ZmLLG1 gene affects the development of the maize root system

ZmLLG1 gene affects the development of the maize root system | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article ZmLLG1 gene affects the development of the maize root system Siyu Sun, Yulu Yang, Richard Shan, Hao Wu, Hongxue Xie This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-2965453/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Nutrient uptake by plant roots seriously affects the growth and yield of maize. AUX/IAA encoding genes play an important role in auxin signal transduction and regulating the growth of plant roots.In this study, AUX/IAA gene ZmLLG1 was cloned from maize. The expression level of the ZmLLG1 gene in different tissues was detected by qRT-PCR, and ZmLLG1 was found to be overexpressed in the roots of maize. Arabidopsis thaliana and mutant maize were both analyzed by statistical method. A yeast two-hybrid experiment, BiFC experiment and pull-down experiment were used to verify the interactions between the protein and ZmLLG1.The CoDing Sequence (CDS) of ZmLLG1 is 684 bp and encodes a protein composed of 227 amino acids. A subcellular localization analysis confirmed that ZmLLG1 was localized in the nucleus. The ZmLLG1 gene was expressed in all tissues of maize, but its expression level was highest in the root. The root length and number of lateral roots of zmllg1 mutant maize were much lower than those of wild-type B73 maize, and ZmLLG1 was responsive to auxin. The yeast two-hybrid assay, BiFC experiment and pull-down assay showed that ZmLLG1 could interact with ZmARF5, ZmARF7, and ZmARF25 to regulate the growth and development of the maize root system. Keywords AUX/IAA, Root, ARF, Maize mutant, Protein AUX/IAA Root ARF Maize mutant Protein Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Full Text Supplementary Files SupplementalTableS1.pdf SupplementalTableS2.pdf rawstatisticsofZmLLG1.xlsx rawstatistics4C4D.xlsx rawstatistics5F5M.xlsx Authorcontributions.pdf Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-2965453","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":206443361,"identity":"3238e221-ef4a-4fe0-a6d0-1e82e9e13aea","order_by":0,"name":"Siyu Sun","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA1klEQVRIie3RsQrCMBCA4QuBuJy4VpS+gRApCEKxr5IidGp3N1sKdim4dvAtBOeTrH2AbuLi1EFxFXEVl8ZNMP/+cXccgM32gw1626t+oL/Ock5mZFjSlHAcsbzYKDMiG+UR+poVZS0NNyOlyIkj3q/iW9PCwp2kHYKlRCRrXwyrZD/fwdKbUQfhLEspLCOcVslhhEDhoYsIzoGOT+0ETXwxIygEHFPUkpW1MCMOImjASLFi48130uCW4NTyO6CvWM7PTbtauJ3kY6Tpa97It8Jms9n+ohcVzEXY3SB8VQAAAABJRU5ErkJggg==","orcid":"https://orcid.org/0009-0000-5536-4942","institution":"Anhui Agriculture University: Anhui Agricultural University","correspondingAuthor":true,"submittingAuthor":false,"prefix":"","firstName":"Siyu","middleName":"","lastName":"Sun","suffix":""},{"id":206443362,"identity":"ef84a70a-6553-44fd-85c8-d0c568ad99bf","order_by":1,"name":"Yulu Yang","email":"","orcid":"","institution":"Anhui Agriculture University: Anhui Agricultural University","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Yulu","middleName":"","lastName":"Yang","suffix":""},{"id":206443363,"identity":"2baec9eb-a8c3-4b29-b64f-376776e75144","order_by":2,"name":"Richard Shan","email":"","orcid":"","institution":"Ingenix Inc Auburndale: Optum Inc","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Richard","middleName":"","lastName":"Shan","suffix":""},{"id":206443364,"identity":"34d97cb5-cd00-4a89-ba07-b20ec96dbf80","order_by":3,"name":"Hao Wu","email":"","orcid":"","institution":"Ingenix Inc Auburndale: Optum Inc","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Hao","middleName":"","lastName":"Wu","suffix":""},{"id":206443365,"identity":"8bac1484-03fe-4fdb-993b-6d7d54b20104","order_by":4,"name":"Hongxue Xie","email":"","orcid":"","institution":"West Anhui University","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Hongxue","middleName":"","lastName":"Xie","suffix":""}],"badges":[],"createdAt":"2023-05-22 08:56:50","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-2965453/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-2965453/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":38098657,"identity":"2e6f1933-ee0d-4a10-a3aa-6a13d7fca6ff","added_by":"auto","created_at":"2023-06-06 14:18:00","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":120529,"visible":true,"origin":"","legend":"\u003cp\u003eStructural analysis of ZmLLG1. (A) Schematic diagram of the ZmLLG1 gene structure. (B) Schematic diagram of the domain of ZmLLG1.