Fine-mapping and primary analysis of candidate genes associated with seed coat color in mung bean(Vigna radiata L.)

Seed coat color affects the appearance and commodity quality of mung beans(Vigna radiata L.).The substances that affect mung bean seed coat color are mainly flavonoids,which have important medicinal value.Mapping the seed coat color gene in mung beans would facilitate the development of new varieties and improve their value.In this study,an F2 mapping population consisting of 546 plants was constructed using Jilv9(black seed coat)and BIS9805(green seed coat).Using bulk segregated analysis(BSA)sequencing and kompetitive allele-specific PCR(KASP)markers,the candidate region related to seed coat color was finally narrowed to 0.66 Mb on chromosome(Chr.)4 and included eight candidate genes.Combined transcriptome and metabolome analyses showed that three of the eight candidate genes(LOC106758748,LOC106758747,and LOC106759075)were differentially expressed,which may have caused the differences in flavonoid metabolite content between Jilv9 and BIS9805.These findings can provide a research basis for cloning the genes related to seed coat color and accelerate molecular markerassisted selection breeding in mung beans.

Segregation Analysis on Genetic System of Quantitative Traits in Plants

Based on the traditional polygene inheritance model of quantitative traits,the author suggests the major gene and polygene mixed inheritance model.The model was considered as a general one,while the pure major gene and pure polygene inheritance model was a specific case of the general model.Based on the proposed theory,the author established the segregation analysis procedure to study the genetic system of quantitative traits of plants.At present,this procedure can be used to evaluate the genetic effect of individual major genes(up to two to three major genes),the collective genetic effect of polygene,and their heritability value.This paper introduces how to establish the procedure,its main achievements,and its applications.An example is given to illustrate the steps,methods,and effectiveness of the procedure.

Integrated genomic and transcriptomic analysis reveals genes associated with plant height of foxtail millet

Foxtail millet(Setaria italica)is an important C4 model crop;however,due to its high-density planting and high stature,lodging at the filling stage resulted in a serious reduction in yield and quality.Therefore,it is imperative to identify and deploy the genes controlling foxtail millet plant height.In this study,we used a semi-dwarf line 263A and an elite high-stalk breeding variety,Chuang 29 to construct an F2 population to identify dwarf genes.We performed transcriptome analysis(RNA-seq)using internode tissues sampled at three jointing stages of 263A and Chuang 29,as well as bulk segregant analysis(BSA)on their F2 population.A total of 8918 differentially expressed genes(DEGs)were obtained from RNA-seq analysis,and GO analysis showed that DEGs were enriched in functions such as‘‘gibberellin metabolic process”and‘‘oxidoreductase activity”,which have previously been shown to be associated with plant height.A total 593 mutated genes were screened by BSA-seq method.One hundred and seventy-six out of the 593 mutated genes showed differential expression levels between the two parental lines,and seven genes not only showed differential expression in two or three internode tissues but also showed high genomic variation in coding regions,which indicated they play a crucial role in plant height determination.Among them,we found a gibberellin biosynthesis related GA20 oxidase gene(Seita.5G404900),which had a single-base at the third exon,leading to the frameshift mutation at 263A.Cleaved amplified polymorphic sequence assay and association analysis proved the single-base in Seita.5G404900 co-segregated with dwarf phenotype in two independent F2 populations planted in entirely different environments.Taken together,the candidate genes identified in this study will help to elucidate the genetic basis of foxtail millet plant height,and the molecular marker will be useful for marker-assisted dwarf breeding.

Joint Analysis Method for Major Genes Controlling Multiple Correlated Quantitative Traits

Based on the major gene and polygene mixed inheritance model for multiple correlated quantitative traits, the authors proposed a new joint segregation analysis method of major gene controlling multiple correlated quantitative traits, which include major gene detection and its effect and variation estimation. The effect and variation of major gene are estimated by the maximum likelihood method implemented via expectation-maximization （EM） algorithm. Major gene is tested with the likelihood ratio （LR） test statistic. Extensive simulation studies showed that joint analysis not only increases the statistical power of major gene detection but also improves the precision and accuracy of major gene effect estimates. An example of the plant height and the number of tiller of F2 population in rice cross Duonieai x Zhonghua 11 was used in the illustration. The results indicated that the genetic difference of these two traits in this cross refers to only one pleiotropic major gene. The additive effect and dominance effect of the major gene are estimated as -21.3 and 40.6 cm on plant height, and 22.7 and -25.3 on number of tiller, respectively. The major gene shows overdominance for plant height and close to complete dominance for number of tillers.

