine, glutamic acid, and alanine (P < 0.05), and inhibited the mammalian target of rapamycin complex 1 (mTORC1) pathway by decreasing the phosphorylation abundance of key proteins. In summary, the dietary electrolyte imbalance could inhibit the activation of the mTORC1 signaling pathway, which might be a key factor in the influence of the dEB on piglet growth performance and intestinal health. Moreover, second-order polynomial (quadratic) regression analysis showed that the optimal dEB of piglets in the low-protein diet was 250 to 265 mEq/kg.

Solid-state fermentation for gossypol detoxification and nutritive enrichment of cottonseed cake: A scale-up of batch fermentation process

Gossypol, a toxic polyphenolic compound, limits the use of cottonseed cake (CSC) in animal feed. Different approaches have been employed to detoxify gossypol and improve the nutritive properties of feed. Microbial fermentation improves the nutritive quality of CSC by increasing lysine content and reducing free and bound gossypol. In this study, microbial fermentation was scaled up under batch conditions using a prototype device at the capacity of 40 kg per day. The mixed fungal culture C. tropicalis + S. cerevisiae was used for fermentation. An industrial trial was taken to ascertain the gossypol detoxification efficiency. The fermented CSC obtained under scale-up process had 60 to 80% and 40 to 60% reduction of free and bound gossypol, respectively, compared with raw CSC. The fermented CSC demonstrated an increase in crude protein (4 to 12%) and lysine (0.3 to 0.4%) and decrease in crude fibre (3 to 11%). The fermented CSC met the standards of US Food and Drug Administration in terms of its nutritional property. Thus, the simple method described in this study could be adopted for the production of detoxified CSC for use in the animal feed industry.

RNA-Seq Revealed the Molecular Mechanism of Nutritional Quality Improvement in o16-wx Double-Mutation Maize

The enhancement of the nutritional composition of maize grains could be achieved by the introduction of a mutation with a heightened lysine content. To obtain double-recessive mutant lines for the o16 and wx genes, a molecular-marker-assisted selection technique was used to backcross them into conventional maize lines. The resultant maize was subsequently utilized to explore the molecular mechanism responsible for the maize’s nutritional quality. Based on this, an RNA-seq investigation was conducted using the employing kernels during the development period of maize kernel (18, 28, and 38 DAP) to examine the gene expression involved in amino metabolism. The results revealed that a total of 27 and 34 differentially expressed genes (DEGs) were identified in tryptophan metabolism and lysine metabolism, respectively, across three time periods. In the lysine synthesis pathway, the genes encoding AK, ASD, and DapF were found to be up-regulated at various stages, encouraging lysine synthesis. Conversely, in the lysine degradation pathway, the genes encoding ALDH7A1 and LKR/SDH were down-regulated, suggesting an increase in lysine content. In the process of tryptophan metabolism, the down-regulation of genes encoding TAA and ALDH led to an increase in tryptophan content. In addition, the down-regulation of genes encoding α-zein resulted in a decrease in zein content, thereby enhancing the nutritional quality of maize. These findings hold substantial significance for elucidating the transcriptional-level molecular mechanism, underlying the accumulation of o16 and wx genes to improve maize grain quality, as well as offering valuable insight for the development of biomarkers and gene editing.

Multi-omics analysis provides insights into lysine accumulation in quinoa (Chenopodium quinoa Willd.) sprouts

Lysine, the first limiting essential amino acid, the deficiency of which seriously affects the health of human and animals. In this study, quinoa germination significantly increased the nutrients, especially lysine content. To better understanding the underlying molecular mechanism of lysine biosynthesis, isobaric tags for relative and absolute quantitation (iTRAQ)-based proteomics, RNA-sequencing (RNA-Seq) technology and liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) platform-based phytohormones analyses were conducted. Through proteome analyses, a total of 11,406 differentially expressed proteins were identified, which were mainly related to secondary metabolites. The lysine-rich storage globulins and endogenous phytohormones probably contributed the increased lysine content in quinoa during germination. Furthermore, aspartic acid semialdehyde dehydrogenase is essential for lysine synthesis in addition to aspartate kinase and dihydropyridine dicarboxylic acid synthase. Protein-protein interaction analysis indicated lysine biosynthesis is associated with “amino metabolism” and “starch and sucrose metabolism”. Above all, our study screens the candidate genes participated in lysine accumulation and explores the factors affected lysine biosynthesis by multi-omics analysis. These information not only paves a foundation for breeding lysine-rich quinoa sprouts but also provides valuable multi-omics resource to explore the characteristic of nutrients during quinoa germination.

