CAJ | 학술논문

为验证甘蓝型油菜甘油-3-磷酸脱氢酶基因（G3PDH ）的功能，采用 PCR 的方法克隆BnG3PDH 557 bp的干扰靶序列，将其正向片段插入到中间载体pHurricane的NotI酶切位点、反向重复序列插入到XhoI酶切位点，构建ihpRNA载体反向重复框，BamHI、EcoRI酶切下反向重复框后组装到由种子特异性表达的napin启动子驱动的pCAMBIAl390植物表达载体上。结果表明：PCR扩增产物、限制性内切酶酶切产物与预期片段长度相符。其中，1．7 kb的片段包括正向干扰靶片段和部分AtFad2内含子序列，2．3 kb大小的片段是干扰反向重复框。种子特异性表达的BnG3PDH ihpRNA载体成功构建。
위험증감람형유채감유-3-린산탈경매기인（G3PDH ）적공능，채용 PCR 적방법극륭BnG3PDH 557 bp적간우파서렬，장기정향편단삽입도중간재체pHurricane적NotI매절위점、반향중복서렬삽입도XhoI매절위점，구건ihpRNA재체반향중복광，BamHI、EcoRI매절하반향중복광후조장도유충자특이성표체적napin계동자구동적pCAMBIAl390식물표체재체상。결과표명：PCR확증산물、한제성내절매매절산물여예기편단장도상부。기중，1．7 kb적편단포괄정향간우파편단화부분AtFad2내함자서렬，2．3 kb대소적편단시간우반향중복광。충자특이성표체적BnG3PDH ihpRNA재체성공구건。
A ihpRNA vector was constructed to study the function of BnG3PDH gene;A 557 bp target BnG3PDH sequence was amplified by PCR,and the positive fragment was transferred into the platform vector pHurricane in enzyme loci of XhoI,and the reverse complementary target DNA fragment was inserted into the enzyme loci of NotI to form the inverse repeat of ihpRNA vector;The inverse repeat fragment was cut off by BamHI、EcoRI and inserted into a modified vector pCAMBIAl390 which under the drive of a rapeseed seed-specific promoter napin.The PCR and restriction enzyme digestion verified the successful construction of the BnG3PDH ihpRNA vector.