Application of upland cotton GhFAM32Al gene in regulation and control of cotton fiber elongation

Abstract

The invention provides application of an upland cotton GhFAM32Al gene in regulation and control of cotton fiber elongation, and relates to the technical field of genetic engineering. According to the invention, a recombinant inbred line group is constructed among parents with significant difference in fiber length and QTL positioning is carried out, so that the GhFAM32Al gene located in the D02 chromosome has significant influence on the cotton fiber length. Gene overexpression and CRISPR / Cas9 knockout means are used for verification and show that high expression of GhFAM32Al in the fiber development stage can remarkably promote fiber elongation, and fiber length reduction is caused by knockout. Further research finds that GhFAM32Al is a nuclear localized natural disordered protein, can form an interaction compound with GhMYBd protein, and synergistically participates in the regulation and control of the fiber cell elongation process. The invention provides a key gene resource and a molecular regulation strategy for high-quality breeding of cotton.

Description

TECHNICAL FIELD

[0001] The application relates to the technical field of genetic engineering, and particularly relates to a Gossypium hirsutum GhFAM32Al gene in regulating the elongation of cotton fibers. BACKGROUND

[0002] With the improvement of people's living standards, the demand for natural pure cotton fabrics is increasing, and the quality requirements are also higher and higher. The structural reform of China's agricultural supply side points out that the focus should be shifted from one-sided pursuit of high yield to consideration of yield, benefit and quality, and the key is to improve the quality of cotton and cultivate new varieties of "double 30" (fiber length is 30 mm, and fiber strength is more than 30 cN / tex). Therefore, improving the fiber quality of Gossypium hirsutum and cultivating varieties with excellent fiber quality are key problems urgently needed to be solved in cotton breeding. Among the main quality traits of cotton, fiber length is considered to be one of the most important qualities affecting yarn quality.

[0003] With the transfer of cotton land to grain to dry and thin saline-alkali land in China and the increasing demand for high-end raw cotton in the textile industry, cultivating high-yield and high-quality new varieties has become a major demand for the development of the cotton industry. Therefore, mining functional genes related to fiber length is the key to cotton molecular breeding improvement, which provides strong technical support for realizing direct selection and effective aggregation of excellent cotton fiber genes, ensuring the sustainable development and production safety of China's cotton industry, and meeting the needs of the rapid development of the cotton textile industry and the improvement of people's living standards.

[0004] FAM32A plays a potential role in regulating cell cycle, apoptosis and tumor occurrence, and has attracted more and more attention in recent years. Although its exact biological function has not been fully elucidated, recent studies have shown that FAM32A may play a key regulatory role in various life processes. For example, in the study of human spliceosome, FAM32A, as an exon connector factor, interacts with CACTIN, SDE2 and NKAP, etc., and promotes exon ligation by bridging 5'-exon and 3'-splicing site to stabilize the catalytic conformation. In plants, FAM32Al protein can be significantly enriched in the resistance process of strawberry fruits as an intrinsically disordered protein, which indicates that it is a potential disorder protein related to resistance. SUMMARY

[0005] In order to overcome the deficiencies of the prior art, the purpose of the present application is to provide a Gossypium hirsutum GhFAM32Al gene in regulating the elongation of cotton fibers, by regulating the expression level of the Gossypium hirsutum GhFAM32Al gene, the positive regulation of the length of cotton fibers is realized, and key gene resources and technical means are provided for the molecular breeding of high-quality cotton varieties.

[0006] To achieve the above-mentioned purpose, the present application provides the following scheme: Gossypium hirsutum GhFAM32Al Application of the gene in regulating cotton fiber elongation, by increasing GhFAM32Al gene expression level to increase cotton fiber length, by reducing or knocking out GhFAM32Al gene to decrease cotton fiber length.

[0007] Preferably, GhFAM32Al The open reading frame of the gene is 348 bases in length, encoding 115 amino acids, and the gene number is GhM_D02G0237 Located on chromosome D02 of Gossypium hirsutum.

