Pub Date : 2020-09-04DOI: 10.5376/mpb.2020.11.0016
Xinrong Yang, Xudong Yu, Fanhua Wu, Zeping Cai, Jiajia Luo, P. Cao
Full-length trantogenic sequencing (PRJNA579273) of jackfruit seedling stems and leaves was performed. On this basis, biological information analysis was performed on Jackfruit ( Artocarpus heterophyllus ) BR INSENSITIVE 1 (BRI1) family members AhBRI1 , AhBRL1 and AhBRL2 to translate genes into proteins. Using online analysis tools, jackfruit, Arabidopsis (Arabidopsis Thaliana), Sichuan mulberry (Morus notabilis) and Poplar (Populus trichocarpa) were used to analyze and obtain different physicochemical properties data and the secondary and tertiary structure of the protein. The bioinformatics prediction and comparison between the BRI1 protein families of different species were obtained. The results showed that the amino acid residues were 1 195, 1 170 and 985, and the theoretical isoelectric points were 6.32, 5.77 and 7.08. The protein was weakly acidic, hydrophilic and stable. There are secretory pathway signal peptides or chloroplast transport peptides with transmembrane structure. The main components of the secondary structure are irregular coil and -helix, and the tertiary structure is spiral tubular structure. This study provides a basis for further functional studies on members of the BRI1 protein family of jackfruit.
{"title":"Bioinformatics Prediction and Comparison of Artocarpus heterophyllus BRI1 Family Members","authors":"Xinrong Yang, Xudong Yu, Fanhua Wu, Zeping Cai, Jiajia Luo, P. Cao","doi":"10.5376/mpb.2020.11.0016","DOIUrl":"https://doi.org/10.5376/mpb.2020.11.0016","url":null,"abstract":"Full-length trantogenic sequencing (PRJNA579273) of jackfruit seedling stems and leaves was performed. On this basis, biological information analysis was performed on Jackfruit ( Artocarpus heterophyllus ) BR INSENSITIVE 1 (BRI1) family members AhBRI1 , AhBRL1 and AhBRL2 to translate genes into proteins. Using online analysis tools, jackfruit, Arabidopsis (Arabidopsis Thaliana), Sichuan mulberry (Morus notabilis) and Poplar (Populus trichocarpa) were used to analyze and obtain different physicochemical properties data and the secondary and tertiary structure of the protein. The bioinformatics prediction and comparison between the BRI1 protein families of different species were obtained. The results showed that the amino acid residues were 1 195, 1 170 and 985, and the theoretical isoelectric points were 6.32, 5.77 and 7.08. The protein was weakly acidic, hydrophilic and stable. There are secretory pathway signal peptides or chloroplast transport peptides with transmembrane structure. The main components of the secondary structure are irregular coil and -helix, and the tertiary structure is spiral tubular structure. This study provides a basis for further functional studies on members of the BRI1 protein family of jackfruit.","PeriodicalId":32255,"journal":{"name":"Journal of Plant Molecular Breeding","volume":"13 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2020-09-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"77351102","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2020-07-19DOI: 10.5376/mpb.2020.11.0015
Zhang Sujun, Xiao-Yi Zhou, Tang Liyuan, Li Xinghe, Wang Haitao, Liu Cunjing, Cai Xiao, Zhang Xiangyun, Zhang Jianhong
‘Ji1518’ is a new hybrid cotton variety suitable for mechanized harvesting, the two parents of ‘Ji228’ and ‘Ji567’ were crossed to established F2, F2:3 and F2:9 (recombinant inbred lines RILs) population. Simple sequence repeats (SSR) was performed to construct two different genetic maps based on the F2 and F2:9 populations respectively. The QTL mapping of five fiber quality traits was performed in three populations above. A genetic map was constructed by F2 population, which contained 15 loci in 4 linkage groups, with a full-length coverage of 237.10 cM. While the other linkage map was constructed by F2:9 population, which contained 45 loci in 11 linkage groups, with a full-length coverage of 554.42. Based on the inclusive composite interval mapping method with QTL IciMapping 4.1, 15 QTLs related to upper half mean length, fiber strength, the micronaire value, the elongation and the uniformity were both detected in F2 and F2:3 segregating populations, among them, the QTL locus qFM-4-2 related to the micronaire value explained the highest phenotypic variation rate at 21.10%. QTLs with dominant or super-dominant effects accounted for 66.7% of the total, which showed that dominant genes were the main source of fiber quality heterosis in ‘Ji1518’. Meanwhile, 6 QTLs related to the above five traits of fiber quality were detected in the RIL (F2:9) population, and the contribution rate was between 5.10% and 10.26%. QTL loci related with FS and MIC were detected near HAU2349 in all three populations, and QTL loci related with FU were detected near HAU2710 in all three populations, and the markers above were linked on A6 chromosome. These stable and common QTLs are beneficial to the MAS breeding, which could improve the breeding efficiency.
