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Article(id=1200500171139305651, tenantId=1146029695717560320, journalId=1189982191388893191, issueId=1200500165426672625, articleNumber=null, orderNo=null, doi=10.16438/j.0513-4870.2023-1259, pmid=null, cstr=null, oa=null, hot=null, price=null, onlineType=0, articleFormat=0, articleType=null, articleTypeStr=research-article, receivedDate=1699200000000, receivedDateStr=2023-11-06, revisedDate=1707148800000, revisedDateStr=2024-02-06, acceptedDate=null, acceptedDateStr=null, onlineDate=1764151145014, onlineDateStr=2025-11-26, pubDate=1718121600000, pubDateStr=2024-06-12, doiRegisterDate=null, doiRegisterDateStr=null, onlineIssueDate=1764151145014, onlineIssueDateStr=2025-11-26, onlineJustAcceptDate=null, onlineJustAcceptDateStr=null, onlineFirstDate=null, onlineFirstDateStr=null, sourceXml=null, magXml=null, createTime=1764151145014, creator=13701087609, updateTime=1764151145014, updator=13701087609, issue=Issue{id=1200500165426672625, tenantId=1146029695717560320, journalId=1189982191388893191, year='2024', volume='59', issue='6', pageStart='1509', pageEnd='1896', issueExtLink='null', onlineDate='null', pubDate='null', beforeIssueId=null, nextIssueId=null, price=null, status=1, issueComplete=1, articleOrder=1, issueType=-1, specialIssue=null, createTime=1764151143651, creator=13701087609, updateTime=1764225143180, updator=13701087609, preIssue=null, nextIssue=null, ext={EN=IssueExt(id=1200810542001680840, tenantId=1146029695717560320, journalId=1189982191388893191, issueId=1200500165426672625, language=EN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=), CN=IssueExt(id=1200810542001680841, tenantId=1146029695717560320, journalId=1189982191388893191, issueId=1200500165426672625, language=CN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=)}, issueFiles=null}, startPage=1864, endPage=1872, ext={EN=ArticleExt(id=1200500172536008961, articleId=1200500171139305651, tenantId=1146029695717560320, journalId=1189982191388893191, language=EN, title=Full-length transcriptome sequencing and bioinformatics analysis of Polygonatum kingianum, columnId=1190335348761793317, journalTitle=Acta Pharmaceutica Sinica, columnName=Original Articles, runingTitle=null, highlight=null, articleAbstract=

The purpose of this study was to enrich the genomic information and provide a basis for further development and utilization of Polygonatum kingianum. The fresh rhizome of P. kingianum was used as the experimental material, and the full-length transcriptome was sequenced by PacBio Sequel platform. The final measured polymerase reads were 1 120 485, with a total of 77.73 GB of data. In NR database, 41 864 homologous sequence alignments were aligned to 5 species; 40 506 were annotated in the KOG database and classified into 26 categories based on their functionality; 69 060 GO annotated 32 functional groups divided into 3 categories: cellular components, molecular functions, and biological processes; 45 779 annotations were added to 145 metabolic pathways in the KEGG database. Among them, there are 127 identified transcripts related to polysaccharide synthesis in the glycolysis/gluconeogenesis metabolic pathway, 144 in the starch and sucrose metabolic pathway, 85 in the amino acid sugar and nucleotide sugar metabolic pathway, and 69 in the fructose and mannose metabolic pathway. In addition, 1 781 transcription factors were detected, distributed in 37 transcription factor families; 180 293 SSR loci were detected, among which single base repeats accounted for the most, accounting for 30.48% of all base repeats, and at least five base repeats, accounting for only 0.14% of single base repeats. The results of this study provide important data supporting for enriching the genomic information of P. kingianum and exploring its effective biosynthetic pathway.

, correspAuthors=Guo-wei ZHENG, Jia CHEN, authorNote=null, correspAuthorsNote=null, copyrightStatement=Copyright ©2024 Acta Pharmaceutica Sinica. All rights reserved., copyrightOwner=null, extLink=null, articleAbsUrl=null, sourceXml=null, magXml=null, pdfUrl=null, pdf=null, pdfFileSize=null, pdfExtLink=null, richHtmlUrl=null, mobilePdfUrl=null, reviewReport=null, pdfFirstPage=null, abstractGraph=null, abstractGraphContent=null, abstractVideo=null, citation=null, cebUrl=null, magXmlContent=null, mapNumber=null, authorCompany=null, fund=null, authors=null, authorsList=Qi MI, Yan-li ZHAO, Ping XU, Meng-wen YU, Xuan ZHANG, Zhen-hua TU, Chun-hua LI, Guo-wei ZHENG, Jia CHEN), CN=ArticleExt(id=1200500176696758751, articleId=1200500171139305651, tenantId=1146029695717560320, journalId=1189982191388893191, language=CN, title=滇黄精全长转录组测序及生物信息学分析, columnId=1190335348896011050, journalTitle=药学学报, columnName=研究论文, runingTitle=null, highlight=null, articleAbstract=

本文通过PacBio Sequel平台对滇黄精根茎进行全长转录组测序, 并对测序结果进行生物信息学分析, 旨在为滇黄精基因功能的研究提供参考.最终测得polymerase reads为1 120 485个, 总数据共77.73 GB.在NR数据库中, 有41 864个同源序列比对到5个物种; 有40 506个在KOG数据库得到注释, 根据其功能分为26类; 有69 060个得到GO注释, 分为细胞组分、分子功能及生物学过程等3大类的32个功能组; 有45 779个注释到KEGG数据库的145条代谢途经中, 其中鉴定与多糖合成有关的转录本在糖酵解/糖异生代谢通路中有127条, 淀粉和蔗糖代谢通路有144条, 氨基酸糖和核苷酸糖代谢通路有85条, 果糖和甘露糖代谢通路有69条; 此外还检测到1 781个转录因子, 分布在37个转录因子家族中; 检测到了180 293个SSR位点, 其中以单碱基重复占比最多, 占所有碱基重复类型的30.48%, 最少的是五碱基重复, 仅为单碱基重复的0.14%.本研究结果为丰富滇黄精基因组信息, 探索其有效成生物合成途径提供重要的数据支持

, correspAuthors=郑国伟, 陈佳, authorNote=null, correspAuthorsNote=
*郑国伟, E-mail: ;
陈佳, E-mail:
, copyrightStatement=版权所有©《药学学报》编辑部2024, copyrightOwner=null, extLink=null, articleAbsUrl=null, sourceXml=KYf1zMRl/kdCe0cPe8R+8g==, magXml=AO9xIoQK0Fp4BCua26xfCw==, pdfUrl=null, pdf=P0WTD/grNQdZDrdTsb72mA==, pdfFileSize=4608439, pdfExtLink=null, richHtmlUrl=null, mobilePdfUrl=null, reviewReport=null, pdfFirstPage=null, abstractGraph=aJONuaJV7M2DRf+ko3K/zw==, abstractGraphContent=null, abstractVideo=null, citation=null, cebUrl=null, magXmlContent=Wk/GF0yGPlpeTz9rGP1bUQ==, mapNumber=null, authorCompany=null, fund=null, authors=

#共同第一作者.

