Acta Horticulturae Sinica ›› 2021, Vol. 48 ›› Issue (9): 1641-1652.doi: 10.16420/j.issn.0513-353x.2021-0580
• Research Papers • Next Articles
WANG Fusheng, LIU Xiaona, XU Yuanyuan, LIU Xiaofeng, ZHU Shiping, ZHAO Xiaochun*()
Received:
2021-06-16
Revised:
2021-08-27
Online:
2021-09-25
Published:
2021-09-30
Contact:
ZHAO Xiaochun
E-mail:zhaoxiaochun@cric.cn
CLC Number:
WANG Fusheng, LIU Xiaona, XU Yuanyuan, LIU Xiaofeng, ZHU Shiping, ZHAO Xiaochun. Cloning and Functional Analysis of Squalene Synthase Gene in Citrus[J]. Acta Horticulturae Sinica, 2021, 48(9): 1641-1652.
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URL: https://www.ahs.ac.cn/EN/10.16420/j.issn.0513-353x.2021-0580
引物名称 Primer | 序列(5′-3′) Sequence |
---|---|
SQS-GSP | F:CATCATGTTGCAACTGCTTTTCTG;R:GATTACGCCAAGCTTCCACATGCATCAGAGCATTGGTGAC |
gene-specific 5′UTR | TCTCTCCTCAAGTCCAATTCAGTCAGTAT |
3′ adapter-UAP | GGCCACGCGTCGACTAGTAC |
SQSRNAi | F:TCTAGAGCTCAGCAGGACTTGTAGGATTAGGT;R:GGATCCGAGCTCTGAGGGATAGCACAGAATCG |
Nos+SQS-sense | F :GATCATGAGCGGAGAATTAA;R:CCTCTTTCTACCTTCCCACA |
pGBiantisense | F:ACTTACTTACACTTGCCTTGGAGT;R:AAGGATCTGAGCTACACATGCTC |
N + P | F:CCTGATGCTCTTCGTCCAGA;R:GGATTGATGTGATAACATGGTGG |
SQS-qPCR | F:TCCAGAAGTTTCGCTCTCGTTA;R:TGGTGTCATCCTCAACAGTGTC |
123βAS | F:CAGCTGCGACTACAGCATTGAG;R:CTTGCGATGCTCAGACTCAGCT |
330βAS | F:ACTGCAATGAAGAGGGCTGCT;R:TCTCTGCGATGCTCAACTGTG |
490CAMS1 | F:ACTTCTAAAGGAGGGTGGACAT;R:TTCTGGTGGCAGCATTGAG |
195LUP2 | F:GGATGGGGATTACACGTTGAG;R:TCGAGCTCTTGCACAAGCGT |
632βAS | F:ATTACACGTTGAGGGTCACAGC;R:AGATGGTATGCTGGTTGCTCC |
789LUP2 | F:CGTTGAAGAGCTAGCTGAGGT;R:CTGAATTTGCCAAAGGAGGTC |
602CAS1 | F:TGCTTTGCTGTTGGGTGGAA;R:CGGCAAAAGCAGTGTCCCAT |
GAPDH | F:GGAAGGTCAAGATCGGAATCAA;R:CGTCCCTCTGCAAGATGACTCT |
Table 1 Primers for gene amplification and qRT-PCR analysis
引物名称 Primer | 序列(5′-3′) Sequence |
---|---|
SQS-GSP | F:CATCATGTTGCAACTGCTTTTCTG;R:GATTACGCCAAGCTTCCACATGCATCAGAGCATTGGTGAC |
gene-specific 5′UTR | TCTCTCCTCAAGTCCAATTCAGTCAGTAT |
3′ adapter-UAP | GGCCACGCGTCGACTAGTAC |
SQSRNAi | F:TCTAGAGCTCAGCAGGACTTGTAGGATTAGGT;R:GGATCCGAGCTCTGAGGGATAGCACAGAATCG |
Nos+SQS-sense | F :GATCATGAGCGGAGAATTAA;R:CCTCTTTCTACCTTCCCACA |
pGBiantisense | F:ACTTACTTACACTTGCCTTGGAGT;R:AAGGATCTGAGCTACACATGCTC |
