CAJ | 학술논문

以枸杞果实为试验材料,利用RACE技术克隆枸杞酸性转化酶基因LbAI的全长cDNA序列,应用生物信息学软件分析LbAI预期编码蛋白质特征；采用气相色谱法和实时荧光定量PCR 技术,分析不同发育阶段枸杞果实及不同颜色成熟果实中可溶性糖含量及LbAI 在果实中的相对表达量。结果显示：LbAI 的cDNA 序列全长2252 bp,开放阅读框(ORF)长1920 bp,编码642个氨基酸,LbAI编码的蛋白质相对分子量和等电点(pI)分别为70600和5.9,GenBank 登录号为KM191309。进化树分析结果显示,枸杞与马铃薯和番茄的亲缘关系最近,相似性在85%以上。随着果实生长发育,枸杞果实中果糖和葡萄糖含量不断升高,而蔗糖呈现出降低趋势；不同颜色枸杞果实中糖含量差异较大,成熟红色和黄色果实中果糖和葡萄糖含量显著高于黑色果实,但蔗糖含量在黑色果实中最高,在红色果实中最低；LbAI相对表达与果实中葡萄糖含量变化基本一致。表明LbAI基因在枸杞果实糖积累过程中具有重要的作用。
이구기과실위시험재료,이용RACE기술극륭구기산성전화매기인LbAI적전장cDNA서렬,응용생물신식학연건분석LbAI예기편마단백질특정；채용기상색보법화실시형광정량PCR 기술,분석불동발육계단구기과실급불동안색성숙과실중가용성당함량급LbAI 재과실중적상대표체량。결과현시：LbAI 적cDNA 서렬전장2252 bp,개방열독광(ORF)장1920 bp,편마642개안기산,LbAI편마적단백질상대분자량화등전점(pI)분별위70600화5.9,GenBank 등록호위KM191309。진화수분석결과현시,구기여마령서화번가적친연관계최근,상사성재85%이상。수착과실생장발육,구기과실중과당화포도당함량불단승고,이자당정현출강저추세；불동안색구기과실중당함량차이교대,성숙홍색화황색과실중과당화포도당함량현저고우흑색과실,단자당함량재흑색과실중최고,재홍색과실중최저；LbAI상대표체여과실중포도당함량변화기본일치。표명LbAI기인재구기과실당적루과정중구유중요적작용。
The acid invertase gene ( LbAI) full-length cDNA sequence from the fruits of wolfberry ( Lycium barbarum L. ) was clone by rapid amplification of cDNA ends ( RACE) , the features of LbAI-encoded protein were analyzed by bioin-formatics software, and the patterns of gene expression and accumulation of soluble sugars at different fruits growth stages and differently-colored fruits of wolfberry were analyzed by real-time fluorescent quantitative PCR ( qRT-PCR ) and gas chromatography ( GC ) . The full length of LbAI cDNA sequence was 2 252 bp, containing a 1920 bp open reading frame ( ORF) which encoded 642 amino acids with a relative molecular mass of 70 600 and isoelectric point (pI) of 5. 9. The phylogenetic tree showed that LbAI had the closest genetic relationship with Solanum tuberosum and S. lycopersicum,with the similarities above 85%. The contents of glucose and fructose increased along with the fruit growth and development, while sucrose content declined. The contents of glucose and fructose in red and yellow fruits were significantly higher than those in black fruit at maturity. The content of sucrose in black fruit was the highest and in red fruit was the lowest. The expression level of LbAI was consistent with the glucose content in fruits. It was suggested that LbAI might play an important role in sugar accumulation of wolfberry fruits.