本研究發展具有內增幅控制（IAC）的多套式聚合酵素鏈鎖反應用以同時偵測三種帶有致病基囚的弧菌（創傷弧菌的vvhA與vvp，腸炎弧菌的tdh，霍亂弧茵的ctx）。在本實驗中IAC製作為一個嵌合式核酸，綠色螢光基因（grp）兩端為增幅vvhA引子序列，在這些弧菌中，增幅片段大小為:IAC（753 bp），vvhA（505 bp），vvp（383 bp），tdh（256 bp）和ctx（167 ...bp）我們使用了28株創傷弧菌、18株腸炎弧菌、14株霍亂弧菌和40株其他弧菌與非弧菌菌株分析中證明本方法的準確性，Kappa index為0.96，在牡蠣均質樣品中，經過8小時的增菌後，可以偵測低至101 CFU/mL添加於樣品中的弧菌。本方法所應用新製作的IAC和新設計的ctx引子，可以用來快速偵測海產中常見的致病性弧菌, This study developed a multiplex-polymerase chain react ion (m-PCR) method that targets the toxin genes, vvhA and vvp for Vibrio vulnificus, tdh for V. parahaemolvticus, ctx for V. cholera, to detect these three species simultaneously. A chimeric DNA consisting of a fragment of the green fluorescent protein gene (gfp) flanked by sequences of the vvhA primers was used as the internal amplification control (IAC). In the presence of these three Vibrio species, amplicons of IAC (753 bp), vvhA (505 bp), vvp (383 bp), tdh (256 bp) and ctx (167 bp) could be simultaneously detected. The accuracy of the m-PCR approach was evaluated with a Kappa index of 0.96 using 2
Kininogens, the precursors of bradykinins, ubiquitously exist in vertebrates, including mammals, birds, amphibians, and fishes. To elucidate the phylogeny of kininogen genes in early vertebrates, we ...cloned the full-length cDNA of kininogen gene from the liver of Lampetra japonica. The open reading frame of this sequence contained 546 bp and encoded 181 amino acids, including a cystatin domain without the canonical binding site for cysteine proteinases and a bradykinin domain. Our results suggested that in lampreys and most of other vertebrates, there might be only one kininogen gene, which was fused by certain sequences during vertebrate evolution and encoded proteins with more functions; however, another special kininogen gene, only encoding the bradykinin domain with multiple copies in some species, arose only in amphibians for adapting themselves to the unique environment. Using reverse transcription PCR, kininogen mRNA was also detected in lamprey gut, kidney, and leukocyte, but absent in lamprey buccal g
