Isolation and identification of starfish gonad-stimulating substance.

对海星促性腺物质的检测程序进行了重新考察，建立了两套程序(I和II)。I-1。神经采集，I-2。GSS提取(酸性乙醇均质)，I-3。Delipid(用氯仿和乙醚分配)，I-4。除盐(用Poragel PN在Proh逐步洗脱中进行分配层析)，I-5。凝胶渗透色谱(Sephacryl S-200型)，I-6。等电势，I-7。正相分配层析(Lmicrosorb Diol Gel in Proh梯度洗脱)，I-8。反相分配层析(ODS凝胶在乙腈梯度洗脱中)，和II-1，2，3。同第一系列，II-4。脱盐和凝胶渗透色谱(Sephadex G-50)，II-5。离子交换层析(ES-50ZN凝胶)，II-6。反相分配层析(乙腈梯度洗脱的ODS凝胶)，II-8。二次反相分配层析(ODS凝胶等梯度洗脱：0.1%三氟乙酸-10%PROH-18%丙酮醇)。通过第一系列分离得到Asterias amurens的GSS，并分析了部分氨基酸序列(N端的AEKYVGMGSYMALVGR)。根据该序列合成了一种多肽。虽然合成的多肽既没有GSS活性，也没有对GSS的抑制活性，但它的抗血清与Asterias和Asterinas的天然GSS相互作用。目前尚未获得阳性DNA片段。上述抗合成多肽抗体将用于检测可能的GSS DNA。

Procedure for starfish gonad-stimulating substance was reexamined and two series (I and II) of the procedure were established. I-1. nerve collection, I-2. GSS extraction (homogenization in acid ethanol), I-3. delipid (partition with chloroform and ether), I-4. desalting (partition chromatography with Poragel PN in PrOH step-wise elution), I-5. gel-permeation chromatography ( Sephacryl S-200), I-6. isoelectric forcusing electrophoresis, I-7. normal phase partition chromatography (Licrosorb diol gel in PrOH gradient elution), I-8. reverse phase partition chromatography (ODS gel in acetoniterile gradient elution), and II-1, 2, 3. the same as the first series, II-4. desalting and gel permiation chromatography (sephadex G-50), II-5. ion-exchange chromatography ( ES-50ZN gel), II-6. reverse phase partition chromatography (ODS gel in acetoniterile gradient elution), II-8. second reverse phase partition chromatography (ODS gel in isocratic elution: 0.1% trifluoroacetic acid-10% prOH-18% acetoniterile).Through the first series, GSS of Asterias amurensis was isolated and partial aminoacid sequence was analyzed (AEKYVGMGSYMALVGR from the N terminal). Based on the sequence a peptide was synthesized. Although the synthesized peptide neither showed the GSS activity nor inhibitory activity for GSS, its antiserum interacted with the native GSS of both Asterias and Asterina.DNA probes based on the aminoacid sequence of Asterias GSS were synthesized (sense and antisense). Positive DNA fragment have not been obtained yet. Above mentioned anti-synthesized peptide IgG will be used for detecting the possible GSS DNA.