Homologous regions (hrs) are repetitive DNA sequences widely distributed in baculovirus genomes, and are known to function as transcriptional enhancers. In this study, we systematically characterized the seven hrs (hr1, hr2L, hr2R, hr3, hr4L, hr4R, and hr5) from a novel isolate of Bombyx mori nucleopolyhedrovirus (strain BmNPV-YT). The seven hrs range from 390 to 880 bp in length and consist of 1–7 repeat units of approximately 72 bp, each containing a 30 bp palindrome with an EcoRI site. Structural diversity among the hrs includes variable numbers of stem–loop motifs, major late transcription factor (MLTF) sites, cAMP response elements (CRE-like), and 12-O-tetradecanoylphorbol 13-acetate (TPA)-response elements (TRE-like). Functional assays using luciferase reporter plasmids revealed that all seven hrs function as potent transcriptional enhancers for both homologous and heterologous promoters in vitro and in vivo. In BmN cells and fifth-instar silkworm larvae, the hrs enhanced the homologous ie1 promoter 11–1258-fold and 43–1545-fold, respectively, with hr2L showing the strongest activity (counts per minute: 4.82 × 10⁸ in cells; 7.08 × 10⁷ in larvae) and hr4R the weakest. Similarly, when driving the heterologous AcMNPV ie1 promoter in Sf21 cells and larvae, enhancement ranged from 19- to 1023-fold and 13- to 449-fold, respectively, again with hr2L and hr3 showing maximal effects and hr4R minimal enhancement. Notably, all the hrs, except hr4R, also stimulated the homologous late p10 promoter, with activation levels of up to 37.7-fold in vitro and 79.2-fold in vivo, significantly above baseline. However, hr4R showed negligible or no enhancement (0.84–1.05-fold). Among all the active hrs, hr5 emerged as the most potent enhancer of the p10 promoter. Spearman’s correlation analyses demonstrated that enhancer efficiency for all three promoters (homologous ie1, heterologous ie1, and homologous p10) correlated significantly with the number of palindromic sequences, the combined count of CRE and TRE motifs, and the total number of characteristic sequences (r = 0.873–0.982, p < 0.05 or p < 0.01). Collectively, our findings clarify the structural features of hrs from the newly isolated strain BmNPV-YT, systematically identify their promoter preferences and host adaptability as transcriptional enhancers, and establish their structure–function relationships. These findings provide novel insights into the transcriptional regulation of baculoviruses and offer valuable tools for optimizing baculovirus expression vector systems.

This study aimed to establish an integrated variety–technology cultivation pathway for the new forage mulberry variety ‘Fengyuan No. 1’, linking salt tolerance mechanisms with practical application. A systematic investigation was conducted via a pot experiment with a 0–5‰ NaCl gradient and a field trial comparing three cultivation modes: Ridge Planting (RP), Furrow Planting (FP), and Flat-Bed Planting (FBP). Key findings are as follows. (1) The salt tolerance threshold was clearly defined: a 100% survival rate at salinity ≤ 4‰ (with no injury symptoms at ≤3‰), and 5‰ identified as the lethal threshold (33.33% survival). Salt stress triggered a resource reallocation strategy, increasing the leaf-to-stem fresh weight ratio from 1.53 (0‰) to 2.78 (5‰) to prioritize leaf photosynthetic function. Stable leaf circularity (0.83–0.87) indicated morphological stress resistance. (2) Optimized cultivation pathways were identified: FBP was the core pathway for maximizing biomass accumulation (root, stem, and leaf fresh weights were 5.0, 2.3, and 1.5 times those of RP, respectively) and resulted in the lowest leaf Na⁺ accumulation (124 mg/kg), making it suitable for lightly to moderately saline–alkali land (≤4‰). FP served as an effective pathway for salt avoidance and height promotion (plant height: 113.18 cm). RP constituted a specialized pathway for high-quality forage, yielding the highest crude protein content (23.3 g/100 g). (3) Cultivation modes significantly affected functional components; FBP activated alkaloid DNJ synthesis (215.16 mg/kg), whereas RP and FP increased osmolyte GABA accumulation (~586 mg/kg). In conclusion, this study integrates a complete technical pathway from salt tolerance mechanism analysis to diversified cultivation options, providing a systematic variety–technology solution for the industrial development of forage mulberry on coastal saline–alkali land.

The shortage of high-quality forage and the need for alternative protein sources are critical challenges for livestock production in northern China, where cold winters and saline–alkaline soils limit crop options. Mulberry (Morusalba L.) is a promising high-protein forage, yet few varieties have been bred for these conditions. Here, we report the breeding of ‘Fengyuan No. 1’, developed through systematic evaluation of 31 mulberry accessions for five forage traits, followed by a 4 × 3 factorial crossing design using four female parents (‘8036’, ‘Dahongpao’, ‘Jinqiang 63’, ‘Hongguo 2’) and three male parents (‘Nongsang 14’, ‘Yu 2’, ‘Yiyuan Huanglu’) selected based on comprehensive evaluation and complementary agronomic traits. Among the nine hybrid families with sufficient seed, family X01 (‘8036’ × ‘Nongsang 14’) ranked first in a three-year (2011–2013) comprehensive evaluation of growth traits using membership function analysis and was subsequently named ‘Fengyuan No. 1’. Multi-location (Shandong, Anhui, Shaanxi) field trials (2018–2021) with ‘Fengchi’ as the control showed that ‘Fengyuan No. 1’ achieved an average annual leaf yield of 12,596.48 kg DM·ha⁻¹ (+14.20%) and total branch–leaf biomass of 25,341.95 kg DM·ha⁻¹ (+13.36%). Leaf crude protein reached 30.87% on a dry matter basis (+25.64% vs. control). Silage prepared from this variety exhibited excellent fermentation quality (pH 4.10) and a relative feed value of 272.46, exceeding alfalfa silage standards. The variety maintained 100% survival at 0.4% NaCl and a 13.74% biomass advantage over the control on saline–alkaline field soils, with winter-desiccated shoot tips reduced by 53%. These results establish ‘Fengyuan No. 1’ as an elite forage mulberry for the Yellow River Basin and similar ecoregions.