水稻短根白化突变体sra1生理生化分析及基因定位

doi:10.3724/SP.J.1006.2019.82041

摘要/Abstract

摘要：

叶色突变体是研究光合作用及叶绿素合成与降解途径的理想材料, 有助于了解高等植物叶绿体发育和光合作用的调控机制。利用甲基磺酸乙酯(EMS)处理西农1B, 获得一个短根白化突变体sra1 (short radicle and albino 1), 从出芽至第三叶期叶片始终为白色, 胚根较同时期野生型明显变短。对相同位置的叶片观察发现, 西农1B叶肉细胞叶绿体含量丰富且膜系统发育完整, 而sra1叶肉细胞液泡化严重, 叶绿体数目明显减少或没有, 基粒类囊体垛叠松散且稀少。生理生化分析发现sra1叶绿素a、叶绿素b和类胡萝卜素含量接近于零, 净光合速率为负值。遗传分析表明该短根白化表型由单个隐性核基因控制, 最终将SRA1定位于水稻第3染色体长臂InDel标记Z-20和Z-42间, 物理距离约657 kb, 是一个未报道的新基因。

关键词: 水稻, 短根白化突变体, 基因定位

Abstract:

The leaf color mutant is an ideal material for studying the process of photosynthesis and the pathways of chlorophyll synthesis and degradation. Studies on rice leaf color mutants are helpful to explain the gene network of chloroplast development and photosynthesis in higher plants. The mutant sra1, derived from the progeny of EMS-treated indica rice Xinong 1B. Leaves of sra1 were white in color from budding to the third leaf, and the radicle of sra1 was significantly shorter than that of wild type in the same stage. The observation of leaves in the same position showed that in Xinong 1B, the chloroplast of mesophyllous cells was abundant and the membrane system was fully developed, while in sra1, the vacuolarization of mesophyll cells was serious, the number of chloroplasts was significantly reduced or absent, and the granule thylakoids were loosely folded. The contents of chlorophyll a, chlorophyll b, and carotenoid in sra1 were close to zero, and the net photosynthetic rate was negative. Genetic analysis indicated that the short-root and albino phenotype was controlled by a single recessive nuclear gene, and SRA1 was localized between the long-arm InDel markers Z-20 and Z-42 of rice chromosome 3. The physical distance between the two InDel markers was about 657 kb. No genes related to chloroplast and root development have been reported in this interval, indicating that sra1 is a novel mutant. The sra1 is a novel mutant with albino and accompanying short roots, suggesting that SRA1 may be involved in regulating chloroplast and root development.

Key words: rice, short radicle and albino mutant, gene mapping

张莉莎,米胜南,王玲,委刚,郑尧杰,周恺,尚丽娜,朱美丹,王楠. 水稻短根白化突变体sra1生理生化分析及基因定位[J]. 作物学报, 2019, 45(4): 556-567.

ZHANG Li-Sha,MI Sheng-Nan,WANG Ling,WEI Gang,ZHENG Yao-Jie,ZHOU Kai,SHANG Li-Na,ZHU Mei-Dan,WANG Nan. Physiological and biochemical analysis and gene mapping of a novel short radicle and albino mutant sra1 in rice[J]. Acta Agronomica Sinica, 2019, 45(4): 556-567.

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材料 Material	净光合速率 Net photosynthetic rate (μmol CO₂ m^-2 s^-1)	气孔导度 Stomatal conductance (mol H₂O m^-2 s^-1)	蒸腾速率 Transpiration rate (mol H₂O m^-2 s^-1)	胞间CO₂浓度 Intercellular CO₂concentration (μmol CO₂L^-1)
WT	11.77±0.1091	0.30±0.0019	3.29±0.0090	298.44±2.9901
sra1	-3.61±0.0680^**	0.18±0.0008^**	1.28±0.0070^**	424.52±5.7801^**