N + P | F:CCTGATGCTCTTCGTCCAGA;R:GGATTGATGTGATAACATGGTGG |
SQS-qPCR | F:TCCAGAAGTTTCGCTCTCGTTA;R:TGGTGTCATCCTCAACAGTGTC |
123βAS | F:CAGCTGCGACTACAGCATTGAG;R:CTTGCGATGCTCAGACTCAGCT |
330βAS | F:ACTGCAATGAAGAGGGCTGCT;R:TCTCTGCGATGCTCAACTGTG |
490CAMS1 | F:ACTTCTAAAGGAGGGTGGACAT;R:TTCTGGTGGCAGCATTGAG |
195LUP2 | F:GGATGGGGATTACACGTTGAG;R:TCGAGCTCTTGCACAAGCGT |
632βAS | F:ATTACACGTTGAGGGTCACAGC;R:AGATGGTATGCTGGTTGCTCC |
789LUP2 | F:CGTTGAAGAGCTAGCTGAGGT;R:CTGAATTTGCCAAAGGAGGTC |
602CAS1 | F:TGCTTTGCTGTTGGGTGGAA;R:CGGCAAAAGCAGTGTCCCAT |
GAPDH | F:GGAAGGTCAAGATCGGAATCAA;R:CGTCCCTCTGCAAGATGACTCT |
Fig. 1 G/C nonsynonymous mutation at14th nucleotide acids of SQS CDS in eighteen citrus accessions G:G with underline is the 14th nucleotide acids in CDS.
Fig. 4 SQS expression level(A,B)and limonoids contents(C,D)in mature seeds and seeds at different developmental stages of pummelos MLZY:Citrus grandis‘Menglun Zao You’;GXMY:C. grandis‘Guanxi Mi You’;PSY:C. grandis‘Pingshan You’;GXSTY:C. grandis‘Guangxi Shatian You’;JLY:C. grandis‘Jinlan You’;MWY:C. grandis‘Meiwan You’;BP:C. grandis‘Burma Pummelo’.
Fig. 6 qRT-PCR results of relative expression level of OSC genes in leaf of SQS RNAi transgenic plants C-:Nontransgenic plants;SiN-1-SiN-4:SQS RNAi transgenic plants.
植株Line | 柠檬苦素Limonin | 诺米林Nomilin | 类柠檬苦素Limonoids |
---|---|---|---|
SiN-1 | 120.99 ± 1.92 c | 235.93 ± 3.95 e | 356.92 ± 5.87 d |
SiN-2 | 80.18 ± 3.18 d | 279.61 ± 9.56 d | 359.79 ± 12.74 d |
SiN-3 | 162.78 ± 5.80 b | 449.87 ± 2.98 a | 611.78 ± 8.78 a |
SiN-4 | 71.51 ± 0.88 d | 398.45 ± 2.81 b | 469.96 ± 3.69 c |
C-(非转基因植株Nontransgenic plants) | 199.57 ± 10.25 a | 294.50 ± 13.72 c | 494.07 ± 1.00 b |
Table 2 Limonoids contents in leaf of transgenic plants SiN-1-SiN-4 mg · kg-1 FW
植株Line | 柠檬苦素Limonin | 诺米林Nomilin | 类柠檬苦素Limonoids |
---|---|---|---|
SiN-1 | 120.99 ± 1.92 c | 235.93 ± 3.95 e | 356.92 ± 5.87 d |
SiN-2 | 80.18 ± 3.18 d | 279.61 ± 9.56 d | 359.79 ± 12.74 d |
SiN-3 | 162.78 ± 5.80 b | 449.87 ± 2.98 a | 611.78 ± 8.78 a |
SiN-4 | 71.51 ± 0.88 d | 398.45 ± 2.81 b | 469.96 ± 3.69 c |
C-(非转基因植株Nontransgenic plants) | 199.57 ± 10.25 a | 294.50 ± 13.72 c | 494.07 ± 1.00 b |
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