Bioinformatic Analysis and Identification of Genes Contributing to Sugarcane Defoliation via BSR-Seq

doi:10.3969/j.issn.1000-2561.2021.10.013

Abstract

Abstract:

Sugarcane (Saccharam spp.), a C4 tropical grass mainly for sugar production, is commercially cultivated in over 100 countries. One of the main barriers to mechanical or manual harvesting of whole sugarcane stalk is the high percentage of impurities, mainly the debris of leaves and sheathes that results in increase of harvest time and cost. The most efficient solution for this issue is to develop sugarcane cultivars that are easy to defoliate in sugarcane breeding program. In order to investigate the genes responsible to sugarcane defoliation, BSR-Seq technology was employed in the current study to analyze pools of the leaves at +6 position of extremely hard-to-defoliate and easy-to-defoliate sugarcane plants derived from the cross of ‘ROC25’ × ‘Yunzhe 89-7’, together with the two parental lines with contrast defoliation phenotypes. A total of 60 Gb of clean reads were obtained and the Q30 values of the four samples were greater than 93% with GC content of 51%. Using R570 genome as reference, the alignment rate was greater than 41%. A total of 4085 differential expressed genes were detected in the 4 samples. The candidate regions were found to locate at 4 loci on chromosome 9, covering a total length of 1.40 Mb in which 86 genes were annotated. There were 15 non-synonymous mutations annotated between the parental lines, and two significant differentially expressed genes (Sh09_g020620, Sh09_g020080) were identified. The expression level of Sh09 g020080 in the easy-to-defoliate genotype was higher than that of the hard-to-defoliate genotype and consistent with the parents but that of Sh09_g020620 was opposite between the offspring pools and the parental lines, suggesting that only Sh09 g020080 may contributing to the phenotype of easy defoliation. Quantitation of Sh09 g020080 gene expression in leaves from position +1 to +7 from the extremely easy-to-defoliate representative accession 40-159 and the extremely hard-to-defoliate accession 5-94 confirmed the BSR-seq results of the two pools. In analysis of another genetic population derived from the cross of ‘B35-9’ × ‘CP08-1553’, Sh09 g020080 was also found to express in a higher level in the extremely easy-to-defoliate accession 16-226 than in the extremely hard-to-defoliate accession 16-224 in +4 to +6 leaves. Analysis of the deduced Sh09_g020080 protein revealed that this protein has four domains conserved in σ factor-like proteins that may function in in red, far red and blue light reactions and accumulation of chlorophyll. Overall, this study shed a new light into the endeavor of exploring essential genes for defoliation in sugarcane.

Key words: sugarcane, BSR-Seq, defoliation

CLC Number:

S566.1

YUE Qu, SHANG Heyang, ZHANG Pin, CHEN Baoshan, HUANG Youzong. Bioinformatic Analysis and Identification of Genes Contributing to Sugarcane Defoliation via BSR-Seq[J]. Chinese Journal of Tropical Crops, 2021, 42(10): 2841-2848.

Figures/Tables 7

Tab. 1 Statistical table of sequencing assembly and comparison

指标Index	T01	T02	T03	T04
有效片段	28 845 092	29 167 652	72 248 183	71 378 769
高质量碱基	8 643 893 628	8 745 375 900	21 666 853 410	21 402 859 902
Q30/%	93.67	93.80	93.97	94.10
GC/%	52.19	53.15	52.24	51.71
总片段	57 690 184	58 335 304	144 496 366	142 757 538
比对片段	23 954 010	23 495 340	63 055 434	61 878 087
比对率/%	41.52	40.28	43.64	43.34

Fig. 1 Clustering heat map of differential expression genes in samples T01 vs T02 and T03 vs T04

Fig. 2 Wayne map of differential expression genes and genes in candidate regions Both T01 vs T02 and T03 vs T04 represent differential gene sets, and CG (candidate gene) represents genes within candidate region.

Tab. 2 Primer sequences of candidate genes and internal reference genes

基因ID Unigene ID	正向引物（5'-3'）Former Primer (5'?3')	反向引物（5'-3'）Reverse Primer (5'?3')
Sh09_g020080	CGTTGGTCTGGTCTTTGGC	AATGGCGTGGACGGAATC
Sh09_g020620	CGTCGTCATCAACCACCCA	TGTTCACGTTCACGTTCTTGC
25s	ATAACCGCATCAGGTCTCCAAG3'	CCTCAGAGCCAATCCTTTTCC

Fig. 3 Relative expression of Sh09_g020620 and Sh09_g020080 in different leaf positions of difficult and easy defoliation lines A: Relative expression of Sh09_g020620 in delamination region of +1-+7 leaf position under condition of the control group; B: Relative expression of Sh09_g020080 in extracellular zone of leaf sheath of +1-+7 leaf position (BSR subpopulation) under condition of control group; C: Relative expression of Sh09_g020080 in isolated area of leaf sheath of +1-+7 leaf position under the condition of the control group (verification population).

Fig. 4 Sh09_g020080 of sugarcane and evolutionary trees of other species

Fig. 5 Tertiary structure model of Sh09_g020080 encoding protein

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