Identification and Expression Analysis of bHLH Transcription Gene Family in Lonicera macranthoides

doi:10.13560/j.cnki.biotech.bull.1985.2024-1105

Abstract

Abstract:

Objective To study the structure and function of bHLH transcription factor family members in Lonicera macranthoides, and to analyze their expression characteristics, so as to lay a foundation for further study on the function of bHLH gene family in flower development of L. macranthoides. Method Based on the flower transcriptome data of two varieties of L. macranthoides, the members of bHLH gene family were screened, and bioinformatics analysis was carried out by online tools to obtain the genes with significant expression differences between the two varieties of L. macranthoides. The expression of bHLH was verified by RT-qPCR. Result A total of 55 bHLH transcription factor family members were identified. Phylogenetic tree analysis showed that the bHLH protein of L. macranthoides was highly conserved with Arabidopsis thaliana, and the functions of some bHLH proteins were predicted based on the phylogenetic tree. Expression analysis showed that there were significant differences in bHLH genes between the two varieties of L.macranthoides. LmbHLH4/21/35 was highly expressed in the ‘Changgui’, and LmbHLH25 was highly expressed in the flowering stage 5-7 of ‘Xianglei’. The results of RT-qPCR showed that the relative expression was basically consistent with the transcriptome sequencing data, which confirmed the reliability of the transcriptome sequencing results. Conclusion LmbHLH4/21/15/25/35 may be a key transcription factor involved in the regulation of flower growth and development of L. macranthoides, which provides a reference for analyzing the molecular mechanism of flower difference phenotype of two varieties of Lonicera macranthoides.

Key words: Lonicera macranthoides, bHLH gene family, bioinformatics, expression analysis

QU Mei-ling, ZHOU Si-min, ZHANG Jing-yu, HE Jia-wei, ZHU Jia-yuan, LIU Xiao-rong, TONG Qiao-zhen, ZHOU Ri-bao, LIU Xiang-dan. Identification and Expression Analysis of bHLH Transcription Gene Family in Lonicera macranthoides[J]. Biotechnology Bulletin, 2025, 41(6): 256-268.

Figures/Tables 12

References 48

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引物名称 Primer name	上游引物 Forward primer（5'-3'）	下游引物 Reverse primer（5'-3'）
LmbHLH4	ACGAATCGGTGGGTGTAAAG	CCCTTCTCCTCCTTTCAGATAAAT
LmbHLH14	CTACTCTGCGCACCTTACTTC	GCACTCCTCCTCAGTTCTTTC
LmbHLH19	GTAAGCCACTGGTGGATGTTAG	GGGTAAGAAGTTTGGAGGGATG
LmbHLH21	CCGCCTTCAATAATCCCTACTC	CCGCTTGTTGTCTGTGTTTC
LmbHLH31	TGTTGAGAGGACGCTTGATATG	GGAAGGATCCCAGGAGTAGTAA
LmbHLH34	GTGGCAACTTCAGCAACAAA	CCCATTCCGTCACCGATAAA
LmbHLH35	ACCTCAGCTGAAATGGAGAAG	TTCGTGGATGGGTAGCATAAC
LmbHLH39	GCTTCTGTGGATCCCATGTT	GTTCCTGTGGGCTGGTTATT
LmbHLH55	AGTCCATCCACCAGAGGTATTA	GAACAACGCCATGAGGAAGA
18S	CTTCGGGATCGGAGTAATGA	GCGGAGTCCTAGAAGCAACA

引物名称 Primer name	上游引物 Forward primer（5'-3'）	下游引物 Reverse primer（5'-3'）
LmbHLH4	ACGAATCGGTGGGTGTAAAG	CCCTTCTCCTCCTTTCAGATAAAT
LmbHLH14	CTACTCTGCGCACCTTACTTC	GCACTCCTCCTCAGTTCTTTC
LmbHLH19	GTAAGCCACTGGTGGATGTTAG	GGGTAAGAAGTTTGGAGGGATG
LmbHLH21	CCGCCTTCAATAATCCCTACTC	CCGCTTGTTGTCTGTGTTTC
LmbHLH31	TGTTGAGAGGACGCTTGATATG	GGAAGGATCCCAGGAGTAGTAA
LmbHLH34	GTGGCAACTTCAGCAACAAA	CCCATTCCGTCACCGATAAA
LmbHLH35	ACCTCAGCTGAAATGGAGAAG	TTCGTGGATGGGTAGCATAAC
LmbHLH39	GCTTCTGTGGATCCCATGTT	GTTCCTGTGGGCTGGTTATT
LmbHLH55	AGTCCATCCACCAGAGGTATTA	GAACAACGCCATGAGGAAGA
18S	CTTCGGGATCGGAGTAATGA	GCGGAGTCCTAGAAGCAACA