Genetic Analysis and QTL Mapping of Large Flag Leaf Angle Trait in Japonica Rice

Genetic segregation analysis for flag leaf angle was conducted using six generations of P1, P2, F1, B1, B2 and F2 derived from a cross of 863B （a maintainer line of japonica rice） and A7444 （a germplasm with large flag leaf angle）. Genotypes and phenotypes of flag leaf angle were investigated in 863B （P1）, A7444 （P2） and 141 plants in BC^F~ （863BIA744411863B） population. An SSR genetic linkage map was constructed and QTLs for flag leaf angle were detected. The genetic map containing 79 information loci was constructed, which covers a total distance of 441.6 cM, averaging 5.6 cM between two neighboring loci. Results showed that the trait was controlled by two major genes plus polygene and the major genes were more important. Fifteen markers showed highly significant correlations with flag leaf angle based on single marker regression analysis. Two QTLs （qFLA2 and qFLA8） for flag leaf angle were detected by both composite interval method in software WinQTLCart 2.5 and composite interval method based on mixed linear model in QTL Network 2.0. The qFLA2 explained 10.50% and 13.28% of phenotypic variation, respectively, and was located at the interval of RM300 and RM145 on the short arm of chromosome 2. The qFLA8 explained 9.59% and 7.64% of phenotypic variation, respectively, and was located at the interval flanking RM6215 and RM8265 on the long arm of chromosome 8. The positive alleles at the two QTLs were both contributed from A7444.

Fine-mapping of a candidate gene for web blotch resistance in Arachis hypogaea L.

Peanut(Arachis hypogaea L.)is a globally important oil crop.Web blotch is one of the most important foliar diseases affecting peanut,which results in serious yield losses worldwide.Breeding web blotch-resistant peanut varieties is the most effective and economically viable method for minimizing yield losses due to web blotch.In the current study,a bulked segregant analysis with next-generation sequencing was used to analyze an F2:3 segregating population and identify candidate loci related to web blotch resistance.Based on the fine-mapping of the candidate genomic interval using kompetitive allele-specific PCR(KASP)markers,we identified a novel web blotch resistance-related locus spanning approximately 169 kb on chromosome 16.This region included four annotated genes,of which only Arahy.35VVQ3 had a non-synonymous single nucleotide polymorphism in the coding region between the two parents.Two markers(Chr.16.12872635 and Chr.16.12966357)linked to this gene were shown to be co-segregated with the resistance of peanut web blotch by 72 randomly selected recombinant inbred lines(RIL),which could be used in marker-assisted breeding of resistant peanut varieties.

A 1-bp deletion in the MC04g1399 is highly associated with failure to produce fruit wart in bitter gourd

Fruit wart is an important appearance trait influencing consumer preferences of bitter gourd(Momordica charantia L.).The molecular genetic mechanisms underlying fruit wart formation in bitter gourd are largely unknown.In this study,genetic analysis based on four generations showed that fruit wart formation in bitter gourd was controlled by a single dominant locus named as Fwa.The Fwa locus was initially mapped into a 4.82 Mb region on pseudochromosome 4 by BSA-seq analysis and subsequently narrowed down to a 286.30 kb region by linkage analysis.A large F2population consisting of 2360 individuals was used to screen recombinants,and the Fwa locus was finally fine mapped into a 22.70 kb region harboring four protein-coding genes through recombination analysis.MC04g1399,encoding an epidermal patterning factor 2-like protein,was proposed as the best candidate gene for Fwa via sequence variation and expression analysis.In addition,a 1-bp insertion and deletion(InDel)variation within MC04g1399 was converted to a cleaved amplified polymorphic sequence(CAPS)marker that could precisely distinguish between the warty and non-warty types with an accuracy rate of 100%among a wide panel of 126 bitter gourd germplasm resources.Our results not only provide a scientific basis for deciphering the molecular mechanisms underlying fruit wart formation but also provide a powerful tool for efficient genetic improvement of fruit wart via marker-assisted selection.

Genetic Analysis of Main Plant Type-related Traits in Bitter Gourd(Momordica charantia L.)