Large and stable genome edits at the sorghum alpha kafirin locus result in changes in chromatin accessibility and globally increased expression of genes encoding lysine enrichment.

Sorghum is a resilient and widely cultivated grain crop used for feed and food. However, it's grain is deficient in lysine, an essential amino acid. This is due to the primary seed storage proteins, the alpha-kafirins, lacking lysine. It has been observed that reductions in alpha-kafirin protein results in rebalancing of the seed proteome and a corresponding increase in non-kafirin proteins which leads to an increased lysine content. However, the mechanisms underlying proteome rebalancing are unclear. This study characterizes a previously developed gene edited sorghum line, with deletions at the alpha kafirin locus. A single consensus guide RNA leads to tandem deletion of multiple members of the gene family in addition to the small target site mutations in remaining genes. RNA-seq and ATAC-seq were utilized to identify changes in gene expression and chromatin accessibility in developing kernels in the absence of most alpha-kafirin expression. Several differentially accessible chromatin regions and differentially expressed genes were identified. Additionally, several genes upregulated in the edited sorghum line were common with their syntenic orthologues differentially expressed in maize prolamin mutants. ATAC-seq showed enrichment of the binding motif for ZmOPAQUE 11, perhaps indicating the transcription factor's involvement in the kernel response to reduced prolamins. Overall, this study provides a resource of genes and chromosomal regions which may be involved in sorghum's response to reduced seed storage proteins and the process of proteome rebalancing.

Effect of number of lysine motifs on the bactericidal and hemolytic activity of short cationic antimicrobial peptides.

Antimicrobial peptides (AMPs) are vital components of the nonspecific immune system that represent a promising broad-spectrum alternative to conventional antibiotics. Several short cationic antimicrobial peptides show highly effective antibacterial activity and low hemolytic activity, which are based on the action of a few critical amino acids, such as phenylalanine (F) and lysine (K). Previous studies have reported that Fmoc-based phenylalanine peptides possess appreciable antibacterial potency against Gram-positive bacteria, but their ability to kill Gram-negative bacteria was suboptimal. In this study, we designed and prepared a series of Fmoc-KnF peptide (n=1-3) series by adding lysine motifs to strengthen their broad-spectrum antibacterial activity. The effect was investigated that the amount of lysine in Fmoc-F peptides on their antibacterial properties and hemolytic activities. Our results showed that the Fmoc-KKF peptide holds the strongest antimicrobial activity against both Gram-positive and negative bacteria among all designed peptides, as well as low hemolytic activity. These results provide support for the general strategy of enhancing the broad-spectrum antibacterial activity of AMPs through increased lysine content.

Effect of defatted soy and peanut flour obtained by new aqueous method on quality of gluten‐free cookies

Gluten-free cookies (GFCs) are promoted for avoiding gluten-related celiac disease. Requirement of protein/lysine enhancement in rice GFCs encouraged this study. Partly substituting rice flour by defatted peanut (DPF) or soy (DSF) and their mixture (1+1) significantly increased protein/lysine content in GFCs. DSF had the most effect of increasing lysine content. Adding DPF or DSF and their mixture significantly improved GFCs hardness and hedonic score. The hedonic score of overall acceptability, taste, crispiness, texture/grain, flavor, and appearance/color of all cookies studied were within very-like scale. Adding DPF or DSF or their mixture produced GFCs with deeper/darker color while DSF had the greatest effect. DPF or DSF or their mixture had dosage-dependent effect of increasing GFCs antioxidant capacity while DSF had the greatest effect. DPF, DSF or DPSFM was also good ingredient to make gluten-free and sugar-free cookies sweetened by meso-erythritol blended with stevia glycosides.

Proteome and Nutritional Shifts Observed in Hordein Double-Mutant Barley Lines.

Lysine is the most limiting essential amino acid in cereals, and efforts have been made over the decades to improve the nutritional quality of these grains by limiting storage protein accumulation and increasing lysine content, while maintaining desired agronomic traits. The single lys3 mutation in barley has been shown to significantly increase lysine content but also reduces grain size. Herein, the regulatory effect of the lys3 mutation that controls storage protein accumulation as well as a plethora of critically important processes in cereal seeds was investigated in double mutant barley lines. This was enabled through the generation of three hordein double-mutants by inter-crossing three single hordein mutants, that had all been backcrossed three times to the malting barley cultivar Sloop. Proteome abundance measurements were integrated with their phenotype measurements; proteins were mapped to chromosomal locations and to their corresponding functional classes. These models enabled the prediction of previously unknown points of crosstalk that connect the impact of lys3 mutations to other signalling pathways. In combination, these results provide an improved understanding of how the mutation at the lys3 locus remodels cellular functions and impact phenotype that can be used in selective breeding to generate favourable agronomic traits.