[0008] Preferably, wherein the increase GhFAM32Al Gene expression is achieved by constructing an overexpression vector containing GhFAM32Al Coding sequence and introducing into cotton plants.

[0009] Preferably, wherein the overexpression vector is introduced into cotton by Agrobacterium-mediated genetic transformation.

[0010] Preferably, wherein the reduction or knockout GhFAM32Al Gene is achieved by CRISPR / Cas9 gene editing technology to generate frameshift mutations in GhFAM32Al Exon region so as to make the protein translation terminate early.

[0011] Preferably, wherein the gene editing mutation results in complete deletion of GhFAM32Al protein in transgenic cotton.

[0012] Preferably, the transcription level and protein level changes are verified by real-time fluorescent quantitative PCR and Western blot at the same time. GhFAM32Al

[0013] Preferably, the transgenic cotton fiber length measured at the development stage of five to twenty days after flowering is significantly different from the corresponding wild type control.

[0014] A transgenic cotton plant obtained by the above application, which comprises GhFAM32Al Expression cassette and exhibits significantly higher cotton fiber length than the corresponding wild type plant.

[0015] Preferably, the GhFAM32Al protein and the GhMYBd protein form an interaction complex in the nucleus, and the presence of the complex is verified by yeast two-hybrid Y2H, bimolecular fluorescence complementation BiFC, dual luciferase complementation LUC or GST pull-down experiment.

[0016] According to the specific embodiments provided by the application, the following technical effects are disclosed: The application locates a key gene controlling cotton fiber elongation by forward genetics​GhFAM32Al , through CRISPR / Cas9 mutants and super-expression transgenic cotton, it is creatively found that GhFAM32Al In the function of regulating cotton fiber elongation, important gene resources are provided for high-quality molecular breeding of cotton.

[0017] The application can obtain cotton plants with fiber elongation by super-expression, Figure 1 The fiber length of the super-expression cotton strain is significantly increased, which provides a new way for high-quality breeding of cotton. BRIEF DESCRIPTION OF DRAWINGS

[0018] In order to more clearly illustrate the technical solutions in the embodiments of the present application or the prior art, the drawings needed in the embodiments will be briefly introduced below. Obviously, the drawings in the following description are only some embodiments of the present application, and other drawings can be obtained by those skilled in the art without creative labor.

[0019] qFL-D02-1 The application provides GhM_D02G0237 Fine mapping and Figure 2 Expression pattern analysis schematic diagram; GhM_D02G0237 The application provides GhFAM32Al ( Positioning schematic diagram on the nucleus; Figure 3 GhFAM32Al The application provides Figure 4 Transcription and protein expression level analysis schematic diagram in super-expression strain and mutant; GhFAM32Al The application provides Figure 5 Positive regulation of fiber elongation schematic diagram; GhFAM32Al The application provides GhFAM32Al and GhMYBd interaction schematic diagram. DETAILED DESCRIPTION

[0020] The technical solutions in the embodiments of the present application will be described clearly and completely below with reference to the drawings in the embodiments of the present application. Obviously, the described embodiments are only some embodiments of the present application, not all. Based on the embodiments in the present application, all other embodiments obtained by those skilled in the art without creative labor are within the scope of protection of the present application.

[0021] The purpose of the present application is to provide a Gossypium hirsutum GhFAM32Al The application of the gene in regulating cotton fiber elongation, by regulating Gossypium hirsutum qFL-D02-1 ​The gene expression level was positively regulated to improve cotton fiber length, providing key gene resources and technical means for molecular breeding of high-quality cotton varieties.

[0022] To make the above-mentioned objects, features and advantages of the present invention more apparent and understandable, the present invention will be further described in detail below with reference to the accompanying drawings and specific embodiments.