In this study,the cultivated bitter gourd inbred line‘Foli 112’and the wild bitter gourd accession THMC170 were used as female(P1)and male(P2)parent,respectively,to obtain 4 generations(P1,P2,F1 and F2)through self-crossing and hybridization.The mixed major gene plus polygene inheritance model of plant quantitative trait was used to perform genetic analysis for 5 plant type-related traits including the number of internode,leaf length,leaf width,stem width and internode length in bitter gourd.Results showed that the inheritance of internode number,leaf length and leaf width was in accordance with E-6 model which involved two pairs of equi-dominant major genes with additive-dominant polygene model;the stem width was controlled by additive-dominance-epistasis polygene inheritance model(C-0 model);the inheritance of internode length involved 2 pairs of equi-additive major genes with additive-dominant polygene model(E-4 model).Meanwhile,the heritability(h2)of stem width and internode number was 20.08%and 39.28%,respectively,indicating that they were vulnerable to environment impacts;leaf length,leaf width and internode length had a higher heritability of 51.50%,52.16%and 64.36%,respectively;the inheritance of leaf length was mainly controlled by major gene(the heritability of major gene was 40.75%),indicating that early-generation selection was better for leaf length,while the inheritance of leaf width and internode length was mainly controlled by mutiple minor genes,indicating that it was better to select these two parameters for late generation.

Bulked Segregant Analysis to Detect QTL Related to Heat Tolerance in Rice(Oryza sativa L.) Using SSR Markers

The study was undertaken to assess the genetic effect of quantitative trait loci （QTLs） conferring heat tolerance at flowering stage in rice. A population consisting of 279 F2 individuals from the cross between 996, a heat tolerant cultivar and 4628, a heat-sensitive cultivar, was analyzed for their segregation pattern of the difference of seed set rate under optimal temperature condition and high temperature condition. The difference of seed set rate under optimal temperature condition and high temperature condition showed normal distribution, indicating the polygenic control over the trait. To identify main effect of QTL for heat tolerance, the parents were surveyed with 200 primer pairs of simple sequence repeats （SSR）. The parental survey revealed 30% polymorphism between parents. In order to detect the main QTL association with heat tolerance, a strategy of combining the DNA pooling from selected segregants and genotyping was adopted. The association of putative markers identified based on DNA pooling from selected segregants was established by single marker analysis （SMA）. The results of SMA revealed that SSR markers, RM3735 on chromosome 4 and RM3586 on chromosome 3 showed significant association with heat tolerance respectively, accounted for 17 and 3% of the total variation respectively. The heat tolerance during flowering stage in rice was controlled by multiple gene. The SSR markers, RM3735 on chromosome 4 and RM3586 on chromosome 3 showed significant association with heat tolerance respectively, accounted for 17 and 3% of the total variation respectively. The two genetic loci, especially for RM3735 on chromosome 4, can be used in marker-assistant-selected method in heat tolerance breeding in rice.

Mapping of wheat stripe rust resistance gene Yr041133 by BSR-Seq analysis

Puccinia striiformis Westend. f. sp. tritici(Pst) pathotype CYR34 is widely virulent and prevalent in China.Here, we report identification of a strpie rust resistance(Yr) gene, designated Yr041133, in winter wheat line 041133. This line produced a hypersensitive reaction to CYR34 and conferred resistance to 13 other pathotypes. Resistance to CYR34 in line 041133 was controlled by a single dominant gene. Bulked segregant RNA sequencing(BSR-Seq) was performed on a pair of RNA bulks generated by pooling resistant and susceptible recombinant inbred lines. Yr041133 was mapped to a 1.7 c M genetic interval on the chromosome arm 7 BL that corresponded to a 0.8 Mb physical interval(608.9–609.7 Mb) in the Chinese Spring reference genome. Based on its unique physical location Yr041133 differred from the other Yr genes on this chromosome arm.