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Acta Pharm Sin (药学学报), 2022, 57: 1193-1202., articleTitle=null, refAbstract=null)], funds=[Fund(id=1201118447024431441, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1200500171139305651, awardId=82360751, language=CN, fundingSource=国家自然科学基金资助项目(82360751), fundOrder=null, country=null), Fund(id=1201118447146066262, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1200500171139305651, awardId=2019YZG07, language=CN, fundingSource=云南中医药大学高层次人才计划(2019YZG07), fundOrder=null, country=null), Fund(id=1201118447322227030, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1200500171139305651, awardId=YNWR-QNBJ-2020-289, language=CN, fundingSource=云南青年精英人才项目(YNWR-QNBJ-2020-289), fundOrder=null, country=null)], companyList=[AuthorCompany(id=1201118434911282019, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1200500171139305651, xref=null, ext=[AuthorCompanyExt(id=1201118434923864933, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1200500171139305651, companyId=1201118434911282019, language=EN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=1. College of Chinese Material Medica, Yunnan University of Chinese Medicine, Kunming 650500, China), AuthorCompanyExt(id=1201118434928059239, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1200500171139305651, companyId=1201118434911282019, language=CN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=1.云南中医药大学中药学院, 云南 昆明 650500)]), AuthorCompany(id=1201118435045499758, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1200500171139305651, xref=null, ext=[AuthorCompanyExt(id=1201118435066471281, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1200500171139305651, companyId=1201118435045499758, language=EN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=2. College of Ethnic Medicines, Yunnan University of Chinese Medicine, Kunming 650500, China), AuthorCompanyExt(id=1201118435074859890, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1200500171139305651, companyId=1201118435045499758, language=CN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=2.云南中医药大学民族医药学院, 云南 昆明 650500)]), AuthorCompany(id=1201118435217466236, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1200500171139305651, xref=null, ext=[AuthorCompanyExt(id=1201118435225854846, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1200500171139305651, companyId=1201118435217466236, language=EN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=3. Gongshan County Agriculture Village Ju Agriculture Comprehensive Development Service Center, Nujiang of the Lisu Autonomous Prefecture, Nujiang 673500, China), AuthorCompanyExt(id=1201118435230049152, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1200500171139305651, companyId=1201118435217466236, language=CN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=3.贡山县农业农村局农业综合发展服务中心, 云南 怒江傈僳族自治州 673500)])], figs=[ArticleFig(id=1201118443299889376, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1200500171139305651, language=EN, label=null, caption=null, figureFileSmall=LLV5yqFmPh3l3YJP9zG6Aw==, figureFileBig=GqhfNV6nm0Qqn1tiivzprg==, tableContent=null), ArticleFig(id=1201118443434107109, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1200500171139305651, language=CN, label=Figure 1, caption= 　Functional annotation of full-length transcripts of <i>P. kingianum</i>. A: Summary of functional annotation results for <i>P. kingianum</i> transcripts; B: The Venn diagram between NR, COG, KEGG, Swissprot and GO; C: Species distribution of annotated transcripts in NR database; D: Classification diagram of GO annotation. BP: Biological process; CC: Cellular component; MF: Molecular function , figureFileSmall=LLV5yqFmPh3l3YJP9zG6Aw==, figureFileBig=GqhfNV6nm0Qqn1tiivzprg==, tableContent=null), ArticleFig(id=1201118443618656492, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1200500171139305651, language=EN, label=null, caption=null, figureFileSmall=VVJSoROAdOJ8JPs+/E+SxA==, figureFileBig=o2mia0AtJQ8q5x0Hnmt++g==, tableContent=null), ArticleFig(id=1201118443710931187, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1200500171139305651, language=CN, label=Figure 2, caption= 　Classification diagram of KOG annotation , figureFileSmall=VVJSoROAdOJ8JPs+/E+SxA==, figureFileBig=o2mia0AtJQ8q5x0Hnmt++g==, tableContent=null), ArticleFig(id=1201118443828371705, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1200500171139305651, language=EN, label=null, caption=null, figureFileSmall=50nGlQsM9uhVKdv6rJYpwA==, figureFileBig=P6Hoj2gYNnfFv+5mE0NJiw==, tableContent=null), ArticleFig(id=1201118443937423613, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1200500171139305651, language=CN, label=Figure 3, caption= 　Schematic diagram of KEGG function distribution , figureFileSmall=50nGlQsM9uhVKdv6rJYpwA==, figureFileBig=P6Hoj2gYNnfFv+5mE0NJiw==, tableContent=null), ArticleFig(id=1201118444038086913, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1200500171139305651, language=EN, label=null, caption=null, figureFileSmall=OvpGd+6yTP81UoN0WqdCXQ==, figureFileBig=ZzJt/aeUoq+qhaWvPzVJjA==, tableContent=null), ArticleFig(id=1201118444168110341, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1200500171139305651, language=CN, label=Figure 4, caption= 　Prediction and analysis of Lnc-RNA transcript of <i>P. kingianum</i>. A: Venn plot of Lnc-RNA prediction results of <i>P. kingianum</i>; B: Distribution map of Lnc-RNA and mRNA length. CPC2: Contrastive predictive coding; CNCI: Coding-non-coding index; PLEK: Predictor of long non-coding RNAs and messenger RNAs based on an improved k-mer scheme; Lnc-RNA: Long non-coding RNA; mRNA: Messenger RNA , figureFileSmall=OvpGd+6yTP81UoN0WqdCXQ==, figureFileBig=ZzJt/aeUoq+qhaWvPzVJjA==, tableContent=null), ArticleFig(id=1201118444310716681, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1200500171139305651, language=EN, label=null, caption=null, figureFileSmall=KvlRQ+yJZPVzZ4xI/isZ8w==, figureFileBig=+SFYFMAiZqx9z6BmGyLfyQ==, tableContent=null), ArticleFig(id=1201118444461711630, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1200500171139305651, language=CN, label=Figure 5, caption= 　Classification and statistics of transcription factor annotations in <i>P. kingianum.</i> TFs: Transcription factors , figureFileSmall=KvlRQ+yJZPVzZ4xI/isZ8w==, figureFileBig=+SFYFMAiZqx9z6BmGyLfyQ==, tableContent=null), ArticleFig(id=1201118444600123668, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1200500171139305651, language=EN, label=null, caption=null, figureFileSmall=KwlYbIjyNuiKJYx+OQJUtg==, figureFileBig=SP5qWgBtZgDBkSKEZVHKXg==, tableContent=null), ArticleFig(id=1201118444709175578, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1200500171139305651, language=CN, label=Figure 6, caption= 　Correlation analyses between <i>INV</i> transcripts and polysaccharide content. <i>INV</i>: Beta-fructofuranosidase , figureFileSmall=KwlYbIjyNuiKJYx+OQJUtg==, figureFileBig=SP5qWgBtZgDBkSKEZVHKXg==, tableContent=null), ArticleFig(id=1201118444872753441, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1200500171139305651, language=EN, label=null, caption=null, figureFileSmall=null, figureFileBig=null, tableContent=
SampleFLNC bases/bpFLNC numberMin numberAverage length /bpMax length /bpN50 /bp
Polygonatum_kingianum1 953 756 490678 465722 88014 7583 212
), ArticleFig(id=1201118444960833827, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1200500171139305651, language=CN, label=Table 1, caption=