Influence of Potato Flour on Dough and Steamed Bread Quality and Correlation Analysis

AbstractPotato offers numerous health benefits that have yet to be utilized by the food industry. To evaluate the use of potato as an ingredient in staple food products, various proportions of potato flour were added to wheat flour (0–50 %) to form potato–wheat flour mixtures. The mixtures were used to make dough and steamed bread that were tested based on several established quality indicators. Specifically, with increasing potato flour proportion above 20 %, dough quality indicators of extensibility, tensile resistance, fermentation volume and fermentation activity exhibited downward trends, with worsening microstructural, while steamed bread exhibited decreasing specific volume, decreasing brightness, worsening textural properties, increasing yellowness and increasing lysine content. However, for dough and steamed bread made from flour mixtures containing 10–20 % potato flour, quality indicators were not significantly different from corresponding control values (100 % wheat flour). Notably, correlation analysis indicated that tensile and fermentative dough properties significantly correlated with resulting steamed bread quality. Therefore, only data measurements for these dough indicators are needed to predict steamed bread quality, in order to reduce testing workload during evaluation of ingredient formulations for steamed bread production.

The Zea mays mutants opaque2 and opaque16 disclose lysine change in waxy maize as revealed by RNA-Seq

In maize, opaque2 (o2) and opaque16 (o16) alleles can increase lysine content, while the waxy (wx) gene can enhance the amylopectin content of grains. In our study, o2 and o16 alleles were backcrossed into waxy maize line (wxwx). The o2o2o16o16wxwx lines had amylopectin contents similar to those of waxy line. Their nutritional value was better than waxy line, but the mechanism by which the o2 and o16 alleles increased the lysine content of waxy maize remained unclear. The o2o2o16o16wxwx lines and their parents on kernels (18th day after pollination) were subjected to RNA sequencing (RNA-Seq). The RNA-Seq analysis revealed 272 differentially expressed genes (DEGs). Functional analyses revealed that these DEGs were mainly related to biomass metabolism. Among them, in o2o2o16o16wxwx lines, 15 genes encoding α-zein were down-regulated, which resulted in the reduction of α-zein synthesis and increased lysine content; lkr/sdh1 and Zm00001d020984.1 genes involved in the lysine degradation pathway were down-regulated, thereby inhibited lysine degradation; sh2, bt2 and ae1 genes involved in starch metabolism were upregulated, leaded to wrinkling kernel and farinaceous endosperm. Our transcriptional-level identification of key genes responsible for increased grain lysine content and farinaceous endosperm formation following introgression of o2 and o16 alleles should promote molecular breeding for maize quality.

Modeling and optimizing the effect of extrusion processing parameters on nutritional properties of soy white flakes-based extrudates using response surface methodology

Thermomechanical effects of extrusion cooking leads to physiochemical changes that may be either beneficial or detrimental. The effects of extrusion processing on the nutritional value of food constituents, specifically protein, as well as the ultimate product are ambiguous. During the extrusion process, peptide bonds are hydrolyzed and molecular structure of protein changes, which can reduce the nutritional value of the protein and amino acids. Our objective was to study the effects of inclusion of soy white flakes (SWF, up to 500 g/kg) and other extrusion processing parameters such as moisture content of feed, barrel temperature and screw speed on the nutritional properties of aquafeed extrudates using a single-screw extruder and to optimize the processing conditions. Response surface regression models were established to correlate the nutritional properties of extrudates to the process variables. Using response surface methodology, extrusion processing was optimized for the maximum values of the analyzed nutritional properties and minimum values of trypsin inhibitor activity (TIA). The proximate composition, lysine content and TIA were significantly (P < 0.05) affected by the process variables. SWF was the most significant variable with quadratic effects on most of the properties. An increase in lysine content, TIA, protein content, ash content and decrease in fat content was observed when SWF was increased from 200 to 500 g/kg in the feed. Increasing temperature resulted in decrease in lysine content and TIA of extrudates. Our study shows that aquafeed with maximum lysine content and minimum trypsin inhibitor activity can be obtained by adding SWF up to 500 g/kg.

Molecular Mechanisms Underlying Increase in Lysine Content of Waxy Maize through the Introgression of the opaque2 Allele.