{"title":"QTL Mapping and Genetic Analysis of Fiber Quality Traits in Hybrid Cotton","authors":"Zhang Sujun, Xiao-Yi Zhou, Tang Liyuan, Li Xinghe, Wang Haitao, Liu Cunjing, Cai Xiao, Zhang Xiangyun, Zhang Jianhong","doi":"10.5376/mpb.2020.11.0015","DOIUrl":"https://doi.org/10.5376/mpb.2020.11.0015","url":null,"abstract":"‘Ji1518’ is a new hybrid cotton variety suitable for mechanized harvesting, the two parents of ‘Ji228’ and ‘Ji567’ were crossed to established F2, F2:3 and F2:9 (recombinant inbred lines RILs) population. Simple sequence repeats (SSR) was performed to construct two different genetic maps based on the F2 and F2:9 populations respectively. The QTL mapping of five fiber quality traits was performed in three populations above. A genetic map was constructed by F2 population, which contained 15 loci in 4 linkage groups, with a full-length coverage of 237.10 cM. While the other linkage map was constructed by F2:9 population, which contained 45 loci in 11 linkage groups, with a full-length coverage of 554.42. Based on the inclusive composite interval mapping method with QTL IciMapping 4.1, 15 QTLs related to upper half mean length, fiber strength, the micronaire value, the elongation and the uniformity were both detected in F2 and F2:3 segregating populations, among them, the QTL locus qFM-4-2 related to the micronaire value explained the highest phenotypic variation rate at 21.10%. QTLs with dominant or super-dominant effects accounted for 66.7% of the total, which showed that dominant genes were the main source of fiber quality heterosis in ‘Ji1518’. Meanwhile, 6 QTLs related to the above five traits of fiber quality were detected in the RIL (F2:9) population, and the contribution rate was between 5.10% and 10.26%. QTL loci related with FS and MIC were detected near HAU2349 in all three populations, and QTL loci related with FU were detected near HAU2710 in all three populations, and the markers above were linked on A6 chromosome. These stable and common QTLs are beneficial to the MAS breeding, which could improve the breeding efficiency.","PeriodicalId":32255,"journal":{"name":"Journal of Plant Molecular Breeding","volume":"3 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2020-07-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"90757490","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2020-06-28DOI: 10.5376/mpb.2020.11.0012
Li Qiang, Li Tao, Shihuai Guo, Yuting Bai, Xing-Long Li
The content of amylose and amylopectin is an important trait affecting cooking and eating quality in millet. During process of kernel grouting, the synthesis of starch involves different pathways and components in non-waxy and waxy millets. Immature grouting grains of waxy millet ‘Gonggu68’ and non-waxy millet ‘Chigu4’ (grouting period 1 and 5 days) were used to analyze their transcriptome sequences by using Illumina Hiseq4000. The results showed that: (1) GBSSⅠ enzymes activity of waxy and non-waxy millet was low-high-low. There were some differences between the two activities. 665 upregulated differentially expressed genes were screened on day 5 and day 1 during grouting period in waxy cultivar ‘Gonggu68’, there were 431 more upregulated genes than downregulated genes. There were 97 more up-regulated genes than down-regulated genes in non-waxy cultivar ‘Chigu4’ on day 5 and day 1 in grouting period. (2) In the A 2 -VS-A 1 waxy comparison group, the differential genes were mainly GO enriched in 7 functions such as the seed oil body biogenic function, 17-β-ketosteroid reductase activity function and so on. it was mainly enriched in biological processes and molecular functions. In the B 2 -VS-B 1 non-waxy comparison group, the differential genes were mainly GO enriched in 8 functions such as the light capturing function and the pigment binding function in the light system I for non-waxy millet. (3) Differentially expressed genes were mainly KEGG enriched in caffeine metabolism pathway, linoleic acid metabolism pathway, anthocyanin biosynthesis pathway, aflatoxin biosynthesis pathway in waxy A 2 -VS-A 1 , but which were mainly KEGG enriched in the synergy-antenna protein pathway, the linoleic acid metabolic pathway, the caffeine metabolic pathway, the brassinosteroid biopathway in non-waxy B 2 -VS-B 1 . These two comparative groups were enriched Caffeine metabolism pathway and linoleic acid metabolism pathway appeared in the process. (4) Three ( SSII-3 , PHO1 , AS ) and four ( PHO1-1 , AS , AGP16 , WAXY ) genes with significant differences and related to waxy and non-waxy millet were screened. With Actin (Si001873) as the internal reference gene, the above seven differentially expressed genes were verified by qRT-PCR, which was consistent with the transcriptome results, indicating that the differentially expressed genes were related to waxy or non-waxy endosperm.