Evaluation and Bulked Segregant Analysis of Major Yield QTL qtl12.1 Introgressed into Indigenous Elite Line for Low Water Availability under Water Stress

Near isogenic lines carrying large-effect QTL （qtl2.1）, which has a consistent influence on grain yield under upland drought stress conditions in a wide range of environments, were evaluated under water stress in the fields. The line which gave higher yield under drought was crossed with a local elite line, PMK3, and forwarded to F2：3 generation. Significant variation was found among the F2：3 lines for agronomic traits under water stress in the fields. Low to high broad sense heritability （H） for investigated traits was also found. Water stress indicators such as leaf rolling and leaf drying were negatively correlated with plant height, biomass and grain yield under stress. Bulked segregant analysis （BSA） was performed with the markers in the vicinity of qUl2.1, and RM27933 was found to be segregated perfectly well in individual components of drought resistant and drought susceptible bulks which were bulked based on yield under water stress among F2：3 lines. Hence, this simple and breeder friendly marker, RM27933, may be useful as a potentially valuable candidate marker for the transfer of the QTL qtl12.1 in the regional breeding program. Bioinformatic analysis of the DNA sequence of the qtl12.1 region was also done to identify and analyze positional candidate genes associated with this QTL and to ascertain the putative molecular basis of qUl2.1.

Quantitative trait loci detection of E dwardsiella tarda resistance in Japanese flounder Paralichthys olivaceus using bulked segregant analysis

In recent years, Edwardsiella tarda has become one of the most deadly pathogens of Japanese fl ounder( Paralichthys olivaceus), causing serious annual losses in commercial production. In contrast to the rapid advances in the aquaculture of P. o livaceus, the study of E. tarda resistance-related markers has lagged behind, hindering the development of a disease-resistant strain. Thus, a marker-trait association analysis was initiated, combining bulked segregant analysis(BSA) and quantitative trait loci(QTL) mapping. Based on 180 microsatellite loci across all chromosomes, 106 individuals from the F1333(♀: F0768 ×♂: F0915)(Nomenclature rule: F+year+family number) were used to detect simple sequence repeats(SSRs) and QTLs associated with E. tarda resistance. After a genomic scan, three markers(Scaffold 404-21589, Scaffold 404-21594 and Scaffold 270-13812) from the same linkage group(LG)-1 exhibited a signifi cant difference between DNA, pooled/bulked from the resistant and susceptible groups( P <0.001). Therefore, 106 individuals were genotyped using all the SSR markers in LG1 by single marker analysis. Two different analytical models were then employed to detect SSR markers with different levels of signifi cance in LG1, where 17 and 18 SSR markers were identifi ed, respectively. Each model found three resistance-related QTLs by composite interval mapping(CIM). These six QTLs, designated q E1–6, explained 16.0%–89.5% of the phenotypic variance. Two of the QTLs, q E-2 and q E-4, were located at the 66.7 c M region, which was considered a major candidate region for E. tarda resistance. This study will provide valuable data for further investigations of E. tarda resistance genes and facilitate the selective breeding of disease-resistant Japanese fl ounder in the future.

Tagging of Brown Planthopper Resistance Genes in F_2s of IR50 × Ptb33 of Rice by Using Bulked Segregant Analysis

Brown planthopper （Nilaparvata lugens Stal） is one of the most damaging pests causing hopper burn in rice, and thereby reducing the productivity and also the quality of the product. The effective management strategy to control this pest is the identification and transfer of desirable genes to local rice cultivars. The most important approach for developing resistant cultivars is the identification of markers, which can help in marker-assisted selection of more durable resistant genotype. The susceptible parent IR50 and the resistant parent Ptb33, and their F2 populations were used in bulked segregant analysis for identification of resistant genes with random amplified polymorphic DNA marker （RAPD） primers. The primers OPC7 and OPAG14 showed both dominant and susceptible specific banding pattern so called co-dominant markers. Moreover, OPC7697 and OPAG14680 showed resistant specific bands and thus being in coupling phase, whereas OPC7846 and OPAG14650 showed susceptible specific genotypic bands in bulked segregant analysis. Therefore, the coupling phase markers, OPC7697 and OPAG14680, are considered to be more useful in marker-assisted selection of rice genotypes in crop improvement.