\u003c/p\u003e","description":"","filename":"Figure1.png","url":"https://assets-eu.researchsquare.com/files/rs-2965453/v1/44d1754539591b6aca26951f.png"},{"id":38098665,"identity":"54a45a06-c6d6-4161-b9cf-2083dedc1c1c","added_by":"auto","created_at":"2023-06-06 14:18:00","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":387052,"visible":true,"origin":"","legend":"\u003cp\u003eTissue expression pattern analysis of the ZmLLG1 gene. (A) Heat map analysis of the ZmLLG1 gene. (B) Histogram of the expression of ZmLLG1 in different maize tissues.\u003c/p\u003e","description":"","filename":"Figure2.png","url":"https://assets-eu.researchsquare.com/files/rs-2965453/v1/67f841e62fce5527df8e22df.png"},{"id":38098658,"identity":"efe5f633-fb6a-4fe7-a61d-7f8e693d3661","added_by":"auto","created_at":"2023-06-06 14:18:00","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":297767,"visible":true,"origin":"","legend":"\u003cp\u003eSubcellular localization of ZmLLG1. (A) pMDC43 empty carrier and ZmLLG1-pMDC43 expression vector infographic; (B) pMDC43 and ZmLLG1 proteins located in the maize nucleus in the protoplast (RFP represents the mCherry fluorescence signal, the green color represents the GFP fluorescence signal, Bright field stands for bright field of view and the yellow color represents the merged fluorescence signal).\u003c/p\u003e","description":"","filename":"Figure3.png","url":"https://assets-eu.researchsquare.com/files/rs-2965453/v1/a227afd0103fc5598cdc0f40.png"},{"id":38103166,"identity":"facc9be0-1d94-4fd6-b22e-4534feea9065","added_by":"auto","created_at":"2023-06-06 14:42:00","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":905784,"visible":true,"origin":"","legend":"\u003cp\u003ePhenotypic analysis of ZmLLG1 overexpression lines in Arabidopsis thaliana. (A) Three transgenic Arabidopsis ZmLLG1-overexpression lines (OE1, OE2 and OE3) and Col-0 grown on MS medium. (B) Three transgenic Arabidopsis ZmLLG1-overexpression lines (OE1, OE2 and OE3) and Col-0 grown on MS medium supplemented with 0.3 μM NAA.(C-D) Root length and lateral root number statistics of three transgenic Arabidopsis ZmLLG1-overexpression lines (OE1, OE2 and OE3) and Col-0. Data represent mean ± SE of three replicates. Significant differences between the transgenic lines and Col-0 control were determined at **P \u0026lt; 0.01 using two-way ANOVA.\u003c/p\u003e","description":"","filename":"Figure4.png","url":"https://assets-eu.researchsquare.com/files/rs-2965453/v1/746de3b8172fae13f8bd3dae.png"},{"id":38098660,"identity":"b3eff8c8-0e17-495f-93d5-d60878d173da","added_by":"auto","created_at":"2023-06-06 14:18:00","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":1098996,"visible":true,"origin":"","legend":"\u003cp\u003ePhenotypic analysis of zmllg1 mutant maize. (A) Total root growth phenotype of wild type maize and mutant zmllg1 maize. The experimental materials were all taken from maize seedlings cultured for 10 days. (B) Schematic diagram of the primary roots of wild-type maize and mutant zmllg1 maize grown for about 10 days. (C) Schematic of the crown root of wild-type maize and mutant zmllg1 maize grown for about 10 days. (D-E) The root traits and aboveground vegetation of mutant zmllg1 maize and wild-type maize were compared at maturity, and the experimental materials were all taken from maize grown in the field for 65 days. (F-M) Statistics of root length and lateral root number at seedling and maturity stage of maize. Data represent mean ± SE of three replicates. Significant differences between the mutant maize and WT control were determined at **P \u0026lt; 0.01 using one-way ANOVA. Bars = 3 cm in A；Bars = 4 cm in B-C；Bars = 5 cm in D；Bars = 20 cm in