直链淀粉和支链淀粉含量是影响谷子蒸煮和食味品质的重要性状。在灌浆过程中，淀粉在非蜡粒和蜡粒中有不同的合成途径和组分。采用Illumina Hiseq4000对灌浆期为1和5 d的糯小米‘公谷68’和非糯小米‘赤谷4’的未成熟灌浆粒进行转录组序列分析。结果表明:(1)糯粒和非糯粒谷子的GBSSⅠ酶活性低-高-低。这两项活动有一些不同之处。在灌浆期第5天和第1天共筛选到665个差异表达上调基因，上调基因比下调基因多431个。无蜡品种赤谷4号在灌浆期第5天和第1天上调基因比下调基因多97个。(2) a2 - vs - a1蜡质对照组的差异基因主要为富含籽油体生物生成功能、17-β-酮类固醇还原酶活性功能等7种功能的氧化石墨烯。它主要富含生物过程和分子功能。在b2 - vs - b1无蜡对照组中，差异基因主要是在无蜡谷子光系统I中富含捕光功能和色素结合功能等8个功能的氧化石墨烯。(3)差异表达基因主要是在蜡质a2 - vs - b1中富集咖啡因代谢途径、亚油酸代谢途径、花青素生物合成途径、黄曲霉毒素生物合成途径的KEGG，而在非蜡质b2 - vs - b1中主要富集协同-触角蛋白途径、亚油酸代谢途径、咖啡因代谢途径、油菜素类固醇生物途径的KEGG。这两个对照组在这一过程中都出现了丰富的咖啡因代谢途径和亚油酸代谢途径。(4)筛选到3个(SSII-3、PHO1、AS)和4个(PHO1-1、AS、AGP16、WAXY)差异显著且与糯小米和非糯小米相关的基因。以Actin (Si001873)为内参基因，通过qRT-PCR验证上述7个差异表达基因，与转录组结果一致，说明差异表达基因与蜡质胚乳或非蜡质胚乳有关。
{"title":"Enrichment Analysis of Differentially Expressed Genes during Endosperm Grouting Periods in Non-waxy and Waxy Foxtail Millets","authors":"Li Qiang, Li Tao, Shihuai Guo, Yuting Bai, Xing-Long Li","doi":"10.5376/mpb.2020.11.0012","DOIUrl":"https://doi.org/10.5376/mpb.2020.11.0012","url":null,"abstract":"The content of amylose and amylopectin is an important trait affecting cooking and eating quality in millet. During process of kernel grouting, the synthesis of starch involves different pathways and components in non-waxy and waxy millets. Immature grouting grains of waxy millet ‘Gonggu68’ and non-waxy millet ‘Chigu4’ (grouting period 1 and 5 days) were used to analyze their transcriptome sequences by using Illumina Hiseq4000. The results showed that: (1) GBSSⅠ enzymes activity of waxy and non-waxy millet was low-high-low. There were some differences between the two activities. 665 upregulated differentially expressed genes were screened on day 5 and day 1 during grouting period in waxy cultivar ‘Gonggu68’, there were 431 more upregulated genes than downregulated genes. There were 97 more up-regulated genes than down-regulated genes in non-waxy cultivar ‘Chigu4’ on day 5 and day 1 in grouting period. (2) In the A 2 -VS-A 1 waxy comparison group, the differential genes were mainly GO enriched in 7 functions such as the seed oil body biogenic function, 17-β-ketosteroid reductase activity function and so on. it was mainly enriched in biological processes and molecular functions. In the B 2 -VS-B 1 non-waxy comparison group, the differential genes were mainly GO enriched in 8 functions such as the light capturing function and the pigment binding function in the light system I for non-waxy millet. (3) Differentially expressed genes were mainly KEGG enriched in caffeine metabolism pathway, linoleic acid metabolism pathway, anthocyanin biosynthesis pathway, aflatoxin biosynthesis pathway in waxy A 2 -VS-A 1 , but which were mainly KEGG enriched in the synergy-antenna protein pathway, the linoleic acid metabolic pathway, the caffeine metabolic pathway, the brassinosteroid biopathway in non-waxy B 2 -VS-B 1 . These two comparative groups were enriched Caffeine metabolism pathway and linoleic acid metabolism pathway appeared in the process. (4) Three ( SSII-3 , PHO1 , AS ) and four ( PHO1-1 , AS , AGP16 , WAXY ) genes with significant differences and related to waxy and non-waxy millet were screened. With Actin (Si001873) as the internal reference gene, the above seven differentially expressed genes were verified by qRT-PCR, which was consistent with the transcriptome results, indicating that the differentially expressed genes were related to waxy or non-waxy endosperm.","PeriodicalId":32255,"journal":{"name":"Journal of Plant Molecular Breeding","volume":"95 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2020-06-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"89902768","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2020-06-19DOI: 10.5376/mpb.2020.11.0002
Pengyu Zhang, Guorui Wang, X. Qiu, Bao Zhang, Li Wei, Tong-chao Wang
Virus-induced gene silencing (VIGS) is an important tool for gene function analysis in plants. In the present study, the function identification of a candidate gene named Tadhn1312 encoding Dehydrin protein obtained from the high-throughput transcriptome was carried out by VIGS. Leaves appeared the chlorotic phenotype and the transcript level of TaPDS decreased rapidly at 7 days after the inoculation by BSMV: TaPDS, which indicated that the virus had successfully infected wheat leaves and the BSMV system was efficient. The chlorophyll content of leaves with BSMV: TaPDS and BSMV: Tadhn1312 inoculation decreased at 7 days, and reached significant level compared with the control. After the inoculation by BSMV: Tadhn1312, the transcript level of Tadhn1312 was rapidly decreased at 7 days, and reached the minimum value at 21 days, indicated that Tadhn1312 had been silenced. The spike differentiation procession of wheat plants inoculated by BSMV: Tadhn1312 was late than that inoculated by BSMV: 00. The result showed that the silencing of Tadhn1312 prolonged the spike differentiation process, illustrating that Tadhn1312 gene was involved in the spike differentiation process of wheat directly or indirectly.