基因名称 Gene name	基因ID GeneID	氨基酸数Number of amino acids	分子量 Molecular weight	等电点 pI	不稳定蛋白 Unstable protein	亲水性 GRAVY	脂肪指数 Fat index	亚细胞定位 Subcellular location
LmbHLH1	TRINITY_DN43160_c3_g1	1 130	122 891.71	8.70	55.20	-0.151	90.38	细胞核Nucleus
LmbHLH2	TRINITY_DN45765_c0_g1	542	59 110.52	6.01	71.84	-0.685	56.86	细胞核Nucleus
LmbHLH3	TRINITY_DN44404_c4_g2	397	43 453.13	5.54	58.14	-0.512	63.12	细胞核Nucleus
LmbHLH4	TRINITY_DN45094_c5_g2	553	60 335.04	5.56	56.51	-0.647	61.57	细胞核Nucleus
LmbHLH5	TRINITY_DN47741_c4_g1	371	41 064.15	4.54	51.80	-0.649	65.98	细胞核Nucleus
LmbHLH6	TRINITY_DN46241_c4_g1	461	50 993.36	5.51	74.35	-0.747	49.11	细胞核Nucleus
LmbHLH7	TRINITY_DN39815_c0_g1	490	54 256.93	6.04	50.07	-0.507	76.39	细胞核Nucleus
LmbHLH8	TRINITY_DN42054_c1_g1	523	57 182.91	5.65	53.37	-0.589	72.16	细胞核Nucleus
LmbHLH9	TRINITY_DN42054_c1_g2	513	56 244.54	4.92	54.06	-0.377	80.02	细胞核Nucleus
LmbHLH10	TRINITY_DN32727_c0_g1	474	53 002.24	7.08	48.26	-0.558	77.91	细胞核Nucleus
LmbHLH11	TRINITY_DN40879_c0_g1	439	49 418.43	5.92	44.35	-0.248	82.6	细胞核Nucleus
LmbHLH12	TRINITY_DN42270_c3_g1	324	36 465.68	7.12	46.27	-0.338	84.54	细胞核Nucleus
LmbHLH13	TRINITY_DN46971_c4_g1	300	33 743.57	8.50	48.42	-0.426	89.7	细胞核Nucleus
LmbHLH14	TRINITY_DN42056_c4_g1	255	28 220.89	7.67	56.87	-0.465	81.84	细胞核Nucleus
LmbHLH15	TRINITY_DN44414_c1_g7	238	26 955.26	9.32	49.17	-0.347	92.94	细胞核Nucleus
LmbHLH16	TRINITY_DN44386_c1_g2	661	72 485.68	5.35	48.40	-0.61	71.27	细胞核Nucleus
LmbHLH17	TRINITY_DN36574_c0_g1	365	40 232.17	5.56	57.74	-0.524	77.18	细胞核Nucleus
LmbHLH18	TRINITY_DN42789_c3_g1	417	47 601.89	4.90	43.74	-0.702	70.82	细胞核Nucleus
LmbHLH19	TRINITY_DN20377_c0_g1	501	55 602.27	6.86	37.66	-0.771	57.41	细胞核Nucleus
LmbHLH20	TRINITY_DN46178_c3_g2	306	34 854.75	5.66	42.40	-0.686	70.36	细胞核Nucleus
LmbHLH21	TRINITY_DN42621_c1_g5	401	44 732.06	8.69	67.88	-0.823	58.83	细胞核Nucleus
LmbHLH22	TRINITY_DN18450_c0_g2	245	28 168.84	4.78	65.71	-0.54	68.53	细胞核Nucleus
LmbHLH23	TRINITY_DN40373_c2_g1	238	26 199.63	8.43	59.20	-0.687	70.13	细胞核Nucleus
LmbHLH24	TRINITY_DN43239_c3_g1	328	36 679.23	5.48	69.01	-0.94	66.04	细胞核Nucleus
LmbHLH25	TRINITY_DN42843_c1_g4	90	10 584.84	7.96	86.02	-0.638	88.06	线粒体Mitochondria
LmbHLH26	TRINITY_DN45117_c2_g1	182	20 540.08	9.69	47.10	-0.267	106.54	细胞质Cytoplasm