[0023] This embodiment first constructs a population of 588 upland cotton recombinant inbred lines (RILs) using the parents Nongda Cotton 13 (NDM13) and Nongda 601 (ND601), which exhibit significant differences in fiber length. Using this RIL population, cotton fiber length phenotypic identification was performed in eight environments. Through linkage analysis, a stable new QTL controlling fiber elongation was located on chromosome D02. qFL-D02-1 Further utilize the RIL population GhM_D02G0234 The remaining heterozygous line RIL189 was self-crossed to obtain a secondary F2 segregating population of nearly 4,000 lines. The QTL interval was narrowed to 64 kb using KASP markers developed based on parental resequencing.

[0024] There are 7 genes in this interval ( GhM_D02G0235 , GhM_D02G0236 , GhM_ , D02G0237 GhM_D02G0238 , GhM_D02G0239 , GhM_D02G0240 and GhM_D02G0238 Using transcriptome analysis of fibers from different tissues and organs of upland cotton and at different developmental stages of the parent plants, we found... GhM_D02G0239 It is hardly expressed in fibers of various organs and different developmental stages; GhM_ It is mainly expressed in leaves and calyxes, and almost not expressed in fibers; D02G0240 GhM_ Although it is expressed in all tissues, the expression level in fibers is extremely low and there is no significant difference between parents; D02G0237 GhM_D02G0237 It is expressed in multiple tissues, especially in ovules, and shows differential expression specific to fiber development stages between the parents NDM13 and ND601. For example, it is highly expressed in fiber tissue on the day of flowering (0 DPA, days post-anther), with higher expression levels in NDM13 than in ND601. However, during fiber elongation and secondary cell wall thickening stages, the expression level in ND601 is 2-3 times that in NDM13, indicating... GhM_D02G0237 It may play a key role in fiber initiation, cell elongation, and secondary wall thickening.

[0025] GhFAM32AlIt encodes a FAM32A-like protein, hence the name. GhFAM32Al Its open reading frame is 348 bp long, consisting of a protein sequence of 115 amino acids, with a molecular weight of approximately 13.18 kDa. Secondary structure prediction indicates that GhFAM32Al is a naturally occurring disordered protein, exhibiting typical characteristics of this type of protein. Subcellular localization analysis shows that GhFAM32Al is located in the cell nucleus, making it a nuclear localized protein, suggesting that GhFAM32Al may play a role in transcriptional regulation or other nuclear processes related to cell development.

[0026] To clarify fam32al-1 To investigate the role of this gene in fiber development, this embodiment constructed gene-edited and overexpressed transgenic lines for this gene. High-throughput sequencing analysis of the target site mutation characteristics yielded three independent knockout lines (named...). fam32al-2 , fam32al-3 and fam32al-1 ).in fam32al-2 It is a biallelic mutant line, with one allele having a 1 bp insertion and the other allele having a 1 bp deletion; fam32al-3 Carrying a homozygous 1 bp deletion; GhFAM32Al It is also a biallelic mutant line, containing a 1 bp insertion and a 5 bp deletion. These deletion mutations are all located in... GhFAM32Al Exon regions and all lead to GhFAM32Al A frameshift mutation occurs, leading to loss of gene function. qRT-PCR analysis showed that in fibers 10 days after flowering (10 DPA), the overexpressing lines... GhFAM32Al Transcriptional levels were significantly increased by 4-6 times compared to wild-type (WT). Western blot analysis further confirmed a significant increase in GhFAM32Al protein content in the overexpression lines, while the knockout lines completely lacked this protein. Fiber dynamic elongation analysis showed that during fiber development at 5-20 DPA, the fiber length in the overexpression lines was consistently higher than that in the wild type, while the fiber length in the mutants (especially in the later stages of fiber development) was significantly shorter than that in the wild type. GhFAM32Al Positive regulation of fiber elongation.