Analysis of bulked segregants to identify molecular markers linked with cocoon weight and cocoon shell weight in the silkworm Bombyx mori L

Two silkworm strains viz, B20 A (high cocoon shell ratio) and C.Nichi (low cocoon shell ratio) were sib mated for 10 generations to determine the homozygosis. Both bulked segregant analysis(BSA) and near isogenic lines (NIL) studies were done to identify the RFLP markers closely linked to cocoon shell parameters. Three hundred and fifty two random clones were identified as the low copy number sequence and used for identification of Restriction Fragment Length Polymorphic (RFLP) marker linked to cocoon weight and cocoon shell character. In the bulk segregant analysis, DNA from the parents (B20 A, C.Nichi), F 1 and F 2 progeny of high shell ratio (HSR) and low shell ratio (LSR) were screened for hybridization with the random clones. Polymorphic banding pattern achieved through southern hybridization with different probes indicated the probable correlation of polymorphism with high and low cocoon shell character which are possible landmarks in identifying the putative marker(s) for the cocoon shell character. Out of the 100 probes tried with parents, F 1, F 2 and their bulks, 10 probes were found to be closely linked to cocoon shell characters.

Major Gene Identification and Quantitative Trait Locus Mapping for YieldRelated Traits in Upland Cotton(Gossypium hirsutum L.)

Segregation analysis of the mixed genetic model of major gene plus polygene was used to identify the major genes for cotton yield-related traits using six generations P1, P2, F1, B1, B2, and F2 generated from the cross of Baimian 1 x TM-1. In addition to boll size and seed index, the major genes for the other five traits were detected： one each for seed yield, lint percentage, boll number, lint index; and two for lint yield. Quantitative trait locus/loci （QTL） mapping was performed in the F2 and F2：3 populations of above cross through molecular marker technology, and a total of 50 QTL （26 suggestive and 24 significant） for yield-related traits were detected. Four common QTL were discovered： qLP-3b（F2）/qLP-3（F2：3） and qLP-19b （F2）/qLP-19（F2：3） for lint percentage, qBN-17（F2）/qBN-17（F2：3） for boll number, and qBS-26b（F2）/qBS-26（F2：3） for boll size. Especially, qLP- 3b（Fz）/qLP-3（F2：3）, not only had LOD scores 〉3 but also exceeded the permutation threshold （5.13 and 5.29, respectively）, correspondingly explaining 23.47 and 29.55% of phenotypic variation. This QTL should be considered preferentially in marker assisted selection （MAS）. Segregation analysis and QTL mapping could mutually complement and verify, which provides a theoretical basis for genetic improvement of cotton yield-related traits by using major genes （QTL）.

Inheritance of marsh spot disease resistance in cranberry common bean(Phaseolus vulgaris L.)

Common bean(Phaseolus vulgaris) is an annual legume crop that is grown worldwide for its edible dry seeds and tender pods. Marsh spot(MS) of the seeds is a physio-genic stress disease affecting seed quality in beans. Studies have suggested that this disease involves a nutritional disorder caused by manganese deficiency, but the inheritance of resistance to this disease has not been reported. A biparental genetic population composed of 138 recombinant inbred lines(RILs) was developed from a cross between an MS resistant cultivar ‘Cran09’ and an MS susceptible cultivar ‘Messina’. The 138 RILs and their two parents were evaluated for MS resistance during five consecutive years from 2015 to 2019 in sandy and heavy clay soils in Morden, Manitoba, Canada. The MS incidence(MSI) and the MS resistance index(MSRI) representing disease severity were shown to be both highly correlated heritable traits that had high broad-sense heritability values(H;) of 86.5% and 83.2%, respectively. No significant differences for MSI and MSRI were observed between the two soil types in all five-(MSI) or four-year(MSRI) data collection, but significant correlations among years were observed despite MS resistance was moderately affected by year. The MSIs and MSRIs displayed a right-skewed distribution, indicating a mixed genetic model involving a few major genes and polygenes. Using the joint segregation analysis method, the same four major genes with additive-epistasis effects showed the best fit for both traits, explaining 84.4% and85.3% of the phenotypic variance for MSI and MSRI, respectively. For both traits, the M1, M2, M3 and m4 acted as the favorable(resistant) alleles for the four genes where M and m represent two alleles of each gene. However, due to epistatic effects, only the individuals of the M1 M2 M3 M4 haplotype appeared to be highly resistant, whereas those of the m1 m2 m3 M4 haplotype were the most susceptible. The m4 allele significantly suppressed the additive effects of M1 M2 M3 on resistance, but decreased susceptibility due to the additive effects of m1 m2 m3. Further quantitative trait locus(QTL) mapping is warranted to identify and validate individual genes and develop molecular markers for marker-assisted selection of resistant cultivars.