LmbHLH27	TRINITY_DN45385_c1_g1	146	16 139.3	8.97	50.05	-0.446	74.18	细胞核Nucleus
LmbHLH28	TRINITY_DN41621_c2_g4	136	15 036.15	9.35	45.97	-0.511	82.43	细胞核Nucleus
LmbHLH29	TRINITY_DN40447_c0_g2	365	40 187.54	9.01	64.24	-0.938	55.89	细胞核Nucleus
LmbHLH30	TRINITY_DN40447_c0_g3	493	54 438.76	6.91	47.43	-0.758	66.92	细胞核Nucleus
LmbHLH31	TRINITY_DN44673_c3_g1	519	56 623.36	6.00	53.66	-0.64	62.95	细胞核Nucleus
LmbHLH32	TRINITY_DN43098_c2_g1	449	46 998.16	6.45	48.96	-0.458	67.19	细胞核Nucleus
LmbHLH33	TRINITY_DN46720_c1_g5	386	42 523	6.08	42.60	-0.451	76.01	细胞核Nucleus
LmbHLH34	TRINITY_DN46720_c1_g2	398	43 975.37	5.69	47.20	-0.668	66.18	细胞核Nucleus
LmbHLH35	TRINITY_DN47048_c1_g1	329	36 614.72	6.14	43.61	-0.75	70.24	细胞核Nucleus
LmbHLH36	TRINITY_DN47048_c1_g5	361	39 136.52	8.20	53.55	-0.658	62.99	细胞核Nucleus
LmbHLH37	TRINITY_DN42352_c4_g1	378	42 296.4	4.78	53.80	-0.839	64.29	细胞核Nucleus
LmbHLH38	TRINITY_DN35329_c0_g1	282	31 833.52	5.60	47.70	-0.649	60.5	细胞核Nucleus
LmbHLH39	TRINITY_DN44560_c7_g3	244	27 872.51	8.37	46.42	-0.946	62.75	细胞核Nucleus
LmbHLH40	TRINITY_DN45264_c3_g1	114	13 132.32	10.45	54.28	-0.275	83.77	叶绿体Chloroplasts
LmbHLH41	TRINITY_DN46281_c1_g1	256	28 313.2	8.91	54.05	-0.511	58.75	细胞核Nucleus
LmbHLH42	TRINITY_DN2359_c0_g1	250	28 089.1	7.86	54.39	-0.34	77.52	细胞核Nucleus
LmbHLH43	TRINITY_DN44645_c1_g1	212	24 266.36	6.04	60.17	-0.758	74.58	细胞核Nucleus
LmbHLH44	TRINITY_DN44510_c4_g1	325	36 719.95	5.52	68.76	-0.625	72	细胞核Nucleus
LmbHLH45	TRINITY_DN22004_c0_g1	303	34 475.81	5.75	69.49	-0.537	70.46	细胞核Nucleus
LmbHLH46	TRINITY_DN42286_c0_g1	307	34 581.25	6.61	84.31	-0.396	80.16	细胞核Nucleus
LmbHLH47	TRINITY_DN38245_c0_g2	286	32 549.83	8.48	66.05	-0.52	78.15	细胞核Nucleus
LmbHLH48	TRINITY_DN36940_c0_g1	218	24 902.56	9.56	54.37	-0.512	101.83	细胞质Cytoplasm
LmbHLH49	TRINITY_DN44940_c4_g1	256	29 471.48	9.36	61.70	-0.796	80.66	细胞核Nucleus
LmbHLH50	TRINITY_DN36667_c0_g1	179	20 359.5	9.03	51.14	-0.309	98.49	细胞核Nucleus
LmbHLH51	TRINITY_DN44808_c6_g1	180	20 647.17	9.49	36.82	-0.32	95.83	细胞核Nucleus
LmbHLH52	TRINITY_DN45353_c0_g1	279	30 516.07	5.32	54.07	-0.574	72.33	细胞核Nucleus
LmbHLH53	TRINITY_DN47214_c0_g2	441	49 404.11	5.79	43.81	-0.53	77.32	细胞核Nucleus
LmbHLH54	TRINITY_DN47214_c0_g1	532	58 538.46	5.78	41.63	-0.398	78.97	液泡膜Vacuole
LmbHLH55	TRINITY_DN45078_c1_g2	884	96 521.93	6.24	38.58	-0.355	81.26	细胞核Nucleus