[0027] To explore further GhFAM32Al The molecular function, in this embodiment will GhMYBd The CDS sequence was ligated into the pGBKT7 vector as bait, and yeast libraries from cotton fibers were used as prey for yeast two-hybrid (Y2H) screening of interacting proteins. A candidate interacting protein was identified through screening and named GhMYBd (encoding myb-like protein D). Transcriptome data from upland cotton NDM8 were analyzed... GhMYBd The expression patterns in cotton roots, stems, leaves, pistils, stamens, calyx, petals, receptacles, and other tissues, as well as ovules and fibers at different developmental stages, showed that...GhFAM32Al It is expressed in all organs of cotton, but the expression level is higher in petals and sepals. In order to verify the interaction between GhFAM32Al and GhMYBd, this embodiment uses BiFC, LUC and GST pull-down experiments to verify the interaction between GhFAM32Al and GhMYBd, which proves that there is an interaction between GhFAM32Al and GhMYBd, indicating that they may act together as part of the cotton fiber development regulation complex.

[0028] The biological materials used in this embodiment are introduced as follows: E. coli Top10 and Agrobacterium GV3101 were purchased from Shengong Biotechnology Co., Ltd., with item numbers B528412 and B528430, respectively. The pGreenII0800-LUC vector was used for transient expression of LUC in tobacco, and the pCAMBIA1304 vector was used for transient expression of GUS in tobacco. Both vectors are commercial vectors, which were preserved in the laboratory in the early stage. The pCambia2300-Cas9 was used to construct the CRISPR / Cas9 gene editing vector, which came from the article “Ge X Y, et al. Efficient genotype-independent cotton genetic transformation and genome editing. Journal of Integrative Plant Biology, 2023, 65(4): 901-917”. The cotton germplasm materials used were collected and preserved in the laboratory, which came from the article “Ma Z Y, et al. High-quality genome assembly and resequencing of modern cotton cultivars provide resources for crop improvement. Nature Genetics, 2021, 53(9): 1385-1391”.

[0029] The nucleotide sequences of the genes or the amino acid sequences of the proteins used in this embodiment are as follows: GhFAM32Al Coding sequence (SEQ ID NO. 1): ATGTCGGATGCATACGAGAAAGTGAAAGGAGGTAGATTAACCTTCAAAGGAGGAAGCTTAGCGACTCGCAAATCAATCGACAAAAGCAAGAAGAAGCACAAGAAGAAAAAGATCGCCGAAGATGATACTCAGCCTACCCTCGACGCCTCCACCGTTTCCGTCGAAGGTTCTGAA TCTGGTGGAGCGGATATCTACACCATCGACCGCTGCTAAAAAGATGAAGTACGAGGAGCTGTTCCCCGTCGAGGCTAGGAAGTTTCGGTTACGACCCTAAAAACCCTAAAGCCACATCAGTGGAACAAGCCCTCGATGATCGTGTTAAGAAGAAAGCCGATCGTTACTGTAAATGA GhFAM32Al amino acid sequence: MSDAYEKVKGGRLTFKGGSLATRKSIDKSKKKHKKKKIAEDDTQPTLDASTVSVEGSESGGADIYTIDAAKKMKYEELFPVEARKFGYDPKNPKATSVEQALDDRVKKKADRYCK Example 1 qFL-D02-1 Related to fiber elongation This invention first constructed a recombinant inbred line population containing 588 lines using the long-fiber variety Nongda Cotton 13 and the general-fiber-length variety Nongda 601 as parents. Phenotypic identification of fiber length was conducted in eight environments across multiple regions between 2017 and 2019. Further whole-genome resequencing was performed on both parents (22.4×) and offspring (4.3×), yielding a total of 1,909,511 polymorphic SNPs, including 232,946 aa×bb SNPs that showed no abnormal genotypes in the offspring population. Through QTL linkage analysis, a stable QTL was located on chromosome D02, detectable in all seven environments and with a phenotypic variance explanation rate of 14.65%. Figure 1 The physical range is between 2.84 Mbp and 3.83 Mbp (TM-1HAU v1.0), and a total of 43 genes were identified within this range, such as... qFL-D02-1 As shown in Part A of the diagram.