E\u003c/p\u003e","description":"","filename":"Figure5.png","url":"https://assets-eu.researchsquare.com/files/rs-2965453/v1/b6b29f86442026cde053f994.png"},{"id":38098659,"identity":"ff61e238-847d-4e9b-b37d-ef5ebde84abb","added_by":"auto","created_at":"2023-06-06 14:18:00","extension":"png","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":122639,"visible":true,"origin":"","legend":"\u003cp\u003eExpression analysis of ZmLLG1 gene treated with auxin. (A) The expression of ZmLLG1 gene in wild-type maize treated with 0.2 μM NAA was analyzed by qRT-PCR. (B) The expression of ZmLLG1 gene in wild-type maize treated with 5 μM NAA was analyzed by qRT-PCR. Maize root samples were taken at 0 h, 1 h, 3 h, 6 h, 9 h and 12 h after adding different concentrations of auxin at seedling stage.\u003c/p\u003e","description":"","filename":"Figure6.png","url":"https://assets-eu.researchsquare.com/files/rs-2965453/v1/7d2c0975235dbb9ec7ca6d0e.png"},{"id":38099687,"identity":"37f73179-3066-46b3-8b18-a303d455e534","added_by":"auto","created_at":"2023-06-06 14:26:00","extension":"png","order_by":7,"title":"Figure 7","display":"","copyAsset":false,"role":"figure","size":791123,"visible":true,"origin":"","legend":"\u003cp\u003eExperimental analysis of interaction between ZmLLG1 and ARFs. (A) Yeast two-hybrid assays of ZmLLG1 and ZmARF 5/7/25. All the experiments were recreated at least three times and yielded similar results. PGADT7-T and PGBKT7-53 were co-transformed into the yeast strain Gold2 system as the positive control, and PGADT7 and PGBKT7 were co-transformed into the yeast strain Gold2 system as the negative control. (B) Bimolecular fluorescence complementation assays for ZmLLG1 and ZmARF 5/7/25 (YFP stands for Yellow Fluorescent Protein, RFP represents the mCherry fluorescence signal, Bright field stands for Bright field of view, Merged represents the fusion protein). Maize protoplasts showed yellow fluorescence, indicating that ZmLLG1 interacted with ZmARF 5/7/25. (C) Pull-down assays between ZmLLG1 and ZmARF5/7/25. After the samples passed through GST beads, they were hybridized with the antibody of MBP, which could be visualized on the PVDF membrane, indicating that ZmLLG1 interacted with ZmARF 5/7/25 (IN stands for internal reference, IP represent the beads class type, IB represents the type of antibody used). The internal reference was the protein expressed by the empty carrier of GST, and the protein size was 26 KDa. The protein size of ZmLLG1-GST was 49KDa. The size of the protein expressed by the empty MBP vector was 40 KDa. The protein size of ZmARF5-MBP was 142.95 KDa, ZmARF7-MBP was 156.79 KDa, ZmARF25-MBP was 147.57 KDa.\u003c/p\u003e","description":"","filename":"Figure7.png","url":"https://assets-eu.researchsquare.com/files/rs-2965453/v1/f983a3c0d2a302545b4c3ab9.png"},{"id":41268628,"identity":"f8f912db-d87b-46cd-a809-f817ef96cc81","added_by":"auto","created_at":"2023-08-08 20:18:42","extension":"pdf","order_by":1,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":854918,"visible":true,"origin":"","legend":"","description":"","filename":"SiyuSun.pdf","url":"https://assets-eu.researchsquare.com/files/rs-2965453/v1_covered_4509a167-70ca-4e0c-9849-2b87a65aa964.pdf"},{"id":38098668,"identity":"c4c767c4-8b9d-456c-ac54-45ae0754ffde","added_by":"auto","created_at":"2023-06-06 14:18:00","extension":"pdf","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":191446,"visible":true,"origin":"","legend":"","description":"","filename":"SupplementalTableS1.pdf","url":"https://assets-eu.researchsquare.com/files/rs-2965453/v1/895a842c95aea769df59da5c.pdf"},{"id":38098669,"identity":"9a7975f7-feaf-443e-afe8-7004ef25b7e8","added_by":"auto","created_at":"2023-06-06 14:18:00","extension":"pdf","order_by":2,"title":"","display":"","copyAsset":false,"role":"supplement","size":165157,"visible":true,"origin":"","legend":"","description":"","filename":"SupplementalTableS2.pdf","url":"https://assets-eu.researchsquare.com/files/rs-2965453/v1/a351bc6b00aa09c8211c8db0.pdf"},{"id":38098662,"identity":"fc7cd9e5-eed5-4f91-ac41-92cda3a6657a","added_by":"auto","created_at":"2023-06-06 