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目的分析厚朴转录组中的SSR位点,并设计引物初步验证,对含SSR的序列进行功能分析,为厚朴分子辅助标记育种和资源保护提供了有利工具。方法将通过厚朴高通量转录组测序获得的Unigene序列进行SSR位点挖掘,利用Primer.03进行引物设计,随机挑选45对引物进行PCR,并利用Blast软件对含有SSR的Unigene进行功能分析。结果在16 369条厚朴转录组序列中共获得8 635个SSR位点,出现频率为52.75%。SSR序列中单核苷酸、二核苷酸和三核苷酸为优势重复类型,重复比例最高为10次,主导重复基元类型为A/T(47.16%)、AG/CT(31.74%)、AAG/CTT(6.53%)。45对引物中22对(48.89%)可以扩增出预期大小的条带。含SSR的Unigene与能量和氧化还原等代谢过程,以及RNA转运、剪接体和植物激素信号转导等通路有关。结论厚朴高通量转录组序列的SSR位点具有类型丰富、特异性强和潜能高等特点,同时SSR序列的功能分析将为厚朴基因挖掘和分子标记辅助育种提供有利的工具。
OBJECTIVE: To analyze the SSR locus in Magnolia officinalis transcriptome, and to design a primer for preliminary validation. The functional analysis of SSR-containing sequences provided a favorable tool for molecular marker-assisted breeding and resource conservation. Methods The Unigene sequence obtained from the Magnolia officinalis high-throughput transcriptome sequencing was used to mine the SSR loci. Primer.03 was used to design the primers. 45 pairs of primers were randomly selected for PCR. The functional analysis of Unigene containing SSR was performed using Blast software. Results A total of 8 635 SSR loci were found in 16 369 Magnoliaceta transcriptomes, with a frequency of 52.75%. Single nucleotide, dinucleotide and trinucleotide repeats were the most common repeat types in SSR sequence with the highest repeat ratio of 10 times. The dominant repeat motifs were A / T (47.16%), AG / CT (31.74%), , AAG / CTT (6.53%). Of the 45 pairs of primers, 22 (48.89%) of the primers amplified bands of the expected size. Unigene with SSR is involved in metabolic processes such as energy and redox, as well as pathways such as RNA transport, spliceosomes and plant hormone signaling. Conclusion The SSR loci of Magnolia officinalis high-throughput transcriptome sequences are rich in type, strong in specificity and high in potential, and the functional analysis of SSR sequences will provide useful tools for gene amplification and molecular marker-assisted breeding of Magnolia officinalis.