[0030] To determine the gene that truly controls fiber elongation in this region, this invention screened for a gene in the RIL population. Figure 1The remaining heterozygous line RIL189 was self-crossed to construct a secondary F2 segregating population. Recombinant single plants were screened using seven pairs of KASP markers developed based on parental resequencing. Phenotypic analysis of the recombinant single plants revealed that the fiber length of lines with the NDM13 genotype (KASP1_01-KASP_11) was significantly higher than that of lines with the ND601 genotype. Therefore, the QTL interval was narrowed down to 230 kb (e.g., ...). Figure 1 (As shown in Part B); Further, using the recombinant heterozygous line LC121, which exhibits heterozygosity in the KASP1_01-KASP_11 state within the secondary population, a new secondary F2 segregating population was constructed through self-pollination. Recombinant single plants were screened using eight newly developed KASP marker pairs. It was found that the fiber length of lines with the NDM13 genotype in KASP1_04-KASP_06 was significantly higher than that of the ND601 genotype lines, narrowing the candidate interval to 64 kb (as shown in Part B). GhM_D02G0234 (As shown in section C).

[0031] There are 7 genes in this interval ( GhM_D02G0235 , GhM_D02G0236 , GhM_ , D02G0237 GhM_D02G0238 , GhM_D02G0239 , GhM_D02G0240 and GhM_D02G0238 Using transcriptome analysis of fibers from different tissues and organs of upland cotton and at different developmental stages of the parent plants, we found... GhM_D02G0239 It is hardly expressed in fibers of various organs and different developmental stages; GhM_ It is mainly expressed in leaves and calyxes, and almost not expressed in fibers; D02G0240 GhM_ Although it is expressed in all tissues, the expression level in fibers is extremely low and there is no significant difference between parents; D02G0237 GhM_D02G0237 It is expressed in multiple tissues, especially in ovules, and shows differential expression specific to the fiber development stage between the parents NDM13 and ND601. High expression is observed in fiber tissue on the day of flowering (0 DPA), with NDM13 showing higher expression than ND601. However, during fiber elongation and secondary wall thickening stages, the expression level in ND601 is 2-3 times that in NDM13, indicating... Figure 1 It may play a key role in fiber initiation and secondary wall thickening (e.g. GhM_D02G0237 (As shown in sections D to F).

[0032] GhFAM32Al It encodes a FAM32A-like protein, hence the name. GhFAM32Al Its open reading frame is 348 bp long, consisting of a protein sequence of 115 amino acids, with a molecular weight of approximately 13.18 kDa. Secondary structure prediction indicates... GhFAM32AlIt belongs to the category of naturally occurring disordered proteins (IDPs) and exhibits typical characteristics of this type of protein. Subcellular localization analysis indicates... Figure 2 It is a nuclear localization protein ( GhFAM32Al ), foreshadowing GhFAM32Al It may play a role in transcriptional regulation or other nuclear processes related to cell development.

[0033] Example 2 GhFAM32Al Positive regulation of fiber elongation To clarify GhFAM32Al Regarding its role in fiber development, this embodiment utilizes Agrobacterium-mediated genetic transformation. fam32al-1 Gene editing and overexpression transformation were performed. High-throughput sequencing analysis was used to identify the mutation characteristics at the target sites, resulting in three independent knockout lines (named...). fam32al-2 , fam32al-3 and fam32al-1 ),in fam32al-2 It is a biallelic mutant line, with both alleles having a 1 bp deletion; fam32al-3 Carrying a homozygous 1 bp deletion; GhFAM32Al It is also a biallelic mutant line, containing a 1 bp insertion and a 5 bp deletion. These mutations are all located in... Figure 3 Mutations in the exon regions, and frameshift mutations that lead to premature termination of translation (e.g.) GhFAM32Al (As shown in Part A of the middle section).