14:18:00","extension":"xlsx","order_by":3,"title":"","display":"","copyAsset":false,"role":"supplement","size":336926,"visible":true,"origin":"","legend":"","description":"","filename":"rawstatisticsofZmLLG1.xlsx","url":"https://assets-eu.researchsquare.com/files/rs-2965453/v1/952be7aa48358ded6d5ec388.xlsx"},{"id":38101280,"identity":"4c57b96e-ebb3-4825-9951-e0de1549ac67","added_by":"auto","created_at":"2023-06-06 14:34:00","extension":"xlsx","order_by":4,"title":"","display":"","copyAsset":false,"role":"supplement","size":12987,"visible":true,"origin":"","legend":"","description":"","filename":"rawstatistics4C4D.xlsx","url":"https://assets-eu.researchsquare.com/files/rs-2965453/v1/e57d5275cc6e7488243669a0.xlsx"},{"id":38099685,"identity":"62851243-2942-4217-9472-ae3ae85c85e1","added_by":"auto","created_at":"2023-06-06 14:26:00","extension":"xlsx","order_by":5,"title":"","display":"","copyAsset":false,"role":"supplement","size":17835,"visible":true,"origin":"","legend":"","description":"","filename":"rawstatistics5F5M.xlsx","url":"https://assets-eu.researchsquare.com/files/rs-2965453/v1/2ad925ff4f5aecb92b5f4cfd.xlsx"},{"id":38098667,"identity":"8cd0afbf-eef1-4540-a249-4cc43d53bf5b","added_by":"auto","created_at":"2023-06-06 14:18:00","extension":"pdf","order_by":6,"title":"","display":"","copyAsset":false,"role":"supplement","size":227524,"visible":true,"origin":"","legend":"","description":"","filename":"Authorcontributions.pdf","url":"https://assets-eu.researchsquare.com/files/rs-2965453/v1/7fcc00afc25ac4f76c724883.pdf"}],"financialInterests":"","formattedTitle":"ZmLLG1 gene affects the development of the maize root system","fulltext":[],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":false,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":true,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":true,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"AUX/IAA, Root, ARF, Maize mutant, Protein","lastPublishedDoi":"10.21203/rs.3.rs-2965453/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-2965453/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"Nutrient uptake by plant roots seriously affects the growth and yield of maize. AUX/IAA encoding genes play an important role in auxin signal transduction and regulating the growth of plant roots.In this study, AUX/IAA gene ZmLLG1 was cloned from maize. The expression level of the ZmLLG1 gene in different tissues was detected by qRT-PCR, and ZmLLG1 was found to be overexpressed in the roots of maize. Arabidopsis thaliana and mutant maize were both analyzed by statistical method. A yeast two-hybrid experiment, BiFC experiment and pull-down experiment were used to verify the interactions between the protein and ZmLLG1.The CoDing Sequence (CDS) of ZmLLG1 is 684 bp and encodes a protein composed of 227 amino acids. A subcellular localization analysis confirmed that ZmLLG1 was localized in the nucleus. The ZmLLG1 gene was expressed in all tissues of maize, but its expression level was highest in the root. The root length and number of lateral roots of zmllg1 mutant maize were much lower than those of wild-type B73 maize, and ZmLLG1 was responsive to auxin. The yeast two-hybrid assay, BiFC experiment and pull-down assay showed that ZmLLG1 could interact with ZmARF5, ZmARF7, and ZmARF25 to regulate the growth and development of the maize root system. Keywords AUX/IAA, Root, ARF, Maize mutant, Protein","manuscriptTitle":"ZmLLG1 gene affects the development of the maize root system","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2023-06-06 14:17:55","doi":"10.21203/rs.3.rs-2965453/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"17268b4b-5318-495c-bdb2-7ce62e85227f","owner":[],"postedDate":"June 6th, 2023","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2023-08-12T09:29:13+00:00","versionOfRecord":[],"versionCreatedAt":"2023-06-06 14:17:55","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-2965453","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-2965453","identity":"rs-2965453","version":["v1"]},"buildId":"7rjqhiLT3MXkJMwkYKINL","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}