{"title":"Functional Analysis of Tadhn1312 by Virus-Induced Gene Silencing (VIGS) in Common Wheat (Triticum aestivum L.)","authors":"Pengyu Zhang, Guorui Wang, X. Qiu, Bao Zhang, Li Wei, Tong-chao Wang","doi":"10.5376/mpb.2020.11.0002","DOIUrl":"https://doi.org/10.5376/mpb.2020.11.0002","url":null,"abstract":"Virus-induced gene silencing (VIGS) is an important tool for gene function analysis in plants. In the present study, the function identification of a candidate gene named Tadhn1312 encoding Dehydrin protein obtained from the high-throughput transcriptome was carried out by VIGS. Leaves appeared the chlorotic phenotype and the transcript level of TaPDS decreased rapidly at 7 days after the inoculation by BSMV: TaPDS, which indicated that the virus had successfully infected wheat leaves and the BSMV system was efficient. The chlorophyll content of leaves with BSMV: TaPDS and BSMV: Tadhn1312 inoculation decreased at 7 days, and reached significant level compared with the control. After the inoculation by BSMV: Tadhn1312, the transcript level of Tadhn1312 was rapidly decreased at 7 days, and reached the minimum value at 21 days, indicated that Tadhn1312 had been silenced. The spike differentiation procession of wheat plants inoculated by BSMV: Tadhn1312 was late than that inoculated by BSMV: 00. The result showed that the silencing of Tadhn1312 prolonged the spike differentiation process, illustrating that Tadhn1312 gene was involved in the spike differentiation process of wheat directly or indirectly.","PeriodicalId":32255,"journal":{"name":"Journal of Plant Molecular Breeding","volume":"10 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2020-06-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"81678840","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2020-06-09DOI: 10.5376/mpb.2020.11.0005
Y. Cheng, W. Deng, Yelei Lu, Han Shaopeng, Y. Lv, Gongjian Zeng, Zhou Chao, Dechun Zhang, Shen Xiangling
Sorghum is one of the world's important crops after wheat, rice, maize, and barley. Although the sorghum genome had been well-sequenced, genetic breeding and functional genome research in sorghum cultivar BTx623 is still limited due to the lack of efficient and stable genetic transformation and regeneration system in sequencing. In this study, the immature embryos of sorghum genome-sequencing cultivar BTx623 was used as the explants material, and the bar gene resistant to phosphoglyphosate was used as the screening marker for Agrobacterium -mediated sorghum genetic transformation. By screening the adaptability of callus to different concentrations of phosphoglyphosate, the appropriate concentration of phosphoglyphosate in the genetic transformation of sorghum cultivar BTx623 was determined to be 2.5 mg/L, and BTx623 immature embryo was used as explants to obtain resistant callus. After screening, regenerated plants were obtained by treating resistant callus with 0.0067 mg/L ZNC in regeneration medium. Therefore, this study successfully obtained resistant callus and regenerating plants, and established a genetic transformation and regeneration system in sorghum cultivar BTx623, which may have great significance for functional genome research and genetic breeding in sorghum.