编号 Code	酶 Enzyme	基因 Gene	水稻 Rice
1	谷氨酰-tRNA合成酶 Glutamyl-tRNAsynthetase	GltX	AK099931
2	谷氨酰-tRNA还原酶 Glutamyl-tRNAreductase	HEMA	AK099393
3	谷氨酸-1-半醛转氨酶 Glutamate-1-semialdehyde aminotransferase	GSA	AK064826
4	5-氨基乙酰丙酸脱水酶 5-aminolevulinate dehydratase	HEMB	AK101836
5	羟甲基后胆色素原合酶 Hydroxymethylbilane synthase	HEMC	AK060914
6	尿卟啉原III合酶 Uroporphyrinogen III synthase	HEMD	AK107127
7	尿卟啉原III脱羧酶 Uroporphyrinogen III decarboxylase	HEME	AK070859
8	粪卟啉原III氧化酶 Copro-porphyrinogen III oxidase	HEMF	AK070391
9	原卟啉原氧化酶 Protoporphyrinogen IX oxidase	HEMG	AK108365
10	镁螯合酶D亚基 Mg chelatase D subunit	CHLD	AK072463
	镁螯合酶H亚基 Mg chelatase H subunit	CHLH	AK067323
	镁螯合酶I亚基 Mg chelatase I subunit	CHLI	AK060389
11	镁原卟啉IX甲基转移酶 Mg-protoporphyrin IX methyltransferase	CHLM	AK059151
12	镁原卟啉 IX单甲酯环化酶 Mg-protoporphyrinogen IX monomethylester cyclase	CHL27	AK069333
13	二乙烯还原酶 3,8-divinyl reductase protochlorophyll a-8-vinyl reductase	DVR	AK103940
14	原叶绿素酸酯氧化还原酶 Protochlorophyllide oxidoreductase	POR	AK068143
15	叶绿素合酶 Chlorophyll synthase	CHLG	AK068855
16	叶绿素a加氧酶1 Chlorophyllide a oxygenase	CAO1	AF284781/AK063367

标记 Marker	正向引物序列 Forward primer sequence (5′-3′)	反向引物序列 Reverse primer sequence (5′-3′)
Z-12	ACGCCGTCTCCTTGGTAATC	CTTACATTGGAAAGCGGAGG
Z-20	TTGTAGTGCGGGCAGTTCTC	GTACTGTTTGCCTCTGCTTGC
Z-42	CCTTAATCTTGTCGCACAAC	GCTTCTCTCCATCAACCATAT
Z-78	ACGCAGGAGAAGCTAGATAGG	AACCATCTTTTAGTGTCGGGT
Z-83	TACCCTTGGAGCACATATAGTTAA	TTGGTCACCAACATATCTTGAA
Z-112	GGCTCTGCTAGCGAAATACTAG	GCTCACCATACTCACCAATTACT
ZMD5-18	TCAGGTCTTACGACGGTATGG	GGACACACTAGAATCTACGCACG
ZMD5-44	GAAGCTATTAGCCGGGATCG	GCCAAGGCAAAGCTCTCTT
ZTQ43	TGCAGAGACAAGGAAGCGG	TCCTGATCGTTGAGCAGGC
RM8208	TGTAAATGCCTGAGTGCCTACCC	AGCTAAACCGCTAGGGCCTTCC
RM3400	GGGTGCACCTTTGTATCTGTGC	TGACAGAGGTAAAGCAGCAGTAGTCG
RM15177	TCCTGTGTTGGACGGAGTATGC	GCCTCAGAGGTTAGAAGACAGACAGC
RM405	TATGCTTTCTGTCAGCTTCC	CTGCTGTGAAAGAGTTGACG
RM18053	GAGACCAGAGGGAGACAAAGAGAGG	CTTAGGTCTCCCGACAGTCACG

