Functional Study on the Key Gene LaLBD37 Related to the Lily Bulblets Formation

Abstract

Oriental hybrid lilies, known for their vibrant colors, diverse flower shapes, and long blooming seasons, require annual bulb propagation in horticultural production. This necessity can lead to higher production costs and limit their use in landscaping. The LA hybrid lily 'Aladdin' has shown strong self-reproduction capabilities in optimal cultivation environments, producing numerous high-quality underground stem bulblets. This makes it a valuable model for studying bulblet formation in lilies under natural conditions. Through transcriptome data analysis of different developmental stages of 'Aladdin' bulblets, the LaLBD37 gene, linked to bulblet formation, was identified. Bioinformatics analysis, subcellular localization studies, and transcriptional activation activity tests were conducted to understand the characteristics of LaLBD37. By introducing the LaLBD37 gene into 'Sorbonne' aseptic seedlings via Agrobacterium-mediated transformation, resistant plants were obtained. Positive plants were identified through various methods such as GUS activity detection, PCR, and fluorescence quantitative PCR. Phenotypic changes in positive plants were observed, and various physiological indicators were measured to confirm the role of LaLBD37 in bulblet formation, including soluble sugar content, starch content, sucrose synthase activity, and endogenous hormone levels. The findings suggest that the LaLBD37 gene plays a significant role in promoting the development of lily bulblets, offering insights for enhancing the reproductive capacity of Oriental hybrid lilies and exploring the molecular mechanisms involved in lily bulb regeneration.

Keywords: LaLBD37; bulblets genesis; functional verification; homologous transformation; oriental hybrid lily.

Figure 1

Full-length cloning and sequence alignment of LaLBD37. (a) Full-length cloning and domain prediction of LaLBD37; (b) domain prediction of LaLBD37; (c) Multiple sequence alignments of the amino acids of LaLBD37.

Figure 2

Analysis of subcellular localization and transcriptional activation activity of LaLBD37. (a) Nuclear localization of LaLBD37; (b) The division of N- and C-terminal regions of LaLBD37; (c) Transcriptional activation assay. Scale bar, 20 µm.

Figure 3

Growth process of transgenic plants. (a) Screening culture process; (b) Germination process; (c) Growth process.

Figure 4

Validation of transgenic ‘Sorbonne’ seedlings. (a) GUS detection in ‘Sorbonne’ transgenic LaLBD37 resistant plants; (b) PCR detection of ‘Sorbonne’ resistant plants. WT: Untransformed ‘Sorbonne’ plants; EV: Empty vector-transformed ‘Sorbonne’ plants; W: Water; C: Carrier vector; (c) Quantitative analysis of LaLBD37 in ‘Sorbonne’ positive plants; each group of data was the mean of three repetitions; ck: empty vector-transformed ‘Sorbonne’ line; L1–L8: LaLBD37-transformed ‘Sorbonne’ lines; * represents significant difference (p < 0.05).

Figure 5

Phenotypic observation and scale count of transgenic ‘Sorbonne’. (a) Phenotypic observation of transformed empty vector ‘Sorbonne’ and transgenic ‘Sorbonne’. ck: empty vector-transformed ‘Sorbonne’; L1, L2, and L4: lines with high expression of LaLBD37 in ‘Sorbonne’; (b) Number of scales for the transforming empty vector ‘Sorbonne’ and transgenic ‘Sorbonne’. ck: empty vector-transformed ‘Sorbonne’ line; L: LaLBD37-transformed ‘Sorbonne’ lines; Different letters represent significant differences among the three groups.

Figure 6

Phenotypic observation and scale count of transgenic ‘Sorbonne’. (a) The change of soluble sugar content in LaLBD37 transgenic lines; (b) The change of starch content in LaLBD37 transgenic lines; (c) The change of sucrose synthase activity in LaLBD37 transgenic lines; each group was the average of three repetitions; ck: empty vector-transformed ‘Sorbonne’; L1, L2, and L4: lines with high expression of LaLBD37 in ‘Sorbonne’; * representing significant difference (p < 0.05); ** represents significant difference (p < 0.01).

Figure 7

Determination of endogenous hormone content in LaLBD37 overexpression lines at the same time. (a) The IAA content in LaLBD37 transgenic lines; (b) The GA3 content in LaLBD37 transgenic lines; (c) The ABA content in LaLBD37 transgenic lines; each group was the average of three repetitions; ck: empty vector-transformed ‘Sorbonne’; L1, L2, and L4: lines with high expression of LaLBD37 in ‘Sorbonne’; ** represents significant difference (p < 0.01).