{"title":"Establishment of Sorghum BTx623 Immature Embryos Genetic Transformation and Regeneration System","authors":"Y. Cheng, W. Deng, Yelei Lu, Han Shaopeng, Y. Lv, Gongjian Zeng, Zhou Chao, Dechun Zhang, Shen Xiangling","doi":"10.5376/mpb.2020.11.0005","DOIUrl":"https://doi.org/10.5376/mpb.2020.11.0005","url":null,"abstract":"Sorghum is one of the world's important crops after wheat, rice, maize, and barley. Although the sorghum genome had been well-sequenced, genetic breeding and functional genome research in sorghum cultivar BTx623 is still limited due to the lack of efficient and stable genetic transformation and regeneration system in sequencing. In this study, the immature embryos of sorghum genome-sequencing cultivar BTx623 was used as the explants material, and the bar gene resistant to phosphoglyphosate was used as the screening marker for Agrobacterium -mediated sorghum genetic transformation. By screening the adaptability of callus to different concentrations of phosphoglyphosate, the appropriate concentration of phosphoglyphosate in the genetic transformation of sorghum cultivar BTx623 was determined to be 2.5 mg/L, and BTx623 immature embryo was used as explants to obtain resistant callus. After screening, regenerated plants were obtained by treating resistant callus with 0.0067 mg/L ZNC in regeneration medium. Therefore, this study successfully obtained resistant callus and regenerating plants, and established a genetic transformation and regeneration system in sorghum cultivar BTx623, which may have great significance for functional genome research and genetic breeding in sorghum.","PeriodicalId":32255,"journal":{"name":"Journal of Plant Molecular Breeding","volume":"3 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2020-06-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"86457973","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2020-01-01DOI: 10.5376/MPB.2020.11.0028
Luo Sainan, Wen Zhang, C. Peng, Han Jian, Li Fei-Fei, Li Xianxin
China has abundant of local citrus resources, which had many seeds. The Seedless is the aim of the local citrus breeding. Triploid citurs resources usually obtained by crosssing breeding between the diploid and tetraploid plants. However, the resources of tetraploid citrus are limited. It has been reported that the hybrid between diploid and diploid plants can produce triploid citrus, which not only saves the breeding time, but also shortens the breeding cycles. In this project, flow cytometry detection assisted by embryo rescue system and SSR identification technology were used to improve the probability of obtaining polyploidy from the hybrid offspring, and provide technical support for obtaining polyploidy from large-scale hybridization between diploid and diploid. plants. Three diploid pollen cultivars were crossbred with Hunan local cultivar 8306, immature seeds were collected for embryo rescue to obtain regenerative plants, plant ploidy was detected by flow cytometry and stoma electron microscope, and offspring genotypes were identified by SSR. After 100 d hybrid between three diploid pollens and Ponkan 8 306, 405 plants were obtained by embryo rescue technique. And 70 plants survived in the greenhouse after transplantation. Through flow cytometry instrument and stomatal electron microscope inspection, the results showed that SSR analysis validated that there are 13 polyploid plants, including 1 tetraploid plant and 12 triploid plants. The ratio of polyploidy was 18.57%. This study is obtained triploid and tetraploid plants by Ponkan embryo rescue from diploid interspecific hybridization. In this study, the embryo rescue technology system and the early identification system for polyploid hybridization were established, and a lot of new polyploid germplasm were obtained.
{"title":"Identification of Ploidy Variation of Ponkan Embryo Rescue Plants","authors":"Luo Sainan, Wen Zhang, C. Peng, Han Jian, Li Fei-Fei, Li Xianxin","doi":"10.5376/MPB.2020.11.0028","DOIUrl":"https://doi.org/10.5376/MPB.2020.11.0028","url":null,"abstract":"China has abundant of local citrus resources, which had many seeds. The Seedless is the aim of the local citrus breeding. Triploid citurs resources usually obtained by crosssing breeding between the diploid and tetraploid plants. However, the resources of tetraploid citrus are limited. It has been reported that the hybrid between diploid and diploid plants can produce triploid citrus, which not only saves the breeding time, but also shortens the breeding cycles. In this project, flow cytometry detection assisted by embryo rescue system and SSR identification technology were used to improve the probability of obtaining polyploidy from the hybrid offspring, and provide technical support for obtaining polyploidy from large-scale hybridization between diploid and diploid. plants. Three diploid pollen cultivars were crossbred with Hunan local cultivar 8306, immature seeds were collected for embryo rescue to obtain regenerative plants, plant ploidy was detected by flow cytometry and stoma electron microscope, and offspring genotypes were identified by SSR. After 100 d hybrid between three diploid pollens and Ponkan 8 306, 405 plants were obtained by embryo rescue technique. And 70 plants survived in the greenhouse after transplantation. Through flow cytometry instrument and stomatal electron microscope inspection, the results showed that SSR analysis validated that there are 13 polyploid plants, including 1 tetraploid plant and 12 triploid plants. The ratio of polyploidy was 18.57%. This study is obtained triploid and tetraploid plants by Ponkan embryo rescue from diploid interspecific hybridization. In this study, the embryo rescue technology system and the early identification system for polyploid hybridization were established, and a lot of new polyploid germplasm were obtained.","PeriodicalId":32255,"journal":{"name":"Journal of Plant Molecular Breeding","volume":"34 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2020-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"75205801","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2020-01-01DOI: 10.5376/mpb.2020.11.0017
Zhan Li, Yajie Wang, Xiaohua Lu, Ruimei Li, Jiao Liu, S. Fu, Xin-wen Hu, Guo Jianchun, Yuan Yao
Starch glucan chain structure of cassava root is the key factor to determine starch quality. Soluble starch synthase III (SSIII) is the key enzyme to regulate the synthesis of long chain in plant amylopectin glucan. Cassava has two MeSSIII homologous genes MeSSIII-1 and MeSSIII-2. To study the effect of cassava MeSSIII on the quality formation of cassava root starch, a double gene editing vector for MeSSIII-1 and MeSSIII-2 was constructed. The sgRNA target for MeSSIII-1 and MeSSIII-2 was designed simultaneously by online software CRISPR-Pv2.0 based on the conserved segments, and the recombinant pCAMBIAP1301-Cas9MeSSIII-gRNA plasmid was constructed by digestion and ligation. The gene editing vector was transformed into LBA4404 Agrobacterium competent cells and used to infect the friable embryogenic callus of cassava, and the their DNA was extracted. The target segments of MeSSIII-1 and MeSSIII-2 were amplified by PCR for Sanger sequencing, and analyzed the editing of target position. The results showed that the target sites of MeSSIII-1 and MeSSIII-2 were successfully edited. This study helps to further obtain mutants of theMeSSIII gene to analyze the role of this gene in the cassava starch synthesis pathway.