　FLNC result statistics. FLNC: Full-length non-concatemer

, figureFileSmall=null, figureFileBig=null, tableContent=
SampleFLNC bases/bpFLNC numberMin numberAverage length /bpMax length /bpN50 /bp
Polygonatum_kingianum1 953 756 490678 465722 88014 7583 212
), ArticleFig(id=1201118445111828778, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1200500171139305651, language=EN, label=null, caption=null, figureFileSmall=null, figureFileBig=null, tableContent=
SSR typecc*p1p2p3p4p5p6
Number1 70710 18010 2754 8155 952259148373
Percent/%5.0630.230.4814.2817.660.770.441.11
), ArticleFig(id=1201118445225074992, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1200500171139305651, language=CN, label=Table 2, caption=

　SSR site analysis of P. kingianum. SSR: Simple sequence repeats; c: Composite SSR; c*: Composite type with overlap between SSRs; p1: Single base repeat; p2: Dibasic repeat; p3: Triple base repeat; p4: Tetra nucleotide duplication; p5: Pent nucleotide duplication; p6: Six base repeat

, figureFileSmall=null, figureFileBig=null, tableContent=
SSR typecc*p1p2p3p4p5p6
Number1 70710 18010 2754 8155 952259148373
Percent/%5.0630.230.4814.2817.660.770.441.11
), ArticleFig(id=1201118445329932596, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1200500171139305651, language=EN, label=null, caption=null, figureFileSmall=null, figureFileBig=null, tableContent=
IDPrimer sequencePrime
TM/℃		Prime GC
percent/%		Primer pair
product
size/bp		SSR
type		SSRSSR
length
/bp		SSR
start		SSR
end
Polygonatum_kingianum_transcript_61128F: CCAGGAGAATGGGTTGGTCC60.03460280p1(A)10102231
R: AGAGAGGAACACAACAGCGG59.96655
Polygonatum_kingianum_transcript_33808F: ATGTTCCTGCAGAGCGTCAA59.96550103c*(CT)12(CTCT)6*24149172
R: CATCCACTCAGGGCATTGGT60.03455
Polygonatum_kingianum_transcript_52353F: GGCGGGGAAGGGTTAAGAAA59.96155200p1(T)10101 9521 961
R: CAACGGAGTGCCTGGATTCT60.03655
Polygonatum_kingianum_transcript_47350F: GAAACAGGGCCGTGACACTA59.96655197p3(AGC)5151 8111 825
R: CTGCAAGGAGAGATGCACCA60.03655
Polygonatum_kingianum_transcript_21981F: AGGCTGCAAATTCCTCCTCC60.03455239p1(T)10101625
R: TTTTGGAGGCGGCAGGTAAT59.96150
Polygonatum_kingianum_transcript_59587F: GAGTACCCTTTTGCCGAGCT60.03655279c*(T)13(TT)6*121 3321 343
R: CTTCTGATCCGCTGCGATCT59.96855
Polygonatum_kingianum_transcript_7136F: AACACCATATCCTGCGTGCA60.03650256c(T)10atctttttcc(GA)63293124
R: GCATTGCAGGCCTCACAAAA59.96650
Polygonatum_kingianum_transcript_2968F: CTTCCCTCTGTCTGCAGCAA59.96455227p1(T)10104 3384 347
R: TAAGAGCAGGTCTCCTCCCC60.03260
Polygonatum_kingianum_transcript_43887F: CTGAGTTCCTTGCTGAGGG60.03660243p2(CT)6121728
R: GAGAGCGGATATGGCGTTGA59.96855
Polygonatum_kingianum_transcript_58976F: CCTCTACGTCAGGACCCAGA60.03460143p1(T)11116373
R: TGTTCAGCGCCTTCAGCATA60.03650
Polygonatum_kingianum_transcript_55221F: GATTCTGAGGCAGCCCACTT60.03555170p3(GAA)824306329
R: CCCGGGAAAGATTGTCCACA59.96255
Polygonatum_kingianum_transcript_9971F: CTGGATGGATTGGTGGGCTT60.03355203p1(T)1010590599
R: ACGAATTGTCTCCGGCTTGT59.96550
Polygonatum_kingianum_transcript_57199F: CTCAAGGGCACTGTGACGAT60.03755108p3(TTC)5151 4981 512
R: AACTCAGATGTCTGCAGGGC60.03655
Polygonatum_kingianum_transcript_9908F: GAGCTACCATCACCTCGCTC59.96960127p2(CA)612978989
R: CGACCAACAATGCAGCCAAA59.96850
Polygonatum_kingianum_transcript_28150F: TTTTTCCCCCTCTCATCGGC60.03455191p2(CT)6122031
R: AGAATTCGCCTTGCCATCCA60.03450
Polygonatum_kingianum_transcript_60339F: CGTGGACGAGGATGTGGAAA60.03855151c*(GT)9(GAGA)5
(GA)11*		401453
R: ATTATGCCCCATGCCCACAA60.03150
Polygonatum_kingianum_transcript_56385F: AGTGGAGGTGAAGACCCTGT60.10555275p1(T)10101019
R: TGCCTTCATCCAATGCCCAT60.03250
Polygonatum_kingianum_transcript_15896F: GTCTGTGCTGCAACACTGTG59.97155117p3(GAG)515458472
R: GCTCGTGAAGTGACACTCCA59.96855
Polygonatum_kingianum_transcript_56614F: CCATCGAGAACCCCTCATCG59.96760146c*(TCTC)7(TCTCTC)
5*(TC)15*		30101130
R: GAAGGGAGCCTTGTCGTTGA59.96555
Polygonatum_kingianum_transcript_63298F: TGTGATCGCTCTGTGCATGT60.03750300p1(A)11111 0811 091
R: ACAAGGACGACGAAGGCAAT59.96550
Polygonatum_kingianum_transcript_44544F: CTGCTGCTGCGATGGAATTC59.97155194c(A)10(GCT)91063108
R: ATCATCAACACCTGGCTCCG60.10855
), ArticleFig(id=1201118445485121850, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1200500171139305651, language=CN, label=Table 3, caption=

