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Acta Agronomica Sinica ›› 2018, Vol. 44 ›› Issue (03): 376-384.doi: 10.3724/SP.J.1006.2018.00376

• CROP GENETICS & BREEDING · GERMPLASM RESOURCES · MOLECULAR GENETICS • Previous Articles     Next Articles

Cloning and Imprinting Characterization Analyses of Paternally Expressed Gene ZmVIL1 in Maize Endosperm

Chao-Xian LIU**, Jiu-Guang WANG**, Xiu-Peng MEI, Ting-Ting YU, Guo-Qiang WANG, Lian ZHOU, Yi-Lin CAI*()   

  1. Maize Research Institute, Southwest University / Engineering Research Center of South Upland Agriculture, Ministry of Education, Chongqing 400715, China
  • Received:2017-07-11 Accepted:2017-11-21 Online:2018-03-12 Published:2017-12-04
  • Contact: Chao-Xian LIU,Jiu-Guang WANG,Yi-Lin CAI E-mail:caiyilin1789@163.com
  • Supported by:
    This study was supported by China Postdoctoral Science Foundation (2014M552303), Fundamental Research Funds for the Central Universities (XDJK2015B009), and the Technology Integration and Demonstration of Zhongkeyu 9699, and Xidabainuo 1 (cstc2015jcsf-nycgzhA80006).

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Abstract:

Imprint genes play important roles in maize seed development. ZmVIL1 is a paternal-preferentially expressed gene. In this study, the full length of ZmVIL1 cDNA was cloned by RACE, which is about 2.2 kb and encodes a protein consisting of 598 amino acids. The analyses of allele expression of ZmVIL1 in endosperm and embryo at 14 days after pollination (DAP) of hybrids from reciprocal crosses between B73 and Mo17 indicated that ZmVIL1 was maternally imprinted in endosperm, while not in embryo. The allele expression was extensively evaluated in 14 DAP endosperm of hybrids form reciprocal crosses of Zheng 58 and Chang 7-2, Huang C and 178, PH6WC and PH4CV, and in 10-28 DAP endosperm of hybrids form reciprocal crosses of B73 and Mo17, suggesting that ZmVIL1 was gene-specific and binary imprinting and consistently imprinted in endosperm from 12 to 28 DAP. Expression pattern analyses demonstrated ZmVIL1 was constitutively expressed from vegetative to reproductive stages. ZmVIL1 was strongly expressed in seed at 10 DAP, immature tassels, immature ears and silk, and secondly expressed in ovule, embryo at 25 DAP and root, suggesting that ZmVIL1 plays important roles in maize seed and flower development.

Key words: ZmVIL1, gene cloning, gene imprinting, imprinting pattern

Table 1

Primer sequences for 5° RACE, 3° RACE, CDS amplification and imprinting characterization analyses of ZmVIL1"

引物名称
Primer name		 正向引物序列
Forward primer sequence (5°-3°)		 反向引物序列
Reverse primer sequence (5°-3°)		 内切酶
Restriction enzyme
ZmVIL1-5° race GCCCATCGCATTCCAACTGCCTTACTAC
ZmVIL1-5° race-nest-1 GCGTTTAGGTCGGGGACAGAAGATGAG
ZmVIL1-5° race-nest-2 GGTCAGGAGAACTCAACCACTCGTCAGC
ZmVIL1-3° race CTCGGCAAGTAAGCAAGGTGAATAGCGT
ZmVIL1-3° race-nest-1 GCAACTGGATGGGGATTATGAGCACTG
ZmVIL1-3° race-nest-2 AGGTAGTAAGGCAGTTGGAATGCGATGG
ZmVIL1-CDS ATGCCCAAAACACCTCCTGT CTCAGCGAACCACAACAACAAA
ZmVIL1-imprinting ATTCAAAGAGCTGACGAGTGGT TCATAATCCCCATCCAGTTGCC Hph I

Fig. 1

5° RACE and 3° RACE for ZmVIL1 cloning A, B, C: the gel electrophoresis for the first, second and third round RACE PCR products. L1, L3, L5: the gel electrophoresis for ZmVIL1 cDNA 5° RACE; L2, L4, L6: the gel electrophoresis for ZmVIL1 cDNA 3°RACE; Marker: D2000 plus."

Fig. 2

Protein sequences alignment for ZmVIL1 in maize, and VIN3, VIL1 in Arabidopsis"

Fig. 3

SNPs in ZmVIL1 CDS of B73 and Mo17 The arrows indicate the SNPs at the 1380th, 1450th, and 1530th base pairs in ZmVIL1 CDS of B73 and Mo17."

Fig. 4

Imprinting characterization analysis of ZmVIL1 in 14 DAP endosperm and embryo of B73 and Mo17 reciprocal hybrids A: imprinting characterization analysis of ZmVIL1 in 14 DAP endosperm; B: imprinting characterization analysis of ZmVIL1 in 14 DAP embryo. Marker: D2000 plus."

Fig. 5

Imprinting characterization analysis of ZmVIL1 in 14 DAP endosperm of Huang C and 178, Zheng 58 and Chang 7-2, PH6WC and PH4CV reciprocal hybrids H: Huang C; C: Chang 7-2; Z: Zheng 58; 4C: PH4CV; 6W: PH6WC; Marker: D2000 plus."

Fig. 6

Imprinting characterization analysis of ZmVIL1 in 10-28 DAP endosperm of B73 and Mo17 reciprocal hybrids A: imprinting characterization analysis of ZmVIL1 in 10-28 DAP endosperm of B73×Mo17; B: imprinting characterization analysis of ZmVIL1 in 10-28 DAP endosperm of Mo17×B73; Marker: D2000 plus."

Fig. 7

Expression pattern of ZmVIL1 in different maize tissues"

Fig. 8

Molecular evolution relationship analysis of ZmVIL1 with VIN3 protein family in Arabidopsis and rice"

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