基因名称 Gene name	基因ID GeneID	氨基酸数Number of amino acids	分子量 Molecular weight	等电点 pI	不稳定蛋白 Unstable protein	亲水性 GRAVY	脂肪指数 Fat index	亚细胞定位 Subcellular location
LmbHLH1	TRINITY_DN43160_c3_g1	1 130	122 891.71	8.70	55.20	-0.151	90.38	细胞核Nucleus
LmbHLH2	TRINITY_DN45765_c0_g1	542	59 110.52	6.01	71.84	-0.685	56.86	细胞核Nucleus
LmbHLH3	TRINITY_DN44404_c4_g2	397	43 453.13	5.54	58.14	-0.512	63.12	细胞核Nucleus
LmbHLH4	TRINITY_DN45094_c5_g2	553	60 335.04	5.56	56.51	-0.647	61.57	细胞核Nucleus
LmbHLH5	TRINITY_DN47741_c4_g1	371	41 064.15	4.54	51.80	-0.649	65.98	细胞核Nucleus
LmbHLH6	TRINITY_DN46241_c4_g1	461	50 993.36	5.51	74.35	-0.747	49.11	细胞核Nucleus
LmbHLH7	TRINITY_DN39815_c0_g1	490	54 256.93	6.04	50.07	-0.507	76.39	细胞核Nucleus
LmbHLH8	TRINITY_DN42054_c1_g1	523	57 182.91	5.65	53.37	-0.589	72.16	细胞核Nucleus
LmbHLH9	TRINITY_DN42054_c1_g2	513	56 244.54	4.92	54.06	-0.377	80.02	细胞核Nucleus
LmbHLH10	TRINITY_DN32727_c0_g1	474	53 002.24	7.08	48.26	-0.558	77.91	细胞核Nucleus
LmbHLH11	TRINITY_DN40879_c0_g1	439	49 418.43	5.92	44.35	-0.248	82.6	细胞核Nucleus
LmbHLH12	TRINITY_DN42270_c3_g1	324	36 465.68	7.12	46.27	-0.338	84.54	细胞核Nucleus
LmbHLH13	TRINITY_DN46971_c4_g1	300	33 743.57	8.50	48.42	-0.426	89.7	细胞核Nucleus
LmbHLH14	TRINITY_DN42056_c4_g1	255	28 220.89	7.67	56.87	-0.465	81.84	细胞核Nucleus
LmbHLH15	TRINITY_DN44414_c1_g7	238	26 955.26	9.32	49.17	-0.347	92.94	细胞核Nucleus
LmbHLH16	TRINITY_DN44386_c1_g2	661	72 485.68	5.35	48.40	-0.61	71.27	细胞核Nucleus
LmbHLH17	TRINITY_DN36574_c0_g1	365	40 232.17	5.56	57.74	-0.524	77.18	细胞核Nucleus
LmbHLH18	TRINITY_DN42789_c3_g1	417	47 601.89	4.90	43.74	-0.702	70.82	细胞核Nucleus
LmbHLH19	TRINITY_DN20377_c0_g1	501	55 602.27	6.86	37.66	-0.771	57.41	细胞核Nucleus
LmbHLH20	TRINITY_DN46178_c3_g2	306	34 854.75	5.66	42.40	-0.686	70.36	细胞核Nucleus
LmbHLH21	TRINITY_DN42621_c1_g5	401	44 732.06	8.69	67.88	-0.823	58.83	细胞核Nucleus
LmbHLH22	TRINITY_DN18450_c0_g2	245	28 168.84	4.78	65.71	-0.54	68.53	细胞核Nucleus
LmbHLH23	TRINITY_DN40373_c2_g1	238	26 199.63	8.43	59.20	-0.687	70.13	细胞核Nucleus
LmbHLH24	TRINITY_DN43239_c3_g1	328	36 679.23	5.48	69.01	-0.94	66.04	细胞核Nucleus
LmbHLH25	TRINITY_DN42843_c1_g4	90	10 584.84	7.96	86.02	-0.638	88.06	线粒体Mitochondria
LmbHLH26	TRINITY_DN45117_c2_g1	182	20 540.08	9.69	47.10	-0.267	106.54	细胞质Cytoplasm
