Identification of the R2R3-MYB Gene and Expression Analysis of Flavonoid Regulatory Genes in Blueberry

doi:10.13560/j.cnki.biotech.bull.1985.2025-0103

Abstract

Abstract:

Objective Elucidating the expression patterns of the R2R3-MYB transcription factor family in blueberry during flavonoid regulation, thereby providing a theoretical foundation for exploring the regulatory mechanisms underlying flavonoid biosynthesis in blueberry. Method Bioinformatics was used to analyze the characteristics of R2R3-MYB genes in blueberry, such as physicochemical properties and subcellular localization, and RT-qPCR was to detect the expression patterns of VcMYBs in different tissues and different fruit development stages. Result A total of 278 R2R3-MYB genes were identified in blueberry, with amino acid sequence lengths ranging from 160 to 970 aa, and most of them were located in the nucleus. The blueberry VcMYBs was divided into 23 subfamilies by phylogenetic tree analysis, and the genes belonging to the same subfamily had similar structural characteristics. Cis-acting element analysis showed that VcMYBs may be involved in biological stress, abiotic stress, hormone induction, plant growth and development, and light response. Fourteen genes (SG4, SG5, SG6 and SG7) that may be involved in the regulation of flavonoid synthesis were randomly selected for further analysis. RT-qPCR analysis showed that the most of the VcMYBs had high expressions in fruits. Among them, VcMYB91(SG6) and VcMYB56(SG5) genes were lowly expressed in the green fruit stage and highly expressed in the mature stage, suggesting that they may be involved in fruit coloring. However, VcMYB88(SG4) and VcMYB96(SG4) showed the opposite expression pattern, suggesting that they may inhibit fruit coloring. In addition, VcMYB236(SG7) and VcMYB44(SG7) genes may be involved in flavonoid synthesis. Conclusion The 278 blueberry R2R3-MYB genes were identified via genome-wide analysis. Functional dissection revealed that four genes (VcMYB56, VcMYB91, etc.) from the SG5/6 subgroups regulate anthocyanin biosynthesis, six genes (VcMYB88, VcMYB96, etc.) from the SG4/5 subgroups participate in proanthocyanidin synthesis, and two genes (VcMYB236, VcMYB44) from the SG7 subgroup may govern flavonol metabolism.

Key words: blueberry, R2R3-MYB, bioinformatics, tissue expression pattern, different fruit development stages, flavonoid synthesis

LIU Jia-li, SONG Jing-rong, ZHAO Wen-yu, ZHANG Xin-yuan, ZHAO Zi-yang, CAO Yi-bo, ZHANG Ling-yun. Identification of the R2R3-MYB Gene and Expression Analysis of Flavonoid Regulatory Genes in Blueberry[J]. Biotechnology Bulletin, 2025, 41(9): 124-138.