　SSR primer design table

, figureFileSmall=null, figureFileBig=null, tableContent=
IDPrimer sequencePrime
TM/℃		Prime GC
percent/%		Primer pair
product
size/bp		SSR
type		SSRSSR
length
/bp		SSR
start		SSR
end
Polygonatum_kingianum_transcript_61128F: CCAGGAGAATGGGTTGGTCC60.03460280p1(A)10102231
R: AGAGAGGAACACAACAGCGG59.96655
Polygonatum_kingianum_transcript_33808F: ATGTTCCTGCAGAGCGTCAA59.96550103c*(CT)12(CTCT)6*24149172
R: CATCCACTCAGGGCATTGGT60.03455
Polygonatum_kingianum_transcript_52353F: GGCGGGGAAGGGTTAAGAAA59.96155200p1(T)10101 9521 961
R: CAACGGAGTGCCTGGATTCT60.03655
Polygonatum_kingianum_transcript_47350F: GAAACAGGGCCGTGACACTA59.96655197p3(AGC)5151 8111 825
R: CTGCAAGGAGAGATGCACCA60.03655
Polygonatum_kingianum_transcript_21981F: AGGCTGCAAATTCCTCCTCC60.03455239p1(T)10101625
R: TTTTGGAGGCGGCAGGTAAT59.96150
Polygonatum_kingianum_transcript_59587F: GAGTACCCTTTTGCCGAGCT60.03655279c*(T)13(TT)6*121 3321 343
R: CTTCTGATCCGCTGCGATCT59.96855
Polygonatum_kingianum_transcript_7136F: AACACCATATCCTGCGTGCA60.03650256c(T)10atctttttcc(GA)63293124
R: GCATTGCAGGCCTCACAAAA59.96650
Polygonatum_kingianum_transcript_2968F: CTTCCCTCTGTCTGCAGCAA59.96455227p1(T)10104 3384 347
R: TAAGAGCAGGTCTCCTCCCC60.03260
Polygonatum_kingianum_transcript_43887F: CTGAGTTCCTTGCTGAGGG60.03660243p2(CT)6121728
R: GAGAGCGGATATGGCGTTGA59.96855
Polygonatum_kingianum_transcript_58976F: CCTCTACGTCAGGACCCAGA60.03460143p1(T)11116373
R: TGTTCAGCGCCTTCAGCATA60.03650
Polygonatum_kingianum_transcript_55221F: GATTCTGAGGCAGCCCACTT60.03555170p3(GAA)824306329
R: CCCGGGAAAGATTGTCCACA59.96255
Polygonatum_kingianum_transcript_9971F: CTGGATGGATTGGTGGGCTT60.03355203p1(T)1010590599
R: ACGAATTGTCTCCGGCTTGT59.96550
Polygonatum_kingianum_transcript_57199F: CTCAAGGGCACTGTGACGAT60.03755108p3(TTC)5151 4981 512
R: AACTCAGATGTCTGCAGGGC60.03655
Polygonatum_kingianum_transcript_9908F: GAGCTACCATCACCTCGCTC59.96960127p2(CA)612978989
R: CGACCAACAATGCAGCCAAA59.96850
Polygonatum_kingianum_transcript_28150F: TTTTTCCCCCTCTCATCGGC60.03455191p2(CT)6122031
R: AGAATTCGCCTTGCCATCCA60.03450
Polygonatum_kingianum_transcript_60339F: CGTGGACGAGGATGTGGAAA60.03855151c*(GT)9(GAGA)5
(GA)11*		401453
R: ATTATGCCCCATGCCCACAA60.03150
Polygonatum_kingianum_transcript_56385F: AGTGGAGGTGAAGACCCTGT60.10555275p1(T)10101019
R: TGCCTTCATCCAATGCCCAT60.03250
Polygonatum_kingianum_transcript_15896F: GTCTGTGCTGCAACACTGTG59.97155117p3(GAG)515458472
R: GCTCGTGAAGTGACACTCCA59.96855
Polygonatum_kingianum_transcript_56614F: CCATCGAGAACCCCTCATCG59.96760146c*(TCTC)7(TCTCTC)
5*(TC)15*		30101130
R: GAAGGGAGCCTTGTCGTTGA59.96555
Polygonatum_kingianum_transcript_63298F: TGTGATCGCTCTGTGCATGT60.03750300p1(A)11111 0811 091
R: ACAAGGACGACGAAGGCAAT59.96550
Polygonatum_kingianum_transcript_44544F: CTGCTGCTGCGATGGAATTC59.97155194c(A)10(GCT)91063108
R: ATCATCAACACCTGGCTCCG60.10855
), ArticleFig(id=1201118445610950974, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1200500171139305651, language=EN, label=null, caption=null, figureFileSmall=null, figureFileBig=null, tableContent=
GeneNo. of transcriptEnzymeKEGG pathwayEnzyme code
PGM22Phosphoglucomutaseko00010, ko005205.4.2.2
ALDO10Fructose-bisphosphate aldolaseko000104.1.2.13
GPI39Glucose-6-phosphate isomeraseko000105.3.1.9
HK19Hexokinaseko00010, ko000512.7.1.1
scrK36Fructokinaseko00010, ko000512.7.1.90
FBP1Fructose-1, 6-bisphosphataseko000103.1.3.11
MPI3Mannose-6-phosphate isomeraseko000515.3.1.8
PMM1Phosphomannomutaseko000515.4.2.8
GMPP5Mannose-1-phosphate guanylyltransferaseko000512.7.7.13
GMDs5Gdpmannose 4, 6-dehydrataseko000514.2.1.47
RHM12UDP-glucose 4, 6-dehydrataseko005204.2.1.76
UXE4UDP-arabinose 4-epimeraseko005205.1.3.5
AXS13UDP-apiose/xylose synthaseko00520-
UGE10UDP-glucuronate 4-epimeraseko005205.1.3.6
GALE10UDP-glucose 4-epimeraseko005205.1.3.2
UGDH14UDP-glucose 6-dehydrogenaseko005201.1.1.22
SPS31Sucrose-phosphate synthaseko005002.4.1.14
SUS62Sucrose synthaseko005002.4.1.13
SPP3Sucrose-6-phosphataseko005003.1.3.24
INV48Beta-fructofuranosidaseko005003.2.1.26
), ArticleFig(id=1201118445740974404, tenantId=1146029695717560320, journalId=1189982191388893191, articleId=1200500171139305651, language=CN, label=Table 4, caption=