木薯根淀粉葡聚糖链结构是决定木薯淀粉品质的关键因素。可溶性淀粉合成酶III (Soluble starch synthase III, SSIII)是调控植物支链淀粉葡聚糖长链合成的关键酶。木薯有两个MeSSIII同源基因MeSSIII-1和MeSSIII-2。为研究木薯MeSSIII对木薯根淀粉品质形成的影响，构建了MeSSIII-1和MeSSIII-2双基因编辑载体。基于保守片段，利用在线软件CRISPR-Pv2.0同时设计MeSSIII-1和MeSSIII-2的sgRNA靶标，通过酶切和连接构建重组pCAMBIAP1301-Cas9MeSSIII-gRNA质粒。将该基因编辑载体转化为LBA4404农杆菌感态细胞，感染木薯脆性胚性愈伤组织，提取其DNA。通过PCR扩增出MeSSIII-1和MeSSIII-2的目标片段进行Sanger测序，并对目标位置进行编辑分析。结果表明，成功编辑了MeSSIII-1和MeSSIII-2的目标位点。本研究有助于进一步获得theMeSSIII基因的突变体，分析该基因在木薯淀粉合成途径中的作用。
{"title":"Construction and Verification of CRISPR/Cas9 Gene Editing Vector for Cassava MeSSIII Gene","authors":"Zhan Li, Yajie Wang, Xiaohua Lu, Ruimei Li, Jiao Liu, S. Fu, Xin-wen Hu, Guo Jianchun, Yuan Yao","doi":"10.5376/mpb.2020.11.0017","DOIUrl":"https://doi.org/10.5376/mpb.2020.11.0017","url":null,"abstract":"Starch glucan chain structure of cassava root is the key factor to determine starch quality. Soluble starch synthase III (SSIII) is the key enzyme to regulate the synthesis of long chain in plant amylopectin glucan. Cassava has two MeSSIII homologous genes MeSSIII-1 and MeSSIII-2. To study the effect of cassava MeSSIII on the quality formation of cassava root starch, a double gene editing vector for MeSSIII-1 and MeSSIII-2 was constructed. The sgRNA target for MeSSIII-1 and MeSSIII-2 was designed simultaneously by online software CRISPR-Pv2.0 based on the conserved segments, and the recombinant pCAMBIAP1301-Cas9MeSSIII-gRNA plasmid was constructed by digestion and ligation. The gene editing vector was transformed into LBA4404 Agrobacterium competent cells and used to infect the friable embryogenic callus of cassava, and the their DNA was extracted. The target segments of MeSSIII-1 and MeSSIII-2 were amplified by PCR for Sanger sequencing, and analyzed the editing of target position. The results showed that the target sites of MeSSIII-1 and MeSSIII-2 were successfully edited. This study helps to further obtain mutants of theMeSSIII gene to analyze the role of this gene in the cassava starch synthesis pathway.","PeriodicalId":32255,"journal":{"name":"Journal of Plant Molecular Breeding","volume":"29 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2020-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"73855189","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2020-01-01DOI: 10.5376/mpb.2020.11.0011
Chao Ma, Y. Bu, Chenghui Zhang, Hong-Xiang Ma, Bao-quan Hu
For the study of the polymorphism and evolutionary characteristics of MYB1 and its allele MYB10 involved in the regulation of anthocyanin biosynthesis in genus Malus. The MYB1 and MYB10 gene sequences amplified by RT-PCR technology in several Malus plants were sequenced and predicted for their protein structures to detect the differences. Six MYB1 sequences were obtained and the full-length CDS were 1 239 bp except for those of Malus cv. Eleyi and Malus sieversii (1 209 bp). While nine MYB10 were sequenced and the full-length CDS were 732 bp (Malus micromalus was 717 bp). The result from the comparison of protein sequences of MYB1 and MYB10 showed that the polymorphism of MYB1 was significantly higher than MYB10 and the polymorphism difference between the alleles may be related to the differentiation of gene function. Two polymorphism sites, the interchange of alkaline arginine and non-polar leucine, located on functional domain which may affect the function of MYB1 in different Malus plants. The result suggested that sequence conservatism of the allele MYB10 and MYB1 significantly differed from each other, and the genetic function responsible for evolution was differentiated. The result can provide further theoretical basis for the study of the gene function and regulation mechanism of apple fruit color.