Figures/Tables 8

References 52

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基因 Gene	引物 Primer （5'-3'）	∆G/∆g （25 ℃）（kcal/mol）	∆H/∆h （kcal/mol）	3'∆G （kcal/mol）	Tm （℃）	GC （%）	Primer score	PCR score
VcMYB44	F： GACCAGACCTAAAACGAGGCAAC	-36.5/-35.0	-190.5/-181.6	-7.3	60.8	52.2	941	920
VcMYB44	R： GGCAATCTTCCTGCTATTAGTGACC	-37.9/-36.7	-201.5/-194.0	-7.1	60.4	48.0	963	920
VcMYB56	F： TCGGAATATCCTAACGACCAACCAC	-37.9/-37.2	-202.2/-194.5	-7.2	60.3	48.0	946	889
VcMYB56	R： GACTTTCTGCTTTCGGGCCTC	-34.0/-32.9	-173.0/-166.5	-7.7	60.8	57.1	856	889
VcMYB77	F： TTCTTCCGATCAGGACAAACCCTC	-36.9/-36.0	-193.2/-185.4	-7.3	60.9	50.0	940	877
VcMYB77	R： TGACACTGCCGTGGTAATACCC	-34.5/-34.0	-179.3/-171.5	-6.7	60.0	54.5	897	877
VcMYB88	F： TTGCCTTTCCCTCTCCTCTAATGC	-36.6/-35.9	-191.7/-184.0	-6.7	60.6	50.0	880	900
VcMYB88	R： TAGGAATCCCAAGACTTTCCGTTT	-34.5/-34.3	-177.7/-181.3	-6.8	60.5	41.7	920	900
VcMYB91	F： CCACCTGACCGACAAATGCAA	-33.3/-32.7	-168.9/-164.8	-7.3	60.6	52.4	930	926
VcMYB91	R： AACCGTGCATATCCCTTACGAC	-34.5/-33.2	-177.3/-172.0	-7.5	60.5	50.0	941	926
VcMYB96	F： CCTTATCAAAACCAATCTGAACCGT	-37.0/-35.6	-194.0/-192.1	-8.0	60.9	40.0	839	867
VcMYB96	R： TTTGCTACTGCCACTATTCCCAC	-35.1/-34.2	-182.7/-177.3	-7.6	60.3	47.8	901	867
VcMYB103	F： TCACGCGCTGAATCCTAACCC	-34.2/-33.5	-174.1/-166.6	-7.1	61.0	57.1	936	883
VcMYB103	R： AACACTGCCGTGGTAATACCC	-32.9/-32.0	-167.4/-162.7	-6.7	60.1	52.4	889	883
VcMYB139	F： GTTGAGAGATCCCTCCGACCA	-32.9/-32.1	-167.1/-161.7	-7.1	60.3	57.1	920	888
VcMYB139	R： CTTTTCGGTCAGTAGGACTTCGTT	-36.0/-35.4	-187.2/-187.6	-7.1	60.8	45.8	933	888
VcMYB172	F： CCTCGCTCTCAAAATCCACGTC	-35.4/-34.2	-183.7/-175.9	-7.5	60.5	54.5	928	879
VcMYB172	R： TCTAGACAACTTGCAGCATCGG	-33.7/-33.4	-171.9/-172.6	-7.3	60.5	50.0	954	879
VcMYB214	F： GAAAGTCCATAACCCAAAGCCCAA	-36.1/-35.4	-188.9/-183.5	-7.1	60.6	45.8	918	893
VcMYB214	R： TCTTTAGGAGGGCAATCAAAGCTAC	-37.0/-36.1	-195.5/-191.3	-6.6	60.3	44.0	867	893
VcMYB229	F： TGAATTACCTCCGTCCGAACGTC	-36.3/-35.5	-191.1/-182.4	-7.5	60.3	52.2	920	793
VcMYB229	R： TCTCCCAGCTATCAGCGACCA	-32.8/-32.9	-166.6/-161.9	-7.1	60.2	57.1	919	793
VcMYB236	F： AGTTGTAATGCCGTCAACACCGA	-35.7/-35.5	-186.5/-180.7	-7.9	60.5	47.8	887	857
VcMYB236	R： ACTAGTGCCGTGATCATTTATTCCG	-38.1/-36.7	-201.0/-195.2	-7.4	61.0	44.0	904	857
VcMYB256	F： ACTACTGGAACACCCATATCCGAA	-35.4/-35.0	-183.1/-181.9	-7.4	60.8	45.8	937	861
VcMYB256	R： ATTAGCAGTCGCAAAAGATATGGT	-34.7/-34.0	-180.2/-183.3	-6.6	60.1	37.5	879	861
VcMYB259	F： TCCTTAATTGCGGGTAGACTTCCA	-35.1/-35.3	-182.2/-182.8	-6.7	60.3	45.8	887	866
VcMYB259	R： TCCTTTCAGTCGTATTCAAGGTCG	-36.1/-35.3	-189.0/-187.4	-7.9	60.4	45.8	866	866

