Potato tuberization-related transcription factor stpdf2 gene and application
By overexpressing the StPDF2 gene in potatoes, the problems of delayed tuber formation and insufficient tuber quantity were solved, enabling earlier tuber formation and increased tuber quantity. This method is suitable for the creation of early-forming tuber materials and efficient breeding.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- SOUTHWEST UNIV
- Filing Date
- 2026-05-06
- Publication Date
- 2026-06-05
AI Technical Summary
The lack of utilization of the potato StPDF2 gene in existing technologies leads to delayed tuber formation and insufficient tuber quantity, affecting potato yield and adaptability to the growing season.
By increasing the expression level of the StPDF2 transcription factor in potatoes, the StPDF2 gene was introduced into potato plants using an overexpression vector, promoting tuber formation and increasing tuber quantity.
It significantly advances tuber formation time, increases the number of tubers per plant, and does not affect the above-ground vegetative growth of the plant, showing good prospects for breeding applications.
Smart Images
Figure CN122146720A_ABST
Abstract
Description
Technical Field
[0001] This invention relates to the field of molecular genetic breeding engineering technology, specifically to a potato tuber formation-related transcription factor. StPDF2 Genes and their applications. Background Technology
[0002] potato (Solanum tuberosum L .） Potatoes are the world's fourth largest food crop, and tubers are not only the main harvested organ but also an important storage organ. The timing of tuber formation and the number of tubers directly affect potato yield, seasonal adaptability, and varietal value. Therefore, identifying functional genes that can stably promote tuber formation and establishing corresponding molecular regulatory technologies are of great significance for potato molecular design breeding.
[0003] The HD-ZIP (Homeodomain-leucine zipper) transcription factor contains a homeodomain (HD) of approximately 60-61 amino acids at its N-terminus, linked to a leucine zipper (LZ). The HD domain is responsible for recognizing cis-elements, and LZ mediates dimerization (Ruberti et al., 1991; Harris et al., 2011; Turchi et al., 2015). Its family classification is mainly based on the homology of the HD-ZIP binding domain sequence, generally divided into four subfamilies: HD-ZIP I–IV (Ariel et al., 2007; Elhitiand Stasolla, 2009). Class I and II proteins are centered around HD+LZ and specifically recognize the pseudo-palindromic sequence CAATNATTG. Class II proteins have a conserved CPSCE motif downstream of LZ and have been reported to be associated with cellular redox status (Elhiti and Stasolla, 2009; Harris et al., 2011). Class III and IV proteins have more complex structures. In addition to HD+LZ, they also carry the START lipid-binding domain and the SAD domain. Their DNA binding sites are GTAAT(G / C)ATTAC and a cis sequence containing a TAAA core, respectively (Elhiti and Stasolla, 2009; Zhao et al., 2011; Chen et al., 2014).
[0004] HD-ZIP I is involved in the regulation of abiotic stress and plant growth and development. For example, Arabidopsis thaliana AtATHB7 / AtATHB12 can be significantly induced to express under water deficiency and ABA treatment; AtATHB6It participates in the ABA signaling pathway and is associated with components such as ABI1 (Elhiti and Stasolla, 2009; Li et al., 2022); HD-ZIP II is associated with changes in light quality, shading avoidance, and auxin response. AtATHB2 / AtHAT4 It is sensitive to changes in red / far-red light and regulates shading-induced hypocotyl elongation. AtHAT2 Members of this group are significantly upregulated under auxin induction and participate in elongation growth (Harriset et al., 2011; Elhiti and Stasolla, 2009; Turchi et al., 2015). HD-ZIP III has the function of regulating organ polarity formation. REV / PHB / PHV and others regulate the development of apical structures and lateral organs in the embryo, and mutants often show organ polarity disorders. At the same time, these factors can positively regulate auxin biosynthesis, transport and response gene expression (Emery et al., 2003; Turchi et al., 2015). HD-ZIP IV induces epidermal differentiation. AtPDF2 / AtATML1 Together they maintain epidermal cell characteristics, and the double mutant exhibits epidermal-related structural abnormalities. Other members also affect trichome morphology, stratum corneum formation, and anthocyanin accumulation (Elhiti and Stasolla, 2009).
