采用RT-PCR和RACE(rapid amplification of cDNA ends)技术,从红花未成熟的种子和叶片中分离到8个FAD2基因和1个FAD6基因片段,并全部提交至GenBank上。在NCBI中Blast结果显示,CtFAD2-1和Ct-FAD2-8与向日葵、大豆、棉花等种子特异表达的F...采用RT-PCR和RACE(rapid amplification of cDNA ends)技术,从红花未成熟的种子和叶片中分离到8个FAD2基因和1个FAD6基因片段,并全部提交至GenBank上。在NCBI中Blast结果显示,CtFAD2-1和Ct-FAD2-8与向日葵、大豆、棉花等种子特异表达的FAD2-1基因的同源性较高。而CtFAD2-2与其他作物中组成型表达的FAD2-2/CtFAD2-3有较高的相似性。将红花ω6脂肪酸脱氢酶氨基酸序列进行同源性比对,CtFAD2-1与CtFAD2-8的同源性最高,为79.9%;其次是CtFAD2-2与CtFAD2-1和CtFAD2-8,分别为70.8%和70.2%;而这3个FAD2基因与其余5个拷贝间的序列相似性均较低,在52.0%～61.0%之间;CtFAD6与CtFAD2之间的相似性极低,在18.0%～21.9%之间。红花8个FAD2均检测到内质网滞留信号,将其定位于内质网膜上,而CtFAD6 N-端检测到67bp的质体信号肽序列,将其定位于叶绿体膜上。疏水性和跨膜分析结果显示,除Ct-FAD2-6以外,其余FAD2均含有6个疏水区,分别跨膜4～6次。展开更多
The full length cDNA encoding growth hormone of a freshwater fish, Lepomis cyanellus, (LcGH) was cloned from pituitary RNA with RT-PCR, 3′and 5′ RACE (rapid amplification of cDNA ends). The LcGH cDNA (Genbank No. AY...The full length cDNA encoding growth hormone of a freshwater fish, Lepomis cyanellus, (LcGH) was cloned from pituitary RNA with RT-PCR, 3′and 5′ RACE (rapid amplification of cDNA ends). The LcGH cDNA (Genbank No. AY530822), about 989nt (nucleotide) long, consisted of a open reading frame with 615nt long, 5′and 3′untranslated regions with 93nt and 224nt long respectively, and a 57nt poly (A) tail. The DNA sequence analysis showed that there are typical Kozak sequence and polyadenylation signal. The pregrowth hormone peptide of 204aa deduced from LcGH cDNA included a putative signal peptide (17aa) locating in its N-terminal. There exist a Asn-Cys-Thr glycosylation site at amino acid 201, and 4 cysteine residues (No. 69, 177, 194, 202) that are essential to construct two S-S bonds in this pregrowth hormone peptide. Homological comparision among LcGH and other species growth hormones showed that There is high homology (more than 85%) between growth hormone of Lepomis cyanellus and that of most perciformes fish, but low homology (less than 70%) in comparison with other species such as Siluriformes and Cypriniformes fish.展开更多
文摘The full length cDNA encoding growth hormone of a freshwater fish, Lepomis cyanellus, (LcGH) was cloned from pituitary RNA with RT-PCR, 3′and 5′ RACE (rapid amplification of cDNA ends). The LcGH cDNA (Genbank No. AY530822), about 989nt (nucleotide) long, consisted of a open reading frame with 615nt long, 5′and 3′untranslated regions with 93nt and 224nt long respectively, and a 57nt poly (A) tail. The DNA sequence analysis showed that there are typical Kozak sequence and polyadenylation signal. The pregrowth hormone peptide of 204aa deduced from LcGH cDNA included a putative signal peptide (17aa) locating in its N-terminal. There exist a Asn-Cys-Thr glycosylation site at amino acid 201, and 4 cysteine residues (No. 69, 177, 194, 202) that are essential to construct two S-S bonds in this pregrowth hormone peptide. Homological comparision among LcGH and other species growth hormones showed that There is high homology (more than 85%) between growth hormone of Lepomis cyanellus and that of most perciformes fish, but low homology (less than 70%) in comparison with other species such as Siluriformes and Cypriniformes fish.