　Unigene related to the synthesis of polysaccharides from P. kingianum

, figureFileSmall=null, figureFileBig=null, tableContent=
GeneNo. of transcriptEnzymeKEGG pathwayEnzyme code
PGM22Phosphoglucomutaseko00010, ko005205.4.2.2
ALDO10Fructose-bisphosphate aldolaseko000104.1.2.13
GPI39Glucose-6-phosphate isomeraseko000105.3.1.9
HK19Hexokinaseko00010, ko000512.7.1.1
scrK36Fructokinaseko00010, ko000512.7.1.90
FBP1Fructose-1, 6-bisphosphataseko000103.1.3.11
MPI3Mannose-6-phosphate isomeraseko000515.3.1.8
PMM1Phosphomannomutaseko000515.4.2.8
GMPP5Mannose-1-phosphate guanylyltransferaseko000512.7.7.13
GMDs5Gdpmannose 4, 6-dehydrataseko000514.2.1.47
RHM12UDP-glucose 4, 6-dehydrataseko005204.2.1.76
UXE4UDP-arabinose 4-epimeraseko005205.1.3.5
AXS13UDP-apiose/xylose synthaseko00520-
UGE10UDP-glucuronate 4-epimeraseko005205.1.3.6
GALE10UDP-glucose 4-epimeraseko005205.1.3.2
UGDH14UDP-glucose 6-dehydrogenaseko005201.1.1.22
SPS31Sucrose-phosphate synthaseko005002.4.1.14
SUS62Sucrose synthaseko005002.4.1.13
SPP3Sucrose-6-phosphataseko005003.1.3.24
INV48Beta-fructofuranosidaseko005003.2.1.26
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滇黄精全长转录组测序及生物信息学分析
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米琪 1, # , 赵艳丽 1, # , 徐萍 1 , 于梦雯 1 , 张萱 1 , 涂振华 1 , 李春华 3 , 郑国伟 1, * , 陈佳 2, *
药学学报 | 研究论文 2024,59(6): 1864-1872
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药学学报 | 研究论文 2024, 59(6): 1864-1872
滇黄精全长转录组测序及生物信息学分析
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米琪1, #, 赵艳丽1, #, 徐萍1, 于梦雯1, 张萱1, 涂振华1, 李春华3, 郑国伟1, * , 陈佳2, *
作者信息
  • 1.云南中医药大学中药学院, 云南 昆明 650500
  • 2.云南中医药大学民族医药学院, 云南 昆明 650500
  • 3.贡山县农业农村局农业综合发展服务中心, 云南 怒江傈僳族自治州 673500

通讯作者:

*郑国伟, E-mail: ;
陈佳, E-mail:
Full-length transcriptome sequencing and bioinformatics analysis of Polygonatum kingianum
Qi MI1, Yan-li ZHAO1, Ping XU1, Meng-wen YU1, Xuan ZHANG1, Zhen-hua TU1, Chun-hua LI3, Guo-wei ZHENG1, * , Jia CHEN2, *
Affiliations
  • 1. College of Chinese Material Medica, Yunnan University of Chinese Medicine, Kunming 650500, China
  • 2. College of Ethnic Medicines, Yunnan University of Chinese Medicine, Kunming 650500, China
  • 3. Gongshan County Agriculture Village Ju Agriculture Comprehensive Development Service Center, Nujiang of the Lisu Autonomous Prefecture, Nujiang 673500, China
出版时间: 2024-06-12 doi: 10.16438/j.0513-4870.2023-1259
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摘要
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本文通过PacBio Sequel平台对滇黄精根茎进行全长转录组测序, 并对测序结果进行生物信息学分析, 旨在为滇黄精基因功能的研究提供参考.最终测得polymerase reads为1 120 485个, 总数据共77.73 GB.在NR数据库中, 有41 864个同源序列比对到5个物种; 有40 506个在KOG数据库得到注释, 根据其功能分为26类; 有69 060个得到GO注释, 分为细胞组分、分子功能及生物学过程等3大类的32个功能组; 有45 779个注释到KEGG数据库的145条代谢途经中, 其中鉴定与多糖合成有关的转录本在糖酵解/糖异生代谢通路中有127条, 淀粉和蔗糖代谢通路有144条, 氨基酸糖和核苷酸糖代谢通路有85条, 果糖和甘露糖代谢通路有69条; 此外还检测到1 781个转录因子, 分布在37个转录因子家族中; 检测到了180 293个SSR位点, 其中以单碱基重复占比最多, 占所有碱基重复类型的30.48%, 最少的是五碱基重复, 仅为单碱基重复的0.14%.本研究结果为丰富滇黄精基因组信息, 探索其有效成生物合成途径提供重要的数据支持

滇黄精  /  全长转录组  /  种质资源  /  多糖合成

The purpose of this study was to enrich the genomic information and provide a basis for further development and utilization of Polygonatum kingianum. The fresh rhizome of P. kingianum was used as the experimental material, and the full-length transcriptome was sequenced by PacBio Sequel platform. The final measured polymerase reads were 1 120 485, with a total of 77.73 GB of data. In NR database, 41 864 homologous sequence alignments were aligned to 5 species; 40 506 were annotated in the KOG database and classified into 26 categories based on their functionality; 69 060 GO annotated 32 functional groups divided into 3 categories: cellular components, molecular functions, and biological processes; 45 779 annotations were added to 145 metabolic pathways in the KEGG database. Among them, there are 127 identified transcripts related to polysaccharide synthesis in the glycolysis/gluconeogenesis metabolic pathway, 144 in the starch and sucrose metabolic pathway, 85 in the amino acid sugar and nucleotide sugar metabolic pathway, and 69 in the fructose and mannose metabolic pathway. In addition, 1 781 transcription factors were detected, distributed in 37 transcription factor families; 180 293 SSR loci were detected, among which single base repeats accounted for the most, accounting for 30.48% of all base repeats, and at least five base repeats, accounting for only 0.14% of single base repeats. The results of this study provide important data supporting for enriching the genomic information of P. kingianum and exploring its effective biosynthetic pathway.