[0005] PDF2 Belonging to the HD-ZIP IV subfamily, it consists of a typical HD domain and a downstream ZIP domain. The HD domain recognizes the TAAATG(C / T)A motif, and the ZIP domain mediates homodimerization or heterodimerization (Nagata et al., 2025;). AtPDF2 It is mainly expressed in the young stage of root tip (Abe et al., 2003; Nagata et al., 2025). AtPDF2 / AtATML1 The promoter contains an L1-box cis element, and previous studies have suggested that its upstream regulation may depend on... AtATML1 Mediated by homologous factors and finely regulated by hierarchical-specific regulatory networks, but regarding AtPDF2 The molecular mechanisms underlying expression regulation remain unclear. Compared to wild-type, single mutations... AtPDF2 The genes did not change significantly, but AtPDF2 / AtATML1 Double mutations can cause embryos to stop developing during the globular stage. AtPDF2 Overexpression leads to epidermal differentiation (Abe et al., 2003). Furthermore, AtATML1 / AtPDF2 Functional defects increase seedlings' sensitivity to stresses such as drought and salinity (Nagata et al., 2025). AtPDF2 Similar factors (AtHDG / GL2 Double mutations cause abnormalities in the floral organ epidermis, accompanied by a reduction in the number of stamens (Kamata et al., 2013). Overexpression AtPDF2 The plants also exhibited delayed flowering (Abe et al., 2003). Furthermore, previous studies have shown that AtbHLH7 and AtPDF2 can form a complex to synergistically suppress the expression of multiple seed lipid synthesis-related genes (Liuet et al., 2025). However, the function of HD-ZIP transcription factors in regulating potato tuber formation and development has not yet been reported. Summary of the Invention
[0006] Addressing the lack of potato technology in existing technologies StPDF2 To address the shortcomings of existing gene function utilization technologies, this invention provides a potato tuber-related transcription factor. StPDF2 Genes and Applications. Improving [something] in potatoes StPDF2 The expression level of this substance can significantly promote tuber formation, advance tuber formation time, and increase the number of tubers per plant.
[0007] To achieve the above objectives, the technical solution provided by this invention is as follows:
[0008] This invention provides a potato tuber formation-related transcription factor. StPDF2 The gene, whose sequence number in the potato database is Soltu.DM.10G000400.1, has a base sequence as shown in SEQ ID NO. 1.
[0009] The present invention also provides a product containing the above-mentioned... StPDF2 Gene overexpression vectors.
[0010] Furthermore, the present invention also provides a method containing the above-mentioned... StPDF2 Plants with gene overexpression vectors.
[0011] Furthermore, the present invention also provides a potato tuber formation-related transcription factor. StPDF2 Applications of genes.
[0012] The present invention has the following beneficial effects: (1) This invention can significantly promote potato tuber formation. Experimental results show that after 28 days of long-day cultivation, the tuber formation is induced under short-day conditions. StPDF2 By day 6 after short-day induction, 50% of the overexpressing gene lines had formed tubers, while wild-type plants had not yet formed tubers; by day 9, StPDF2 All plants in the overexpression line formed tubers, while only 50% of the wild-type plants formed tubers, indicating that... StPDF2 Gene overexpression can significantly advance tuber formation.
[0013] (2) This invention is beneficial to increasing the number of tubers per plant. StPDF2 Gene overexpression not only promotes the initiation of tuber formation but also increases the number of tubers per plant, thus possessing high breeding value.
[0014] (3) This invention has little impact on the vegetative growth of the aboveground parts of the plant. StPDF2 The aboveground parts of the gene-overexpressing plants showed no significant morphological differences compared to the wild-type plants, indicating that the gene mainly acts on the developmental transition stage of underground organs, which is beneficial for targeted improvement of tuber formation traits.