基因 Gene	基因产物 Product	RAP位点 RAP_locus
OsPOLP	PolI-like DNA polymerase, plastidal DNA polymerase 1	Os08g0175300
FtsZ	Plastid division protein FtsZ1	Os04g0665400
OsRPOTP	RNA polymerase	Os06g0652000
RpoB	RNA polymerase beta subunit	CAA33986
V2	Virscent 2	Os03g0320900
V3	Virscent 3	Os06g0168600
HEMA1	Glutamyl-tRNA reductase 1	Os10g0502400
HEMD	Uroporphyrinogen-III synthase	Os03g0186100
CHLD	Magnesium chelatase subunit	Os03g0811100
CHLM	Mg-protoporphyrin IX methyltransferase	Os06g0132400
CAO1	Chlorophyll a oxygenase 1	Os10g0567400
PSY1	Phytoene Synthase 1	Os06g0729000
PSY2	Phytoene synthase 2	Os12g0626400
carb1R	Chlorophyll A/B binding protein 1R	Os09g0346500
carb2R	Chlorophyll A/B binding protein 2R	Os01g0600900
petA	petA, cytochrome f	CAA33961
petB	petB, cytochrome B6	CAA33977
psaA	psaA, PSI P700 apoprotein A1	CAA33996
psbA	psbA, PSII 32 kDa protein	CAA34007
RbcL	RbcL, Ribulose bisphosphate carboxylase large chain	CAA34004
RbcS	Rubisco small subunit	Os12g0274700

基因 Gene	正向引物 Forward sequence (5′-3′)	反向引物 Reverse sequence (5′-3′)
Actin	GACCCAGATCATGTTTGAGACCT	CAGTGTGGCTGACACCATCAC
OsPOLP	ACCGGTGCTTTCAGGCTTG	GCTGACTGATAATCACACG
FtsZ	AAAGGACATAACCTTGCAAG	AGTTTTCCTATTGAACCGTG
OsRPOTP	TCCTCATGTCGAGCAAGGAT	GAAAGAATGTCTGGACTTTG
RpoB	TATGGTCTAATTCCGAGCGGT	TATGGTCTAATTCCGAGCGG
V2	GAGGAGTTCCTCACGATGAT	AGCATCAATGATAGACTCC
V3	GTTAGATGCTTCACTACACAG	GTACCATTGCCAACATGGCAAC
HEMA1	CGCTATTTCTGATGCTATGGGT	TCTTGGGTGATGATTGTTTGG
HEMD	AGGGATGGAAGGCTGCTGGA	CAGTGGTCCTTGGAAGCTCTG
CHLD	GCTTGCAGAAAGCTACACAAGC	AGGCCGTGAGCTAAAGGAGA
CHLM	CCATCCATTGGTCTCCTTATGACA	GTAGCCTACTTACCATCAATGAGTC
CAO1	GACACCTTCATCTGGGCTTCAA	CGAGAGACATCCGGTAGAGC
PSY1	GTGACCGAGCTCAGCCAG	TTAATCGGAAGGCAGATGCT
PSY2	GGGCGTTGCGCATCTAGA	CAAGAAATCGTGCGAAATCA
carb1R	AGATGGGTTTAGTGCGACGAG	TTTGGGATCGAGGGAGTATTT
carb2R	TGTTCTCCATGTTCGGCTTCT	GCTACGGTCCCCACTTCACT
petA	TGAATGTGGGTGCTGTTCTTATTT	TCGGGCGGCGCTAAT
petB	TCCTCGGTTCAATACATAATGACCG	CGTGCAGGATCATCATTAGAACCA
psaA	GCGAGCAAATAAAACACCTTTC	GTACCAGCTTAACGTGGGGAG
psbA	CCCTCATTAGCAGATTCGTTTT	ATGATTGTATTCCAGGCAGAGC
RbcL	CTTGGCAGCATTCCGAGTAA	ACAACGGGCTCGATGTGATA
RbcS	TCCGCTGAGTTTTGGCTATTT	GGACTTGAGCCCTGGAAGG