[0034] In order to evaluate GhFAM32Al To investigate the expression of overexpression in cotton lines, this example utilizes real-time quantitative PCR (qRT-PCR) analysis of overexpressed and wild-type (WT) 10 DPA fibers. The results showed that, compared to WT, all overexpression lines... Figure 3 The expression level was approximately 4 to 6 times that of the WT strain (e.g., Figure 3 (As shown in Part B). Meanwhile, Western blot results showed that the GhFAM32Al protein content was significantly increased in the overexpression lines (e.g., ...). GhFAM32Al (as shown in section C), but not observed in mutant lines.

[0035] To clarify GhFAM32Al The role of fiber elongation in mutant, overexpressing, and wild-type cotton lines was dynamically observed at 5, 10, 15, and 20 DPA after flowering. Results showed that during 5-20 DPA, the fiber length of the overexpressing lines was consistently longer than that of the wild type, while the fibers of the mutants (especially in the later stages) were significantly shorter than those of the wild type. This confirms... Figure 4 Positive regulation of fiber elongation ( Figure 5 ).

[0036] Example 3: The interaction between GhFAM32Al and GhMYBd jointly regulates fiber elongation. Since the functional annotation of GhFAM32Al could not be retrieved from public protein databases, this embodiment investigated its potential cellular role by analyzing its subcellular localization. Therefore, this embodiment constructed a fusion protein expression vector (35S::GhFAM32Al-GFP) containing green fluorescent protein (GFP) and GhFAM32Al and transformed it into GV3101. Through transient expression in the lower epidermal cells of tobacco leaves, the expression location of GFP in tobacco leaves was observed using confocal fluorescence microscopy. The results showed that the green fluorescence signal of the empty GFP control was distributed throughout the nucleus and the entire cytoplasm, while the fluorescence signal of the GhFAM32Al-GFP fusion protein was only distributed in the nucleus, indicating that GhFAM32Al is a nuclear-localized protein (e.g., GFP is found in the nucleus and cytoplasm). GhFAM32Al (As shown in Part A). This result suggests that GhFAM32Al may play a role in transcriptional regulation or other nuclear processes related to cell development.

[0037] To explore further GhFAM32Al The molecular function, in this embodiment will GhMYBd The CDS sequence was ligated into the pGBKT7 vector as bait, and yeast libraries from cotton fibers were used as prey for screening interacting proteins using yeast two-hybrid Y2H. A candidate interacting protein was identified through screening and named GhMYBd (encoding myb-like protein D). Transcriptome data from upland cotton NDM8 were analyzed... GhMYBd The expression patterns in cotton roots, stems, leaves, pistils, stamens, calyx, petals, receptacles, and other tissues, as well as ovules and fibers at different developmental stages, showed that... Figure 5 It is expressed in all organs of cotton, but the expression level is higher in petals and sepals (e.g., GhMYBd (As shown in Part B).

[0038] To verify the interaction between GhFAM32Al and GhMYBd, GhFAM32Al The CDS sequence was ligated to the pGADT7 vector and then ligated to the pGBKT7 vector. Figure 5The recombinant vectors were co-transformed into Y2H-gold cells and cultured on SD / -Leu / -Trp / -His / -Ade medium. Results showed that the positive control pGBKT7-53+pGADT7-T plates exhibited colony growth, indicating the effectiveness of the yeast experimental system. However, the negative controls pGBKT7-lam+pGADT7-T, pGBKT7-GhFAM32Al+pGADT7-T, and pGBKT7+pGADT7-GhMYBd plates showed no colony growth, while pGBKT7-GhFAM32Al+pGADT7-GhMYBd showed colony growth, indicating an interaction between GhFAM32Al and GhMYBd (e.g., ...). Figure 5 (As shown in section C).

