DEVELOPMENT OF EST-SSR MARKERS AND ANALYSIS OF GROWTH TRAIT IN SILVER CARP
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摘要: 研究采用Illumina高通量测序技术对鲢(Hypophthalmichthys molitrix)卵组织进行转录组测序分析, 筛选多态EST-SSR标记并与生长性状进行关联性分析。研究结果显示: (1)测序组装总Unigene 77129个, SSR共24458个, 占所测EST数据的31.7%。鲢包含SSR的EST序列主要有二碱基、三碱基、四碱基和五碱基组成。二碱基中, AC含量最丰富; (2)随机挑选175条EST序列设计SSR引物, 筛选出多态EST-SSR标记36个, Blast结果显示有10条已知基因功能; (3)全同胞家系检测结果显示有两个标记与鲢的生长性状相关。SCE26与体长和体重显著相关(P<0.05), SCE65与肥满度显著相关(P<0.05)。研究结果将为鲢的分子标记辅助育种的应用提供基础理论指导。Abstract: Silver carp is one of important breeding fish in China. It is necessary to study the growth traits markers to accelerate the molecular marker-assisted breeding techniques. The simple sequence repeat (SSR) from the transcriptome of egg in silver carp were characterized by high-throughput Illumina sequencing, and genotypes associated growth traits were analyzed. The main results are as follows: (1) 77129 unigenes were detected by assembly, 31.7% of 24458 ESTs contained repeat motifs of various types (dinucleotide, trinucleotide, tetranucleotide and pentanucleotide) and lengths with AC being the most abundant dinucleotides one. (2) 175 EST-SSR primers were randomly characterized and 36 polymorphic EST-SSR were developed in silver carp. The results of Blast indicated that 10 EST-SSR genes are known. (3) The results showed that two loci were significantly correlated with growth traits in full-sibling family in silver carp. One locus SCE26 was significantly (P<0.05) correlated with body length and body weight, and SCE65 was significantly (P<0.05) correlated with condition factor. This study will provide theoretical basis and technical methods for the development of the marker-assisted breeding system in silver carp.
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Keywords:
- Hypophthalmichthys molitrix /
- EST-SSR /
- Molecular marker /
- Family /
- Growth traits
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图 1 鲢转录组数据微卫星类型特征
Figure 1. Characterization of SSR from silver carp transcriptome data
表 1 鲢36对EST-SSR引物特征
Table 1 Characterization of 36 EST-SSR in silver carp
引物位点Locus 退火温度Tm 片段大小
Size (bp)核心重复类型
Motif引物序列Primer (5′－3′) 基因Gene SCE26 55 148 (GGA)8 CAATGAAGGTGCAAATATTCCTC
ACTCTCTGTCTTCAAAATGGCTGUnknown SCE29 55 178 (TTC)5 CATGGGGATATTTGTGACTTCTT
ACTACAGAGCAAGGACACCTCTGUnknown SCE41 50 185 (ATG)5 CATATATGAGCAGAGCTGAGGCT
AAGTGACGATGGTCTCTGTGAGTUncharacterized protein isoform X1 SCE42 60 168 (TCC)7 AGTACTTTTTGACTGCTCCTTCG
CGGAGATTTTCAGTGTACAGGACTranscription factor 3C polypeptide 2 SCE58 48 173 (TAT)5 AAAATGCACCTGAATCTTTCGTA
AACGTTGATGGTGATGATAATTTGUnknown SCE60 50 176 (CA)10 CTCATATCAGGGCAGCTACAATC
CGTATTAGCATCTCTTCCCGTCSugar phosphate exchanger 2 SCE65 50 157 (TG)10 TGAACTGGATCAGAAGACACTCA
GCAAACTGCAAAAATGATTCTGUnknown SCE77 50 165 (AT)8 GTGATCGCTCCCTAAAGTTAAGA
GGCCTCTATGATCAGCTTCTGTUnknown SCE78 60 386 (TGC)6 ATCTACGCGTCTGCCAGTATC
ACTTCACGTGATCTTTACGAACGSolute carrier family 12 member SCE79 50 158 (TG)11 GCAGCTGTTTTCATAGTTACACG
GGAATTTACAGAATCACACAGCCUnknown SCE84 50 350 (TCCCAT)4 GAAATCGTTTGATGCACCATCT