{"title":"Analysis of Variation in MYB1 and MYB10 in Several Malus Plants","authors":"Chao Ma, Y. Bu, Chenghui Zhang, Hong-Xiang Ma, Bao-quan Hu","doi":"10.5376/mpb.2020.11.0011","DOIUrl":"https://doi.org/10.5376/mpb.2020.11.0011","url":null,"abstract":"For the study of the polymorphism and evolutionary characteristics of MYB1 and its allele MYB10 involved in the regulation of anthocyanin biosynthesis in genus Malus. The MYB1 and MYB10 gene sequences amplified by RT-PCR technology in several Malus plants were sequenced and predicted for their protein structures to detect the differences. Six MYB1 sequences were obtained and the full-length CDS were 1 239 bp except for those of Malus cv. Eleyi and Malus sieversii (1 209 bp). While nine MYB10 were sequenced and the full-length CDS were 732 bp (Malus micromalus was 717 bp). The result from the comparison of protein sequences of MYB1 and MYB10 showed that the polymorphism of MYB1 was significantly higher than MYB10 and the polymorphism difference between the alleles may be related to the differentiation of gene function. Two polymorphism sites, the interchange of alkaline arginine and non-polar leucine, located on functional domain which may affect the function of MYB1 in different Malus plants. The result suggested that sequence conservatism of the allele MYB10 and MYB1 significantly differed from each other, and the genetic function responsible for evolution was differentiated. The result can provide further theoretical basis for the study of the gene function and regulation mechanism of apple fruit color.","PeriodicalId":32255,"journal":{"name":"Journal of Plant Molecular Breeding","volume":"6 3 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2020-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"90418335","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2020-01-01DOI: 10.5376/mpb.2020.11.0014
Yang Shen, Bowei Jia, Jinyu Wang, Xiaoxi Cai, Bingshuang Hu, Yan Wang, Yue‐Ying Chen, Mingzhe Sun, Sun Xiaoli
Soil salt-alkalization is one of the adverse factors limiting crop yields. Identification of key salt-alkaline tolerant genes is of great significance for molecular breeding of stress-resistant crops. In this study, a T-DNA insertion Arabidopsis mutant atgols2 showing higher sensitivity to bicarbonate salt-alkaline stress was screened out against NaHCO3 treatment. Further bioinformatic analysis revealed that the AtGolS2 gene encoded a galactinol synthase, which is a member of the glycosyltransferase family A superfamily. We predicted the protein interaction network of AtGolS2 via SMART online analysis, and found that these AtGolS2 interacting proteins were related to lipid metabolism, galactose biosynthesis and raffinose biosynthesis, and participated in abiotic stress responses. By using the online expression data, we showed that AtGolS2 expression responded to salt, osmotic, drought and ABA stress. PCR amplification by using the three primers method verified the homozygous T-DNA insertion in atgols2. Phenotypic assays further uncovered that atgols2 mutant was more sensitive to high salt, osmotic and ABA stresses than the wild type Arabidopsis. Taken together, results in this study revealed the positive function of AtGolS2 in bicarbonate salt-alkaline, high salt, osmotic and ABA stresses, which will facilitate further research regarding the function and molecular mechanism of the GolS family genes in stress responses.
{"title":"Functional Analysis of Arabidopsis thaliana Galactinol Synthase AtGolS2 in Response to Abiotic Stress","authors":"Yang Shen, Bowei Jia, Jinyu Wang, Xiaoxi Cai, Bingshuang Hu, Yan Wang, Yue‐Ying Chen, Mingzhe Sun, Sun Xiaoli","doi":"10.5376/mpb.2020.11.0014","DOIUrl":"https://doi.org/10.5376/mpb.2020.11.0014","url":null,"abstract":"Soil salt-alkalization is one of the adverse factors limiting crop yields. Identification of key salt-alkaline tolerant genes is of great significance for molecular breeding of stress-resistant crops. In this study, a T-DNA insertion Arabidopsis mutant atgols2 showing higher sensitivity to bicarbonate salt-alkaline stress was screened out against NaHCO3 treatment. Further bioinformatic analysis revealed that the AtGolS2 gene encoded a galactinol synthase, which is a member of the glycosyltransferase family A superfamily. We predicted the protein interaction network of AtGolS2 via SMART online analysis, and found that these AtGolS2 interacting proteins were related to lipid metabolism, galactose biosynthesis and raffinose biosynthesis, and participated in abiotic stress responses. By using the online expression data, we showed that AtGolS2 expression responded to salt, osmotic, drought and ABA stress. PCR amplification by using the three primers method verified the homozygous T-DNA insertion in atgols2. Phenotypic assays further uncovered that atgols2 mutant was more sensitive to high salt, osmotic and ABA stresses than the wild type Arabidopsis. Taken together, results in this study revealed the positive function of AtGolS2 in bicarbonate salt-alkaline, high salt, osmotic and ABA stresses, which will facilitate further research regarding the function and molecular mechanism of the GolS family genes in stress responses.","PeriodicalId":32255,"journal":{"name":"Journal of Plant Molecular Breeding","volume":"41 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2020-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"86198302","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2020-01-01DOI: 10.5376/MPB.2020.11.0027