Polygonatum kingianum  /  full-length transcriptome  /  germplasm resource  /  polysaccharide synthesis
米琪, 赵艳丽, 徐萍, 于梦雯, 张萱, 涂振华, 李春华, 郑国伟, 陈佳. 滇黄精全长转录组测序及生物信息学分析. 药学学报, 2024 , 59 (6) : 1864 -1872 . DOI: 10.16438/j.0513-4870.2023-1259
Qi MI, Yan-li ZHAO, Ping XU, Meng-wen YU, Xuan ZHANG, Zhen-hua TU, Chun-hua LI, Guo-wei ZHENG, Jia CHEN. Full-length transcriptome sequencing and bioinformatics analysis of Polygonatum kingianum[J]. Acta Pharmaceutica Sinica, 2024 , 59 (6) : 1864 -1872 . DOI: 10.16438/j.0513-4870.2023-1259
正文
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滇黄精(Polygonatum kingianum Coll. et Hemsl) 为百合科黄精属植物, 与黄精(P. sibiricum) 和多花黄精(P. cyrtonema) 共同收录于《中华人民共和国药典》 (2020版) 中[1], 是云南重要的道地药材之一, 具有补气养阴、健脾润肺等功效, 用于治疗脾胃气虚、体倦乏力等症。主要分布在云南、四川、贵州等西南部省份, 海拔1 400~2 100 m的亚热带地区[2]。滇黄精根茎中含有大量的糖分、淀粉、蛋白质以及维生素等多种营养成分, 具有保健功能, 久服可以强筋健骨、坚实肌肉, 对人体机能恢复十分有益[3]。作为药食同源植物, 早在2002年就被卫生部列入《既是食品又是药品的物品名单》, 其保健功效得到国家权威部门认可[4]。
滇黄精根茎中富含多糖、甾体皂苷、生物碱、总酚等活性成分以及可溶性糖、蛋白质、还原糖、黄酮等营养成分[5]。其中黄精多糖作为中国药典规定的唯一含量测定成分, 具有广泛的生物学功能, 如调节血糖血脂、调节免疫、抗氧化、抗肿瘤、调节免疫等功能[6]。多糖的生物合成是一个十分复杂的过程, 滇黄精多糖主要由甘露糖、半乳糖、葡萄糖、阿拉伯糖、木糖、氨基半乳糖、岩藻糖、氨基葡萄糖、鼠李糖、葡萄糖醛酸以及半乳糖醛酸11种单糖组成[7]。已有研究报道了20余种可能参与黄精多糖生物合成的关键酶基因[8], 推动了滇黄精活性成分生物合成的研究。但是目前对于黄精多糖生物合成的研究依然处于前期探索阶段, 对于滇黄精的遗传信息以及多糖、皂苷、营养成分等合成通路研究报道较少。
随着转录组测序技术的发展, 二代转录组测序技术广泛应用于植物研究, 在植物研究中发挥着重要作用。Wang等[9]通过HiSeq 2500转录组测序, 确定β-呋喃果糖苷酶(INV)、己糖激酶(HK) 和甘露糖-1-磷酸鸟苷转移酶(GMPP) 是滇黄精多糖代谢途径中酶的关键基因。Wang等[10]利用Illumina Hiseq测序平台对黄精全株进行转录组测序, 发现UDP-葡萄糖基转移酶、β-葡萄糖苷酶等参与了黄精皂苷元生物合成途径。但是二代测序存在读长短, 在建库的时候需要将转录本打成碎片; 在转录本异构体的组装过程中存在较多的嵌合体错误, 丢失可变剪切等重要信息等缺点, 大大制约了转录组数据的深度利用。三代测序的出现弥补了这一缺点, 相比于二代测序技术边合成边测序[11], 三代测序克服了二代测序在鉴别重复元素等方面的局限性, 有效减少时间, 同时原理上避免了PCR的扩增错误, 纠正GC偏好性等缺点, 具有更高的通量和测序效率[12, 13]。本研究采用PacBio测序平台对滇黄精进行全长转录组测序, 利用生物信息学软件进行组装和注释, 以期丰富滇黄精转录组数据, 为滇黄精活性成分的生物合成途径奠定基础。
材料与方法
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材料  供试材料为百合科黄精属滇黄精(P. kingianum), 采自云南省红河州石屏县龙武镇黄精种植基地(23.93 N, 102.41 E, 海拔2 174.50 m), 快速清洗泥土后取当年新生部分根茎, 45 ℃烘干后测得其多糖含量为199.80 mg·g-1, 其余部分液氮速冻, -80 ℃冰箱保存, 用于后续提取RNA。
RNA提取与检测  利用琼脂糖凝胶电泳检测样本基因组是否污染; Nanodrop检测所提RNA的纯度; 使用Nano Drop和Agilent 2100生物分析仪(Thermo Fisher Scientific, USA) 检测RNA的完整性; Qubit对RNA浓度进行精确定量。
转录组测序文库构建 Oligo (dT) 富集含有polyA的mRNA, 使用SMARTer PCR cDNA试剂盒将mRNA反转录为cDNA; PCR扩增富集合成的cDNA, 通过循环优化确定PCR的最佳条件; 利用磁珠筛选片段进行大规模PCR, 以获得足够的cDNA总量; 全长cDNA进行损伤修复、末端修复、链接SMRT哑铃型接头, 构建全长转录组文库; 核酸外切酶消化, 去除cDNA两端未连接接头的序列; 最后结合引物、绑定DNA聚合酶, 形成完整的SMRT ball文库。库检合格后, 使用PacBio SM-RT Iso-seq平台进行全长转录组测序。测序工作委托武汉迈维生物科技有限公司。
生物信息分析流程  使用PacBio SMRTlink对下机后的数据进行处理, 首先对单分子测序序列polymerase reads进行拆分得到Subreads, 基于同一polymerase reads得到的Subreads经过自我纠错形成循环共识序列(circular consensus sequence, CCS), 再根据序列是否包含5′、3′端引物以及polyA尾将序列分为全长序列(full-length non-concatemer, FLNC) 与非全长序列, 最后通过FLNC聚类, 得到一致性转录本系列(consensus transcripts)。
将去除冗余后的转录本比对到非冗余蛋白序列数据库(NR, http://ftp.ncbi.nlm.nih.gov/blast/db)、SwissProt蛋白序列数据库(SwissProt, http://ftp.ebi.ac.uk/pub/databases/swissprot)、基因本体数据库(GO, http://geneontology.org)、京都基因与基因组百科全书(KEGG, http://www.genome.jp/kegg) 等数据库, 得到转录本的注释结果; 对去除冗余后的转录本进行简单重复序列(simple sequence repeats, SSR) 分析和编码潜能预测得到长链非编码RNA (long non-coding RNA, Lnc-RNA)。
多糖合成相关基因挖掘  基于文献报道的多糖合成通路, 根据转录本在KEGG中糖酵解/糖异生(ko00010)、淀粉和蔗糖代谢(ko00500)、氨基酸糖和核苷酸糖代谢(ko00520)、果糖和甘露糖代谢(ko00051) 的注释信息, 结合滇黄精多糖含量, 通过多糖含量与基因表达量的相关性分析, 初步挖掘与滇黄精多糖合成相关的基因和转录本。