The opaque2 (o2) mutation in maize is associated with high lysine content in endosperm and good nutritional value. To improve the nutritional quality of waxy maize, the o2 allele was introgressed into the wxwx line using marker-assisted backcrossing selection technology. The lysine content of o2o2wxwx lines was higher than that of the wxwx line. To reveal the mechanism of increasing lysine content through introgression of the o2 in waxy maize, the transcriptome on kernels (18th day after pollination) of the o2o2wxwx and parent lines was analyzed using RNA-sequencing (RNA-Seq). The RNA-Seq analysis revealed 49 differentially expressed genes (DEGs). Functional analysis showed that these DEGs were mostly related to the catalytic activity and metabolic processes. The O2 gene regulated multiple metabolic pathways related to biological processes (BP) and molecular function (MP) during waxy maize endosperm development. In particular, in the o2o2wxwx lines, the two genes that encode the EF-1α and LHT1 were up-regulated, but the gene that encodes sulfur-rich proteins was down-regulated, raising the grain lysine content. These findings are of great importance for understanding the molecular mechanism underlying the lysine content increase due to o2 allele introgression into waxy maize.

Mapping the Increased Protein Digestibility Trait in the High‐Lysine Sorghum Mutant P721Q

Sorghum [Sorghum bicolor (L.) Moench] is a dietary staple for &gt;500 million people in over 30 countries of the semiarid tropics. Unlike other major cereals, proteins in sorghum grain become much less digestible after wet cooking. P721Q is a high‐lysine sorghum mutant that exhibits a three‐ to fourfold increase in protein digestibility after cooking as compared with other sorghum cultivars. While its increased lysine content has been traced to decreased levels of a kafirin storage protein, the goal of this study was to identify the genetic determinant of the increased digestibility. Genome sequencing of bulked segregants from a P721Q × BTx623 mapping population revealed the highly digestible protein trait maps to the same cluster of kafirin genes that contain the high‐lysine mutation.

Biofortification of rice with the essential amino acid lysine: molecular characterization, nutritional evaluation, and field performance.

Rice (Oryza sativa L.), a major staple crop worldwide, has limited levels of the essential amino acid lysine. We previously produced engineered rice with increased lysine content by expressing bacterial aspartate kinase and dihydrodipicolinate synthase and inhibiting rice lysine ketoglutarate reductase/saccharopine dehydrogenase activity. However, the grain quality, field performance, and integration patterns of the transgenes in these lysine-enriched lines remain unclear. In the present study, we selected several elite transgenic lines with endosperm-specific or constitutive regulation of the above key enzymes but lacking the selectable marker gene. All target transgenes were integrated into the intragenic region in the rice genome. Two pyramid transgenic lines (High Free Lysine; HFL1 and HFL2) with free lysine levels in seeds up to 25-fold that of wild type were obtained via a combination of the above two transgenic events. We observed a dramatic increase in total free amino acids and a slight increase in total protein content in both pyramid lines. Moreover, the general physicochemical properties were improved in pyramid transgenic rice, but the starch composition was not affected. Field trials indicated that the growth of HFL transgenic rice was normal, except for a slight difference in plant height and grain colour. Taken together, these findings will be useful for the potential commercialization of high-lysine transgenic rice.

Solubility Behavior of Cyanophycin Depending on Lysine Content

Study of the synthesis of cyanophycin (CGP) in recombinant organisms focused for a long time mostly on the insoluble form of CGP, due to its easy purification and its putative use as a precursor for biodegradable chemicals. Recently, another form of CGP, which, in contrast to the insoluble form, was soluble at neutral pH, became interesting due to its high lysine content, which was also assumed to be the reason for the solubility of the polymer. In this study, we demonstrate that lysine incorporated into insoluble CGP affected the solubility of the polymer in relation to its lysine content. Insoluble CGP can be separated along a temperature gradient of 90°C to 30°C, where CGP showed an increasing lysine content corresponding to a decreasing temperature needed for solubilization. CGP with less than 3 to 4 mol% lysine did not become soluble even at 90°C, while CGP with 31 mol% lysine was soluble at 30°C. In lysine fractions at higher than 31 mol%, CGP was soluble. The temperature separation will be suitable for improving the downstream processing of CGP synthesized in large-scale fermentations, including faster and more efficient purification of CGP, as well as enrichment and separation of dipeptides and CGP with specific amino acid compositions.