LmbHLH27	TRINITY_DN45385_c1_g1	146	16 139.3	8.97	50.05	-0.446	74.18	细胞核Nucleus
LmbHLH28	TRINITY_DN41621_c2_g4	136	15 036.15	9.35	45.97	-0.511	82.43	细胞核Nucleus
LmbHLH29	TRINITY_DN40447_c0_g2	365	40 187.54	9.01	64.24	-0.938	55.89	细胞核Nucleus
LmbHLH30	TRINITY_DN40447_c0_g3	493	54 438.76	6.91	47.43	-0.758	66.92	细胞核Nucleus
LmbHLH31	TRINITY_DN44673_c3_g1	519	56 623.36	6.00	53.66	-0.64	62.95	细胞核Nucleus
LmbHLH32	TRINITY_DN43098_c2_g1	449	46 998.16	6.45	48.96	-0.458	67.19	细胞核Nucleus
LmbHLH33	TRINITY_DN46720_c1_g5	386	42 523	6.08	42.60	-0.451	76.01	细胞核Nucleus
LmbHLH34	TRINITY_DN46720_c1_g2	398	43 975.37	5.69	47.20	-0.668	66.18	细胞核Nucleus
LmbHLH35	TRINITY_DN47048_c1_g1	329	36 614.72	6.14	43.61	-0.75	70.24	细胞核Nucleus
LmbHLH36	TRINITY_DN47048_c1_g5	361	39 136.52	8.20	53.55	-0.658	62.99	细胞核Nucleus
LmbHLH37	TRINITY_DN42352_c4_g1	378	42 296.4	4.78	53.80	-0.839	64.29	细胞核Nucleus
LmbHLH38	TRINITY_DN35329_c0_g1	282	31 833.52	5.60	47.70	-0.649	60.5	细胞核Nucleus
LmbHLH39	TRINITY_DN44560_c7_g3	244	27 872.51	8.37	46.42	-0.946	62.75	细胞核Nucleus
LmbHLH40	TRINITY_DN45264_c3_g1	114	13 132.32	10.45	54.28	-0.275	83.77	叶绿体Chloroplasts
LmbHLH41	TRINITY_DN46281_c1_g1	256	28 313.2	8.91	54.05	-0.511	58.75	细胞核Nucleus
LmbHLH42	TRINITY_DN2359_c0_g1	250	28 089.1	7.86	54.39	-0.34	77.52	细胞核Nucleus
LmbHLH43	TRINITY_DN44645_c1_g1	212	24 266.36	6.04	60.17	-0.758	74.58	细胞核Nucleus
LmbHLH44	TRINITY_DN44510_c4_g1	325	36 719.95	5.52	68.76	-0.625	72	细胞核Nucleus
LmbHLH45	TRINITY_DN22004_c0_g1	303	34 475.81	5.75	69.49	-0.537	70.46	细胞核Nucleus
LmbHLH46	TRINITY_DN42286_c0_g1	307	34 581.25	6.61	84.31	-0.396	80.16	细胞核Nucleus
LmbHLH47	TRINITY_DN38245_c0_g2	286	32 549.83	8.48	66.05	-0.52	78.15	细胞核Nucleus
LmbHLH48	TRINITY_DN36940_c0_g1	218	24 902.56	9.56	54.37	-0.512	101.83	细胞质Cytoplasm
LmbHLH49	TRINITY_DN44940_c4_g1	256	29 471.48	9.36	61.70	-0.796	80.66	细胞核Nucleus
LmbHLH50	TRINITY_DN36667_c0_g1	179	20 359.5	9.03	51.14	-0.309	98.49	细胞核Nucleus
LmbHLH51	TRINITY_DN44808_c6_g1	180	20 647.17	9.49	36.82	-0.32	95.83	细胞核Nucleus
LmbHLH52	TRINITY_DN45353_c0_g1	279	30 516.07	5.32	54.07	-0.574	72.33	细胞核Nucleus
LmbHLH53	TRINITY_DN47214_c0_g2	441	49 404.11	5.79	43.81	-0.53	77.32	细胞核Nucleus
LmbHLH54	TRINITY_DN47214_c0_g1	532	58 538.46	5.78	41.63	-0.398	78.97	液泡膜Vacuole
LmbHLH55	TRINITY_DN45078_c1_g2	884	96 521.93	6.24	38.58	-0.355	81.26	细胞核Nucleus