数据分析  使用Diamond[14] Blastx软件对转录本进行KEGG、NR、Swiss-Prot、GO、COG、KOG功能注释, 使用CNCI[15]、CPC2[16]、PLEK[17]等编码潜能预测软件进行编码潜能预测, 使用iTAK[18]软件进行转录因子注释, 使用MISA[19]进行SSR位点分析; 使用Origin 2019和迈维云平台进行绘图及Pearson相关性分析(P value阈值: 0.05, cor. value阈值: 0.80)。
结果与分析
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1 全长转录本数据统计
通过提取滇黄精根茎的总RNA, 在PacBio平台上建立了1个PacBio ISO-Sep文库。共获得77.73 GB原始数据(1 120 485 reads), 平均长度为69 375 bp。在SMRT cell的测序中得到CCS bases: 2 295 228 275 bp, 平均长度为2 932 bp, 最长序列为14 832 bp。
插入片段识别处理后, 序列可以分成三类: 全长序列、全长非嵌合序列(FLNC) 和具有polyA的全长非嵌合序列。FLNC用于后续分析, 具体数据如表 1所示。
利用CD-HIT软件进行聚类去冗余, total bases: 159 858 002 bp; total number: 55 454; 最短序列: 101 bp; 平均长度: 2 883 bp; 最长序列: 10 346 bp; N50: 3 201 bp。
2 基因功能注释和功能分类
使用Diamond Blastx软件将滇黄精三代转录本去冗余后得到的转录本序列与KEGG、NR、Swiss-Prot、GO以及KOG数据库比对, 结果显示共有52 690 (95.00%) 个基因得到注释(图 1A)。滇黄精转录组在与NR、GO、KOG、KEGG和SwissProt 5个数据库比对后, 均注释到的基因有29 975个, 单独注释到的基因分别有2 069、4、5、46和0个(图 1B)。滇黄精转录组在NR数据库中注释得到52 346个基因, 占比94.40%, 其中与石刁柏(Asparagus offoconalis L.) 的基因配比率最高, 有36 733个基因, 占比为70.17%, 其他基因配比率高的物种分别为油棕(Elaeis guineensis Jacq.)、海枣(Phoenix dactylifera L.)、椰子(Cocos nucifera L.) 和大车前(Planago major L.), 占比分别为3.77%、3.72%、1.54%和0.77% (图 1C)。在GO数据库中根据基因功能分为生物学过程、细胞成分和分子功能三大类, 其中生物学过程比对到161 262个基因, 细胞成分比对到49 755个基因, 分子功能比对到69 060个基因(图 1D)。
将滇黄精转录本比对KOG数据库的25个功能组进行分类统计, 其中滇黄精转录本比对到最多的功能为“一般功能预测”, 数目达到7 841个, 其次为“翻译后修饰、蛋白质转运、伴侣蛋白”, 数目达4 335, 最少的为“细胞活性”, 仅为21 (图 2)。
在KEGG数据库中, 将转录本参与的KEGG代谢通路分为5个分支: 细胞过程、环境信息处理、遗传信息处理、代谢以及有机系统, 测得的基因数分别为1 837、2 880、10 352、29 084和1 626个(图 3)。
3 Lnc-RNA转录本预测分析
Lnc-RNA为一类长度大于200 bp的长链非编码RNA。在获得可靠的滇黄精转录本后, 将转录本的编码能力进行预测, 对7大数据库都没有注释到的转录本, 根据Lnc-RNA的不编码蛋白的功能特点, 通过CNCI、CPC2、PLEK软件对滇黄精非全长序列进行Lnc-RNA预测, 取3种分析软件鉴定得到的非编码转录本的交集, 共得到1 250个共有Lnc-RNA (图 4A)。将mRNA与预测出的Lnc-RNA进行长度分布密度比较, 可知在滇黄精非全长序列中, Lnc-RNA与mRNA转录本长度相似, 集中在0~600 bp (图 4B)。
4 转录因子注释
对滇黄精样本中具有转录因子(transcription factor, TF) 功能的转录本进行注释, 共鉴定到属于37个基因家族的1 781个(图 5)。其中含有转录本数量最多的10个家族分别是: C3H (219条)、FAR1 (164条)、C2H2 (118条)、bZIP (97条)、bHLH (90条)、MYB (90条)、NAC (86条)、SBP (86条)、TUB (79条)、GRAS (75条)。研究显示, C3H、FAR1、C2H2、bZIP等转录因子在植物生长发育及逆境胁迫应答过程发挥重要作用; MYB和bHLH等转录因子参与了植物次生代谢的调控, 包括萜类、黄酮、木质素、糖类等物质的生物合成[20]。这些转录因子家族的鉴定为今后滇黄精次生代谢产物的生物合成, 以及逆境响应等提供了条件。
5 SSR检测及引物设计
5.1 SSR检测
通过MISA (https://webblast.ipk-gatersleben.de/misa/) 对滇黄精转录本进行SSR检测, 共检测到33 709个SSR位点, 所有碱基重复类型中, 单碱基重复最多(p1: 10 257); 最少的是五碱基重复(p5: 148), 仅为单碱基重复的0.14%, 可见不同类型的核苷酸在数量上差距较大。其他SSR类型二碱基重复(p2)、三碱基重复(p3)、四碱基重复(p4)、六碱基重复(p6)、复合型重复(s) 和SSR之间有重叠的复合型SSR (c*) 的数量分别为4 815、5 952、259、373、1 707和10 180 (表 2)。
5.2 SSR引物设计
为进一步筛选可以在试验中应用的滇黄精SSR分子标记, 采用Primer 3对滇黄精转录本进行SSR引物设计, 共设计引物24 034对, 统计了其中17条与滇黄精多糖合成有关的转录本, 结果如表 3所示。
6 多糖代谢酶基因分析
黄精多糖的生物合成是一个较为复杂的过程, 首先卡尔文循环的产物3-磷酸甘油醛(GAP) 在果糖二磷酸醛缩酶(ALDO)、果糖-1, 6-二磷酸酶(FBP)、蔗糖磷酸合成酶(SPS)、蔗糖-6-磷酸酶(SPP) 蔗糖合成酶(SUS)、磷酸葡萄糖变位酶(PGM) 等酶的作用下生成蔗糖[21]; 蔗糖在INV、PGM、尿苷二磷酸葡萄糖焦磷酸化酶(galU) 的作用下生成UDP-葡萄糖, 成为组成黄精多糖的第一类单糖; 蔗糖在HK、果糖激酶(scrK)、甘露糖-6-果糖激酶(MPI)、磷酸甘露糖突变酶(PMM) 和GMPP的作用下生成的GDP-甘露糖是组成黄精多糖的第二类单糖; GDP-甘露糖在甘露糖4, 6-脱水酶(GMDS)、甘露糖-L-岩藻糖合酶的作用下生成的GDP-岩藻糖是组成黄精多糖的第3种单糖; UDP-葡萄糖在尿苷二磷酸葡萄糖-4, 6脱水酶(RHM) 和3, 5差向异构-4-还原酶(UER1) 的作用下生成UDP-鼠李糖, 构成黄精多糖的第4种单糖, 并且在尿苷二磷酸葡萄糖-4-表异构酶(GALE) 和尿苷二磷酸-D-半乳糖脱氢酶(UGD) 的作用下生成UDP-半乳糖, 构成黄精多糖的第5种单糖; UDP-半乳糖在尿苷二磷酸-4-差向异构酶(UGE)、尿苷二磷酸芹糖/木糖合酶(AXS)、尿苷二磷酸阿拉伯糖-4-差向异构酶(UXE) 的进一步作用下生成的UDP-D-吡喃木糖和UDP-L-阿拉伯糖是构成黄精多糖的两种多糖。这些单糖最后在糖基转移酶的作用下形成黄精多糖。本实验中, 通过KEGG代谢通路富集分析, 共发现与滇黄精多糖合成相关的19个基因或基因家族的转录本348个, 其中糖酵解/糖异生代谢通路的转录本有127条, 淀粉和蔗糖代谢通路的转录本有144条, 氨基酸糖和核苷酸糖代谢通路的转录本有85条, 果糖和甘露糖代谢通路的转录本有69条。在这些基因家族或基因中, 转录本数量最多的基因家族是SUS (62条), 其次是INV (48条), 最少的是FBP和PMM, 只有1条(表 4)。