GGATGTATTCGAGCTTTCATCACUnknown SCE86 55 167 (AAAGA)4 TGGGATTTTGTAGTTCAGTGGAT
ACTTGCTGCTGCATTTGTAGTTTUnknown SCE91 50 170 (GCC)6 GCAGGGTGAAAACATTGATTC
AGCAGGGCAGATCGAACCAbelson tyrosine-protein kinase 2 SCE92 50 186 (CA)8 AACACAACGATCCAACAGAGAAT
GGGTCTATGGATTCTTCCTTGTCUnknown SCE94 50 175 (GCT)5 TCACAGGGATCTAGGTGTTTTTC
AAACCAGCAAAGGTTAAAAGGACUnknown SCE97 55 178 (GAA)5 GCATCCAGAGTCTGAAAGAGAAG
TTCTCTCCGTCTGTCTTTTTGGUnknown SCE101 52 172 (CCA)5 TTATCCAGTCCATGTCCCTCTAC
CTCTTTTAGTGCCCTTTCCACTCSerine/threonine-protein kinase X1 SCE102 55 182 (TAT)6 GTCCAGTGCCAACTGAGTAGTTT
ATCAAGTGGCACAACCAAGTATTUnknown SCE109 50 170 (CT)10 TGGTGATGGAGCTCATCTTCTAC
ACAGCTTTCAGGCTCTTTGAAGTUnknown SCE111 55 173 (GAG)5 AGCGTACGTGCTCTTCTACCAG
CTGCTCGTTGTCGTTGAGATUbiquitin carboxy1-terminal hydrolase SCE118 50 252 (TGA)8 CAGAGCTGGTTGTGTTTGTGTTA
CAGACAGATGTGTCAGAGGGAACBifunctional lysine-specific emethylase SCE128 55 175 (TTC)7 AACCCACAGTTGTCAAGATTGTT
TGACAAGTGAACTGTTGAAGTGCUnknown SCE132 50 158 (TTTC)5 CAAGTGTTTGTGACACTCATTCC
TGGTTTTTCATTCTTATCAACCTTCUnknown SCE133 50 171 (ATT)5 TGAATCTAAAGAGGGAATGATGC
CCAGTGTGCTGTTGTATAAGTCGUnknown SCE136 55 190 (GAAA)5 GTGATAGTTTACCACCCCCTGTT
ATGGCCTTATCATAGGCTTCTTTUnknown SCE142 48 173 (TGA)6 TCTCCTTATTCTGCTGTGGTTGT
ATATGTTCAGCCTTGAGATCGAAUnknown SCE145 48 168 (GAT)8 GGATAATCTGGGGGTGTTAAGTT
CTTCCATCGATGTCTCATTTCATUnknown SCE155 52 170 (CAT)5 ATACAGAAGAGCCAAAACCATCA
AGTTTGATCACCATTCCAAACACUnknown SCE161 55 280 (TCTGA)4 TCAAACCCTCGTGTGATTAATGT
CGTGTGCGAGTGTCTAATATCTGUnknown SCE162 50 171 (CAA)5 GCTCGACTTGTGCCTAATTATTG
AAAATGACAATGTTTGGTCTTGGUnknown SCE163 60 176 (GT)10 AGGACTGTTTGATTGTCTGATGG
ATGAGTGAGAGGTTGTTGAGCAGInterleukin-12 subunit alpha SCE167 50 167 (ATG)6 CACAGCATGAGCTGAAGTGTAAC
AGTTCAAACCGCACACATGTAAUnknown SCE170 55 170 (GAT)6 AATCAAGAAACAATGTGGCCTT
GGTTTGTCGTAGTGGTCTTTCAGZinc finger SCE171 50 175 (ACA)6 GAAGAACAAAAAGAGAATAGGAAAAGA
CTGAAATGAGCAGAATGGAAAATUnknown SCE174 55 163 (TGAT)5 GTTGAAGATCCAGCTCTTCTCTG
TTGTTAATTTGGGTTCACCAGTTUnknown SCE175 50 169 (TCC)6 TGTTATTTTCCTTCAGCTTCTGG
TAGAAGACACCAACAAGCCAAATUncharacterized protein C9orf85 
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表 2 144尾鲢测量指标
Table 2 Measurements from 144 silver carp individuals
项目Item 体长Body length
BL (cm)体重Body
weight BW (g)肥满度K 最小值Min 11.2 25.8 1.41 最大值Max 17 82.3 2.07 平均值Average 14.1 49.4 1.7
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表 3 鲢生长性状有关的EST-SSR多态位点信息
Table 3 Effects of EST-SSR polymorphisms on growth traits of silver carp
位点Locus 基因型Genotype 数目No. 基因型频率Genotype frequency 体长Body length (cm) 体重Body weigh (g) 肥满度K SCE26 A148-148 3 0.02 14.47±0.50ab 52.03±4.89ab 1.75±0.63 A148-154 98 0.68 14.21±0.51a 50.28±4.89a 1.74±0.64 A154-154 43 0.30 13.89±0.49b 47.17±4.98b 1.73±0.61 SCE65 A149-157 36 0.25 14.12±0.48 49.55±5.05 1.74±0.60 ab A157-159 48 0.33 14.20±0.52 49.89±5.10 1.73±0.56 ab A149-159 32 0.22 13.96±0.50 48.63±5.23 1.77±0.58a A159-159 28 0.19 14.18±0.44 49.21±4.79 1.71±0.62b 注: 平均值±标准误; a, b (P<0.05)Note: Mean±SD; a, b (P<0.05)
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