蛋白 Protein	α-螺旋α-helix （%）	折叠延伸链 Turn loop （%）	无规则卷曲 Random coil （%）	蛋白 Protein	α-螺旋α-helix （%）	β-转角 β-sheet （%）	折叠延伸链 Turn loop （%）	无规则卷曲 Random coil （%）
LmbHLH1	46.02	1.86	52.12	LmbHLH29	16.99	0.00	2.47	80.55
LmbHLH2	13.84	0.74	85.42	LmbHLH30	20.69	0.00	1.42	77.89
LmbHLH3	18.14	0.76	81.11	LmbHLH31	13.68	0.00	2.12	84.20
LmbHLH4	14.29	0.72	84.99	LmbHLH32	23.39	0.00	1.56	75.06
LmbHLH5	19.95	2.96	77.09	LmbHLH33	23.83	0.00	1.30	74.87
LmbHLH6	14.75	3.04	82.21	LmbHLH34	21.86	0.00	1.26	76.88
LmbHLH7	30.41	8.98	60.61	LmbHLH35	25.53	0.00	3.04	71.43
LmbHLH8	22.56	5.93	71.51	LmbHLH36	22.71	0.00	3.05	74.24
LmbHLH9	17.54	7.02	75.44	LmbHLH37	17.20	0.00	2.12	80.69
LmbHLH10	30.38	8.44	61.18	LmbHLH38	30.85	0.00	1.77	67.38
LmbHLH11	30.30	10.93	58.77	LmbHLH39	29.10	0.00	2.05	68.85
LmbHLH12	35.80	5.25	58.95	LmbHLH40	39.47	0.00	9.65	50.88
LmbHLH13	44.67	6.67	48.67	LmbHLH41	28.91	0.00	1.17	69.92
LmbHLH14	38.43	9.41	52.16	LmbHLH42	31.20	0.00	0.00	68.80
LmbHLH15	44.12	7.98	47.90	LmbHLH43	31.13	0.00	0.47	68.40
LmbHLH16	27.23	8.17	64.60	LmbHLH44	32.62	0.00	6.46	60.92
LmbHLH17	29.86	5.21	64.93	LmbHLH45	34.98	0.00	7.92	57.10
LmbHLH18	32.13	6.24	61.63	LmbHLH46	35.18	0.00	6.51	58.31
LmbHLH19	13.97	4.19	81.84	LmbHLH47	34.27	0.00	7.34	58.39
LmbHLH20	28.76	1.96	69.28	LmbHLH48	41.28	0.00	10.55	48.17
LmbHLH21	21.95	0.75	77.31	LmbHLH49	35.94	0.00	8.98	55.08
LmbHLH22	32.65	0.82	66.53	LmbHLH50	50.28	0.00	11.17	38.55
LmbHLH23	40.34	0.00	59.66	LmbHLH51	49.44	0.00	11.67	38.89
LmbHLH24	25.00	2.44	72.56	LmbHLH52	22.94	0.00	5.73	71.33
LmbHLH25	62.37	1.08	36.56	LmbHLH53	32.65	0.00	3.63	63.72
LmbHLH26	48.35	9.34	42.31	LmbHLH54	29.14	0.00	4.89	65.98
LmbHLH27	29.45	8.22	62.33	LmbHLH55	22.85	0.00	5.88	71.27
LmbHLH28	31.62	9.56	58.82