[0039] Because the GhMYBd-GFP fusion protein is located in the cell nucleus (e.g. Figure 5 As shown in section D), to verify whether the interaction between GhFAM32Al and GhMYBd exists in plants, this example performed a bimolecular fluorescence complementation (BiFC) experiment in tobacco leaves. The results showed that when the fusion proteins GhFAM32Al-nYFP and GhMYBd-cYFP or GhMYBd-nYFP and GhFAM32Al-cYFP were co-expressed in the lower epidermal cells of tobacco leaves, YFP fluorescence could be specifically observed in the cell nucleus. This nuclear-localized fluorescence signal confirmed the physical interaction between GhFAM32Al and GhMYBd in the cell nucleus (e.g., Figure 5 (As shown in section D); and the Dual-Luciferase Complementation Assay (LUC) also yielded similar results (as shown in section D). GhFAM32Al (As shown in section E). Furthermore, GST-labeled GhFAM32Al protein was co-incubated with His-labeled GhMYBd, unbound protein was washed away, and SDS-PAGE electrophoresis was performed on the boiled beads to reveal GST-... Figure 5 The His-GhMYBd protein was successfully pulled down, resulting in a protein band of approximately 41.6 kDa, indicating a specific and direct interaction between the two proteins (e.g., ​ (As shown in section F).

[0040] In summary, the results of Y2H, BiFC, LUC, and Pull-down experiments indicate that there is an interaction between GhFAM32Al and GhMYBd in the cell nucleus, suggesting that they may act as part of the cotton fiber development regulatory complex in regulating fiber elongation.

[0041] The various embodiments in this specification are described in a progressive manner, with each embodiment focusing on its differences from other embodiments. Similar or identical parts between embodiments can be referred to interchangeably. For the systems disclosed in the embodiments, since they correspond to the methods disclosed in the embodiments, the descriptions are relatively simple; relevant parts can be referred to the method section.

[0042] This document uses specific examples to illustrate the principles and implementation methods of the present invention. The descriptions of the above embodiments are only for the purpose of helping to understand the method and core ideas of the present invention. Furthermore, those skilled in the art will recognize that, based on the ideas of the present invention, there will be changes in the specific implementation methods and application scope. Therefore, the content of this specification should not be construed as a limitation of the present invention.

Claims

1. An application of the upland cotton GhFAM32Al gene in regulating cotton fiber elongation, characterized in that, Cotton fiber length can be increased by increasing the expression level of the GhFAM32Al gene, and cotton fiber length can be decreased by decreasing or knocking out the GhFAM32Al gene.

2. The application according to claim 1, characterized in that, The open reading frame of the GhFAM32Al gene is 348 bases long and encodes 115 amino acids. The gene number is GhM_D02G0237 and it is located on chromosome D02 of upland cotton.

3. The application according to claim 1, characterized in that, The expression of the GhFAM32Al gene was enhanced by constructing an overexpression vector containing the GhFAM32Al coding sequence and introducing it into cotton plants.

4. The application according to claim 3, characterized in that, The overexpression vector was introduced into cotton through Agrobacterium-mediated genetic transformation.

5. The application according to claim 1, characterized in that, The reduction or knockout of the GhFAM32Al gene involves using CRISPR / Cas9 gene editing technology to create frameshift mutations in the exon region of GhFAM32Al, thereby prematurely terminating protein translation.

6. The application according to claim 5, characterized in that, The GhFAM32Al protein, resulting from the gene editing mutation, is completely absent in the transgenic cotton.

7. The application according to any one of claims 3 to 6, characterized in that, The transcriptional and protein levels of GhFAM32Al were simultaneously verified by real-time quantitative PCR and Western blot.

8. The application according to any one of claims 3 to 6, characterized in that, The fiber length of transgenic cotton measured during the developmental stage from five to twenty days after flowering showed a consistently significant difference compared to the corresponding wild-type control.

9. A transgenic cotton plant obtained by application according to any one of claims 3 to 6, characterized in that, This plant contains the GhFAM32Al expression cassette and exhibits significantly longer cotton fiber lengths than the corresponding wild-type plant.

10. The application according to any one of claims 1 to 6, characterized in that, GhFAM32Al protein and GhMYBd protein form an interaction complex in the cell nucleus. The presence of the complex was verified by yeast two-hybrid Y2H, bimolecular fluorescence complementary BiFC, dual luciferase complementary LUC, or GST pull-down assays.

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