Mapping a leaf senescence gene els1 by BSR-Seq in common wheat

Leaf senescence is normally the last stage of plant development. Early senescence of functional leaves significantly reduces the photosynthetic time and efficiency, seriously affecting grain yield and quality in wheat. Discovering genes responsible for early leaf senescence(els) are necessary for developing novel germplasms and cultivars with delayed leaf-senescence through molecular manipulation and marker assisted selection. In this study, we identified an early leaf senescence line M114 in a derivative of a wheat breeding population. Genetic analysis indicated that early leaf senescence in M114 is controlled by a single recessive gene, provisionally designated els1. By applying bulked segregant analysis and RNA-Seq(BSR-Seq), seven polymorphic markers linked to els1 were developed and the gene was located on chromosome arm 2 BS in a 1.5 c M genetic interval between markers WGGB303 and WGGB305. A co-segregating marker, WGGB302, provide a starting point for fine mapping and map-based cloning of els1.

Development of Sequence Characterized Amplified Region (SCAR) Primers for the Detection of Resistance to Sporisorium reiliana in Maize

Head smut of maize （Zea mays L.）, which was caused by Sporisorium reiliana, occurred in most of the maize growing areas of the world. The purpose of this study was to develop SCAR markers for map-based cloning of resistance genes and MAS. Two sets of BC3 progenies, one （BC3Q） derived from the cross Qi319 （resistance）×Huangzao 4 （susceptible）, the other （BC3M） from Mol7 （resistance）× Huangzao 4 （susceptible）, were generated. Huangzao 4 was the recurrent parent in both progenies. A combination of BSA （bulked segregant analysis） with AFLP （amplified fragment length polymorphism） method was applied to map the genes involving the resistance to S. reiliana, and corresponding resistant and susceptible bulks and their parental lines were used for screening polymorphic AFLP primer pairs. One fragment of PI3M61-152 was converted into SCAR （sequence charactered amplified fragment） marker S130. The marker was mapped at chromosome bin 2.09, the interval of a major QTL region previously reported to contribute to S. reiliana resistance. Furthermore, S130 was highly and facilitate map-based cloni associated with resistance to S. reiliana, and could be useful for marker-assisted selection ng of resistance genes.

A Co-Dominant Marker BoE332 Applied to Marker-Assisted Selection of Homozygous Male-Sterile Plants in Cabbage (Brassica oleracea var.capitata L.)

The dominant genic male sterility （DGMS） gene CDMs399-3 derived from a spontaneous mutation in the line 79-399-3 of spring cabbage （Brassica oleracea var. capitata L.）, has been successfully applied in hybrid seed production of several cabbage cultivars in China. During the development of dominant male sterility lines in cabbage, the conventional identification of homozygous male-sterile plants （CDMs399-3/CDMs399-3） is a laborious and time-consuming process. For marker-assisted selection （MAS） of the gene CDMs399-3 transferred into key spring cabbage line 397, expressed sequence tag-simple sequence repeats （EST-SSR） and SSR technology were used to identify markers that were linked to CDMs399-3 based on method of bulked segregant analysis （BSA）. By screening a set of 978 EST-SSRs and 395 SSRs, a marker BoE332 linked to the CDMs399-3 at a distance of 3.6 cM in the genetic background of cabbage line 397 were identified. 7 homozygons male-sterile plants in population P1170 with 20 plants were obtained finally via MAS of BoE332. Thus, BoE332 will greatly facilitate the transferring of the gene CDMs399-3 into the key spring cabbage line 397 and improve the application of DGMS in cabbage hybrid breeding.

Identification RAPD Markers Linked to Texture Quality of Chinese Cabbage

[ Objectives] This study was conducted to identify the random amplification of polymorphic DNA （RAPD） markers linked to chewy texture-controlling gene of Chinese cabbage. [ Methods] The RAPD markers associated with chewy texture of Chinese cabbage were identified via bulked segregant analysis （BSA） in an F2 population derived from the cross between Hua 273 （female parent） and 114 Fushan （male parent）. [ Results] OPA06-1400 was identified to he linked to the chewy texture-controlling gene of Chinese cabbage. The genetic distance between the target gene and the RAPD marker was 24.8 cM. [ Conclusions] The resuits provide experimental evidence for breeding of Chinese cabbage.