N. Duan, Yumin Ma, Kun Wang, Xiaomu Wang, Xie Kun, Bai Jing, Yongyi Yang, Yan-yan Pu, Yongchao Gong
In this article, we carried out genome resequencing and SNP mining for cultivated apples in Shandong Province for the sake of the rapid identification of apple varieties, germplasm evaluation, and utilization. Genomic DNA was extracted immediately from leaves of each sample, and Paired-end Illumina genomic libraries were prepared and sequenced on an Illumina Hiseq 4 000 platform following the manufacturer's instructions. Resequencing of the 31 apple genomes generated a total of 363 Gb high-quality cleaned sequences, with an average of 12.5 Gb per accession that represented approximately 15.9x coverage of the apple genome. The data volume fully meets the needs of downstream analysis and SNP mining. When we used the nucleotide mismatch parameter from 1~12, the mapping rate gradually increased to saturation. There was a highly significant correlation (p<0.0001) between the total mapping rate, mapping rate of pair-end data, and mismatch parameter. Univariate fourth-order equation (regression coefficient r>0.99) were predicted. As the mismatch rate increases, the accuracy of mapping decreases; the genome coverage gradually increases, and heterozygous sites' accuracy gradually increases. In this study, two algorithms were used in SNP mining. The intersection was further taken based on the 'chromosome+site information' as the eigenvalues to obtain a highly reliable single nucleotide variant dataset. A total of 374 404 SNP locus were detected. On average, one variation can be identified from 1 896 bp. The accuracy of the Sanger verification test is as high as 98.1%. Annotation analysis shows that among the 373 763 SNPs, 25 047 (6.7%) are located in the gene coding region, 143 269 (38.27%) are located in the intergenic region, and 179 426 (47.92%) are located in the 2 kb region upstream or downstream of the corresponding genes. Among the coding region SNPs, 13 422 are non-synonymous, while 11 625 are synonymous variations. The ratio of non-synonymous to synonymous SNP is 1.15: 1. Using the filtered 4DTV sites, population clustering analysis results constructed using neighbor-joining algorithms are in line with the trend of the classification of cultivated apples in Shandong province.
{"title":"SNP Mining by Genome Resequencing of 30 Apple Varieties in Shandong Province","authors":"N. Duan, Yumin Ma, Kun Wang, Xiaomu Wang, Xie Kun, Bai Jing, Yongyi Yang, Yan-yan Pu, Yongchao Gong","doi":"10.5376/MPB.2020.11.0027","DOIUrl":"https://doi.org/10.5376/MPB.2020.11.0027","url":null,"abstract":"In this article, we carried out genome resequencing and SNP mining for cultivated apples in Shandong Province for the sake of the rapid identification of apple varieties, germplasm evaluation, and utilization. Genomic DNA was extracted immediately from leaves of each sample, and Paired-end Illumina genomic libraries were prepared and sequenced on an Illumina Hiseq 4 000 platform following the manufacturer's instructions. Resequencing of the 31 apple genomes generated a total of 363 Gb high-quality cleaned sequences, with an average of 12.5 Gb per accession that represented approximately 15.9x coverage of the apple genome. The data volume fully meets the needs of downstream analysis and SNP mining. When we used the nucleotide mismatch parameter from 1~12, the mapping rate gradually increased to saturation. There was a highly significant correlation (p<0.0001) between the total mapping rate, mapping rate of pair-end data, and mismatch parameter. Univariate fourth-order equation (regression coefficient r>0.99) were predicted. As the mismatch rate increases, the accuracy of mapping decreases; the genome coverage gradually increases, and heterozygous sites' accuracy gradually increases. In this study, two algorithms were used in SNP mining. The intersection was further taken based on the 'chromosome+site information' as the eigenvalues to obtain a highly reliable single nucleotide variant dataset. A total of 374 404 SNP locus were detected. On average, one variation can be identified from 1 896 bp. The accuracy of the Sanger verification test is as high as 98.1%. Annotation analysis shows that among the 373 763 SNPs, 25 047 (6.7%) are located in the gene coding region, 143 269 (38.27%) are located in the intergenic region, and 179 426 (47.92%) are located in the 2 kb region upstream or downstream of the corresponding genes. Among the coding region SNPs, 13 422 are non-synonymous, while 11 625 are synonymous variations. The ratio of non-synonymous to synonymous SNP is 1.15: 1. Using the filtered 4DTV sites, population clustering analysis results constructed using neighbor-joining algorithms are in line with the trend of the classification of cultivated apples in Shandong province.","PeriodicalId":32255,"journal":{"name":"Journal of Plant Molecular Breeding","volume":"30 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2020-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"88054674","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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