Increased Lysine Content Is the Main Characteristic of the Soluble Form of the Polyamide Cyanophycin Synthesized by Recombinant Escherichia coli

Cyanophycin, a polyamide of cyanobacterial or noncyanobacterial origin consisting of aspartate, arginine, and lysine, was synthesized in different recombinant strains of Escherichia coli expressing cphA from Synechocystis sp. strain PCC 6308 or PCC 6803, Anabaena sp. strain PCC 7120, or Acinetobacter calcoaceticus ADP1. The molar aspartate/arginine/lysine ratio of the water-soluble form isolated from a recombinant strain expressing CphA6308 was 1:0.5:0.5, with a lysine content higher than any ever described before. The water-insoluble form consisted instead of mainly aspartate and arginine residues and had a lower proportion of lysine, amounting to a maximum of only 5 mol%. It could be confirmed that the synthesis of soluble cyanobacterial granule polypeptide (CGP) is independent of the origin of cphA. Soluble CGP isolated from all recombinant strains contained a least 17 mol% lysine. The total CGP portion of cell dry matter synthesized by CphA6308 from recombinant E. coli was about 30% (wt/wt), including 23% (wt/wt) soluble CGP, by using terrific broth complex medium for cultivation at 30°C for 72 h. Enhanced production of soluble CGP instead of its insoluble form is interesting for further application and makes recombinant E. coli more attractive as a suitable source for the production of polyaspartic acid or dipeptides. In addition, a new low-cost, time-saving, effective, and common isolation procedure for mainly soluble CGP, suitable for large-scale application, was established in this study.

Role of arginine and lysine in the antimicrobial mechanism of histone‐derived peptides

Antimicrobial peptides can kill microbes via various mechanisms including cell lysis or translocation and interaction with intracellular components. Understanding how particular residues affect peptide mechanisms would advance the development of improved drugs. Previous research has shown translocating ability to be promoted by increased arginine content and impaired by increased lysine content in cell‐pentetrating peptides. However, there is little research in this area on antimicrobial peptides. To this end, this study analyzed the antimicrobial mechanism of three histone‐derived peptides – buforin II, DesHDAP1 and parasin – in comparison to their R→K and K→R mutants that contained only lysine or arginine, respectfully, for their positive residues. Analogous to cell‐penetrating peptides, the antibacterial activity of these peptides increased with increased arginine content. These changes can be explained through differences in the translocating, membrane permeabilizing and DNA‐binding ability of peptides. Experimental results were reinforced with explicit molecular dynamics simulations using a 3:1 POPC:POPG membrane. This work was supported by National Institute of Allergy and Infectious Diseases (NIH/NIAID) award R15AI079685.

Increasing Lysine Content of Waxy Maize through Introgression of Opaque-2 and Opaque-16 Genes Using Molecular Assisted and Biochemical Development

The low lysine content of waxy maize cannot meet the nutritional requirements of humans, livestock, or poultry. In the present study, the high-lysine genes o2 and o16 were backcrossed into wx lines using the maize high-lysine inbreds TAIXI19 (o2o2) and QCL3021 (o16o16) as donors and the waxy maize inbred line QCL5019 (wxwx) as a receptor. In the triple-cross F1, backcross, and inbred generations, the SSR markers phi027 and phi112 within the wx and o2 genes and the SSR marker umc1121 linked to the o16 gene were used for foreground selection. Background selection of the whole-genome SSR markers was performed for the selected individuals. The grain lysine content was determined using the dye-binding lysine method. The waxiness of the grain was determined with the I2-KI staining and dual-wavelength spectrophotometric analysis. The BC2F2 generation included 7 plants of genotype wxwxo2o2O16_, 19 plants of genotype wxwxo16o16O2_, and 3 plants of genotype wxwxo2o2o16o16. In these seeds, the average amylopectin content was 96.67%, 96.87%, and 96.62%, respectively, which is similar to that of QCL5019. The average lysine content was 0.555%, 0.380%, and 0.616%, respectively, representing increases of 75.1%, 19.9%, 94.3%, respectively, over QCL5019. The average genetic background recovery rate of the BC2F3 families was 95.3%, 94.3%, 94.2%, respectively. Among these 3 wxwxo2o2O16O16 families, 4 wxwxo2o2O16o16 families, and 3 wxwxo2o2o16o16 families, the longest imported parent donor fragment was 113.35 cM and the shortest fragment was 11.75 cM. No significant differences in lysine content were found between the BC2F4 seeds and the BC2F3 seeds in these 10 families. This allowed us to increase the lysine content of waxy corn and produce seeds with excellent nutritional characteristics suitable for human consumption, animal feed, and food processing. This may be of significance in the breeding of high-quality corn and in improvement of the nutrition of humans, livestock, and poultry.