INV将蔗糖转化为葡萄糖和果糖, 是植物合成多糖的关键酶之一。将转录组测序得到的48个INV转录本表达量与滇黄精多糖含量进行相关性分析, 其中INV12、INV23、INV25、INV36、INV44、INV46与滇黄精多糖含量具有显著的正相关, 因此推测其可能在滇黄精多糖合成过程中起重要作用(图 6)。
讨论
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目前人们对于滇黄精的基因测序研究较少, 限制了滇黄精的进一步开发利用, 本研究通过PacBio测序平台首次对滇黄精的根茎进行全长转录组进行测序和分析, 去除冗余后共获得55 454个转录本, 其完整性和质量(N50, 3 212) 远高于二代(N50, 1 260[22]; N50, 1 350[23]), 可以为及后续滇黄精基因资源挖掘提供基础。通过NR数据库对比发现, 有94.40%的基因得到注释, 其中与滇黄精基因配比率最高的为天门冬科植物石刁柏(70.17%), 此外共注释到黄精属植物18个, 但其所占比例仅为0.41%, 表明黄精属植物的已有基因资源相对匮乏。GO注释和KEGG注释结果显示滇黄精的转录组功能主要集中于生物学过程与代谢过程。这些基因注释和分类信息将有助于对滇黄精基因功能的理解与深入研究。
SSR分子标记技术是常用的遗传标记方法, 具有重复性好、稳定性强等优点, 广泛用于植物种质资源评价、遗传多样性分析等领域[24]。Song等[25]利用18对SSR分子标记, 对来自山西及周边省份的62个柴胡栽培和野生居群共619个植株进行遗传多样性以及居群遗传结构分析, 为柴胡的种质资源利用、遗传变异研究以及柴胡优质种质资源开发提供理论依据。本文在滇黄精转录本中共检测到33 709个SSR位点, 单核苷酸重复到六核苷酸重复以及复合型重复和SSR之间有重叠的复合型SSR均有检测到, 其重复类型主要单碱基重复和SSR之间有重叠的复合型SSR为主, 占总SSR位点的60.68%。利用Primer 3.0软件对获得的滇黄精SSR进行引物设计, 筛选得到了24 034对SSR引物, 显示了滇黄精SSR位点的多态性。本文统计了其中与滇黄精多糖合成相关酶的转录本的SSR引物设计信息, 以期为滇黄精SSR分析以及后续的多糖合成奠定理论基础。
作为药食同源植物, 滇黄精具有十分广阔的发展前景。中国药典规定多糖作为滇黄精唯一质控标准, 因此探究滇黄精中多糖类成分的生物合成途径对于滇黄精的进一步开发利用具有重要意义。本文利用第三代转录组测序技术获得的非冗余转录本中共初步挖掘得到参与滇黄精多糖生物合成的19个基因或基因家族的348条转录本, 远远超过Wang等[10]通过二代转录组测序获得的参与滇黄精多糖生物合成的相关基因或基因家族数量, 说明本实验中使用全长转录组测序测得的转录本更加准确。在得到的19个相关基因中, 除了FBP和PMM两个基因只检测到1条转录本外, 其余基因均得到多条转录本, 其中SUS转录本高达62条, 这为后续多糖生物合成关键酶基因的挖掘提供了更多的候选基因。
随着转录组测序技术的不断发展以及数据库的不断完善。将结果与数据库进行比对, 可得到更多的滇黄精遗传信息及有助于滇黄精多糖合成途径的揭秘。本研究首次利用PacBio Sequel平台获得了滇黄精全长转录组, 分析了滇黄精多糖类成分的生物合成途径, 为滇黄精多糖生物合成的分子机制奠定了基础。
作者贡献: 米琪与赵艳丽共同完成实验内容及论文撰写工作; 徐萍负责实验样品采集以及论文撰写工作; 于梦雯、张萱和涂振华协助完成部分实验内容; 李春华为本实验提供了实验样品; 陈佳指导实验和平台支撑; 郑国伟设计方案并指导论文的撰写和修改。
利益冲突: 所有作者均声明不存在利益冲突。
基金
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  • 国家自然科学基金资助项目(82360751)
  • 云南中医药大学高层次人才计划(2019YZG07)
  • 云南青年精英人才项目(YNWR-QNBJ-2020-289)
文献
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doi: 10.16438/j.0513-4870.2023-1259
  • 接收时间：2023-11-06
  • 首发时间：2025-11-26
  • 出版时间：2024-06-12
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  • 收稿日期：2023-11-06
  • 修回日期：2024-02-06
基金
国家自然科学基金资助项目(82360751)
云南中医药大学高层次人才计划(2019YZG07)
云南青年精英人才项目(YNWR-QNBJ-2020-289)
作者信息
    1.云南中医药大学中药学院, 云南 昆明 650500
    2.云南中医药大学民族医药学院, 云南 昆明 650500
    3.贡山县农业农村局农业综合发展服务中心, 云南 怒江傈僳族自治州 673500

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*郑国伟, E-mail: ;
陈佳, E-mail:
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2种不同金属材料的力学参数
科
Family		 属数
Number of
genus		 种数
Number of
species		 占总种数比例
Percentage of
total species (%)		 属
Genus		 种数
Number of
species		 占总种数比例
Percentage of total
species (%)
鹅膏菌科Amanitaceae 2 11 5.26 鹅膏菌属 Amanita 10 4.78
小菇科 Mycenaceae 2 12 5.74 丝盖伞属 Inocybe 5 2.39
多孔菌科 Polyporaceae 8 14 6.70 蜡蘑属 Laccaria 5 2.39
红菇科 Russulaceae 3 23 11.00 小皮伞属 Marasmius 6 2.87
小菇属 Mycena 11 5.26
光柄菇属 Pluteus 5 2.39
红菇属 Russula 17 8.13
栓菌属 Trametes 5 2.39
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