[0015] (4) This invention has good breeding application prospects and can be used for the creation of early tuber forming materials, screening of materials with high tuber forming efficiency, and potato molecular design breeding. Attached Figure Description
[0016] Figure 1 : StPDF2 Positive detection of gene overexpression lines; where lane M is the marker, the lanes corresponding to the numbers in the figure represent different transgenic lines, "-" represents the negative control lane, and "+" represents the positive control lane.
[0017] Figure 2 : StPDF2 Results of qRT-PCR detection of gene overexpression lines; significance analysis was performed using one-way ANOVA, n = 3, with error bars ± SEM. p<0.0001.
[0018] Figure 3 : StPDF2 Gene overexpression and aboveground phenotypic analysis of wild-type lines.
[0019] Figure 4 Wild-type strains and StPDF2 Tuber formation phenotype in gene overexpression lines; where B: proportion of tuber-forming plants; C: number of stolons; D: number of tubers per plant. One-way ANOVA was used for significance analysis, n = 10, with error bars ± SEM. p<0.1, p<0.001, p<0.0001. Detailed Implementation
[0020] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the description of the embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are only embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on the provided drawings without creative effort.
[0021] Example 1: StPDF2 Acquisition of genes Obtained from the potato database (http: / / spuddb.uga.edu / index.shtml) StPDF2 The CDS sequence (Soltu.DM.10G000400.1) is shown in SEQ ID NO.1.
[0022] Using Premier 5.0 according to StPDF2 Primers pRC26B-StPDF2-F (SEQ ID NO. 2) and pRC26B-StPDF2-R (SEQ ID NO. 3) were designed based on the CDS sequence of the gene and synthesized by Sangon Biotech (Shanghai) Co., Ltd. Using cDNA from tuber samples as templates, the following steps were performed: StPDF2 PCR amplification of the full-length CDS. Amplification conditions were: 98℃ pre-denaturation for 5 min; 98℃ denaturation for 10 s, 58℃ annealing for 10 s, 72℃ extension for 20 s, 35 cycles (denaturation-extension); 72℃ extension for 5 min, and storage at 4℃. The amplified product was gel-cleaved, purified, and ligated into the pRC26B vector (obtained by our laboratory by modifying plant binary expression vectors, mainly by introducing an enhanced CaMV 35S promoter, AtADH 5′-UTR, eGFP reporter gene, and multiple cloning site to form a vector suitable for plant gene overexpression analysis) digested with KpnI and BamHI. The vector was transformed into DH5α competent cells, and positive clones were screened by PCR. Single positive clones were sent to Sangon Biotech (Shanghai) Co., Ltd. for sequencing. Plasmids of positive clones were extracted and confirmed by sequencing. StPDF2 The CDS sequence was ligated onto the vector.
[0023] Table 1. Used for amplification StPDF2 Primers for gene CDS sequence
[0024] Example 2: StPDF2 Genetic transformation of gene overexpression vectors The pRC26B-StPDF2 plasmid was transformed into Agrobacterium strain GV3101. The Agrobacterium GV3101 transformed with pRC26B-StPDF2 plasmid was used to infect sterile potato chips (the selected potato variety was "E3 potato") with a thickness of about 0.2 cm. After infection, the chips were cultured on MS30 plates containing antibiotics until resistant buds differentiated on the chips. The resistant buds were cut off and cultured in MS medium containing antibiotics to obtain resistant buds that could root.
[0025] Example 3: Potato Cultivation Wild-type potatoes and the obtained StPDF2 All gene overexpression lines were preserved as sterile potato tissue culture seedlings. The tissue culture subculture method was as follows: the shoot tips and stem segments of potato tissue culture seedlings were cut and cultured separately in ordinary MS medium in a tissue culture room at 22℃. After 14 days of subculture, they were removed from MS medium, hydroponically cultured for two days, and then planted in plastic pots (approximately 24 cm in diameter).