基因 Gene	引物 Primer （5'-3'）	∆G/∆g （25 ℃）（kcal/mol）	∆H/∆h （kcal/mol）	3'∆G （kcal/mol）	Tm （℃）	GC （%）	Primer score	PCR score
VcMYB44	F： GACCAGACCTAAAACGAGGCAAC	-36.5/-35.0	-190.5/-181.6	-7.3	60.8	52.2	941	920
VcMYB44	R： GGCAATCTTCCTGCTATTAGTGACC	-37.9/-36.7	-201.5/-194.0	-7.1	60.4	48.0	963	920
VcMYB56	F： TCGGAATATCCTAACGACCAACCAC	-37.9/-37.2	-202.2/-194.5	-7.2	60.3	48.0	946	889
VcMYB56	R： GACTTTCTGCTTTCGGGCCTC	-34.0/-32.9	-173.0/-166.5	-7.7	60.8	57.1	856	889
VcMYB77	F： TTCTTCCGATCAGGACAAACCCTC	-36.9/-36.0	-193.2/-185.4	-7.3	60.9	50.0	940	877
VcMYB77	R： TGACACTGCCGTGGTAATACCC	-34.5/-34.0	-179.3/-171.5	-6.7	60.0	54.5	897	877
VcMYB88	F： TTGCCTTTCCCTCTCCTCTAATGC	-36.6/-35.9	-191.7/-184.0	-6.7	60.6	50.0	880	900
VcMYB88	R： TAGGAATCCCAAGACTTTCCGTTT	-34.5/-34.3	-177.7/-181.3	-6.8	60.5	41.7	920	900
VcMYB91	F： CCACCTGACCGACAAATGCAA	-33.3/-32.7	-168.9/-164.8	-7.3	60.6	52.4	930	926
VcMYB91	R： AACCGTGCATATCCCTTACGAC	-34.5/-33.2	-177.3/-172.0	-7.5	60.5	50.0	941	926
VcMYB96	F： CCTTATCAAAACCAATCTGAACCGT	-37.0/-35.6	-194.0/-192.1	-8.0	60.9	40.0	839	867
VcMYB96	R： TTTGCTACTGCCACTATTCCCAC	-35.1/-34.2	-182.7/-177.3	-7.6	60.3	47.8	901	867
VcMYB103	F： TCACGCGCTGAATCCTAACCC	-34.2/-33.5	-174.1/-166.6	-7.1	61.0	57.1	936	883
VcMYB103	R： AACACTGCCGTGGTAATACCC	-32.9/-32.0	-167.4/-162.7	-6.7	60.1	52.4	889	883
VcMYB139	F： GTTGAGAGATCCCTCCGACCA	-32.9/-32.1	-167.1/-161.7	-7.1	60.3	57.1	920	888
VcMYB139	R： CTTTTCGGTCAGTAGGACTTCGTT	-36.0/-35.4	-187.2/-187.6	-7.1	60.8	45.8	933	888
VcMYB172	F： CCTCGCTCTCAAAATCCACGTC	-35.4/-34.2	-183.7/-175.9	-7.5	60.5	54.5	928	879
VcMYB172	R： TCTAGACAACTTGCAGCATCGG	-33.7/-33.4	-171.9/-172.6	-7.3	60.5	50.0	954	879
VcMYB214	F： GAAAGTCCATAACCCAAAGCCCAA	-36.1/-35.4	-188.9/-183.5	-7.1	60.6	45.8	918	893
VcMYB214	R： TCTTTAGGAGGGCAATCAAAGCTAC	-37.0/-36.1	-195.5/-191.3	-6.6	60.3	44.0	867	893
VcMYB229	F： TGAATTACCTCCGTCCGAACGTC	-36.3/-35.5	-191.1/-182.4	-7.5	60.3	52.2	920	793
VcMYB229	R： TCTCCCAGCTATCAGCGACCA	-32.8/-32.9	-166.6/-161.9	-7.1	60.2	57.1	919	793
VcMYB236	F： AGTTGTAATGCCGTCAACACCGA	-35.7/-35.5	-186.5/-180.7	-7.9	60.5	47.8	887	857
VcMYB236	R： ACTAGTGCCGTGATCATTTATTCCG	-38.1/-36.7	-201.0/-195.2	-7.4	61.0	44.0	904	857
VcMYB256	F： ACTACTGGAACACCCATATCCGAA	-35.4/-35.0	-183.1/-181.9	-7.4	60.8	45.8	937	861
VcMYB256	R： ATTAGCAGTCGCAAAAGATATGGT	-34.7/-34.0	-180.2/-183.3	-6.6	60.1	37.5	879	861
VcMYB259	F： TCCTTAATTGCGGGTAGACTTCCA	-35.1/-35.3	-182.2/-182.8	-6.7	60.3	45.8	887	866
VcMYB259	R： TCCTTTCAGTCGTATTCAAGGTCG	-36.1/-35.3	-189.0/-187.4	-7.9	60.4	45.8	866	866