[0026] Example 4: Detection of gene overexpression efficiency Leaves were collected from tissue culture seedlings of overexpression lines obtained through genetic transformation and wild-type lines. Genomic DNA and total RNA were extracted from the samples, and the total RNA was reverse transcribed into cDNA. Based on the cloned... StPDF2 The gene sequence was used to design qRT-PCR primers StPDF2-qF (SEQ ID NO. 4) and StPDF2-qR (SEQ ID NO. 5), as shown in Table 2. The specificity of the primers was ensured using the BLASTn test in NCBI. PCR was performed on the genomic DNA using the overexpression fragment primers pRC26B-StPDF2-F and pRC26B-StPDF2-R to detect whether the target fragment was inserted. StPDF2 In the genome of gene overexpression lines, the results are as follows: Figure 1 As shown, the OE-PDF2-14 / 15 / 16 / 19 / 20 / 21 / 23 / 27 / 34 / 35 lines are transgenic positive lines. Using potato... StEF1a The gene was used as an internal reference gene, and the reverse-transcribed cDNA was used as a template for real-time quantitative PCR. 2 -ΔCt Method Analysis StPDF2 Gene overexpression efficiency, results as follows Figure 2 As shown, the expression level of StPDF2 was significantly upregulated by 10 to 60 times in four strains, namely OE-PDF2- / 19 / 20 / 27 / 35.
[0027] Table 2 StPDF2 qRT-PCR primers for gene sequences
[0028] Example 5: Overexpression StPDF2 Genes do not affect the above-ground growth of plants To clarify the effect of StPDF2 overexpression on potato plants, StPDF2 overexpressing lines and wild-type plants were planted in a long-day growth chamber (conditions set as 16 h light, 8 h darkness, and temperature 22 ± 2℃) for 28 days, and the aboveground phenotypes were investigated. The results are as follows: Figure 3 As shown: There were no significant differences in the aboveground parts of the StPDF2 overexpressing plants compared with wild-type plants.
[0029] Example 6: Overexpression StPDF2 Genes can cause premature potato tuber formation To clarify StPDF2 The function of genes in potato tuber formation was observed in wild-type plants and... StPDF2 The overexpression lines were grown under long-day conditions for 28 days, and then subjected to short-day induction to produce tuber phenotypes on days 3, 6, and 9 after tuber formation. The results are as follows: Figure 4 -A、 Figure 4 As shown in Figure -B, no tubers formed in any of the lines after 3 days of short-day induced tuber formation culture; however, on day 6, two lines showed tuber formation. StPDF2 In the overexpression lines, 50% of the plants had formed tubers, while the wild-type plants had not yet formed tubers; at day 9, 2 StPDF2 All plants in the overexpression lines formed tubers, while only 50% of the wild-type lines formed tubers. These results demonstrate that overexpression... StPDF2 Genes also cause potatoes to form tubers earlier.
[0030] Example 7: StPDF2 Gene overexpression leads to an increase in the number of tubers per plant. Furthermore, this invention examined wild-type plants and StPDF2 The number of stolons and tubers in overexpression lines after short-day induction were measured. Results are as follows: Figure 4 As shown, compared with wild-type plants, overexpression StPDF2 The gene significantly increases the number of stolons. Figure 4 -C) and the number of tubers ( Figure 4 -D).
Claims
1. A potato tuber formation-related transcription factor StPDF2 Genes are characterized by: The gene has the sequence number Soltu.DM.10G000400.1 in the potato database, and its base sequence is shown in SEQ ID NO.
1.
2. A device comprising the form described in claim 1 StPDF2 Gene overexpression vectors.
3. A device comprising the form described in claim 1 StPDF2 Plants with gene overexpression vectors.
4. A potato tuber formation-related transcription factor as described in claim 1 StPDF2 New applications of genes.