Two gonadotropin releasing hormones and a method to isolate the same
AssigneeCouncil of Scientific and Industrial Research
InventorsAbhijit Chatterjee, Partha Ray, Subrata Dasgupta, Samir Hattacharya, Santosh Pasha
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Abstract
The present invention relates to two novel Gonadotropin releasing hormones muGnRH I and muGnRH II of amino acid SEQ ID 1 as QHWSAWRLPG, and SEQ ID 2 QHWSWGILPG respectively, useful for induced breeding in fish both in combination and alone, by activating production of Gonadotropin, and a method of isolating the same from Indian Murrel brain, and further, a method of inducing breeding in fishes using the said novel gonadotropin releasing hormones.
Claims7
A method of isolating a gonadotropin releasing hormone selected from the group consisting of muGnRH I (SEQ ID NO:1) and muGnRH II (SEQ ID NO:2) said method comprising the steps of:
- (a) homogenizing hypothalami under acidic conditions to obtain a homogenate;
- (b) filtering said homogenate to remove fat and cell debris;
- (c) adding acetone in a stepwise manner to the filtrate to obtain three precipitates AC I, AC II, and AC III;
- (d) estimating gonadotropin releasing hormone (GnRH) activity in each of the precipitates obtained in step (c);
- (e) fractionating active precipitate AC II using Molecular Exclusion Chromatography to obtain fractions SG I and SG II;
- (f) estimating gonadotropin releasing hormone (GnRH) activity in the fractions obtained in step (e);
- (g) fractionating fraction SG II using ion exchange chromatography using an anionic column to collect fractions MQ I, MQ II, MQ III, and MQ IV;
- (h) estimating gonadotropin releasing hormone (GnRH) activity in the fractions MQ I, MQ II, MQ III, and MQ IV obtained in step (g);
- (i) fractionating the fraction MQ I obtained in step (h) using ion exchange chromatography using a cationic column to obtain fractions MS I, MS II, MS III, and MS IV;
- (j) estimating gonadotropin releasing hormone (GnRH) activity in the fractions MS I, MS II, MS III, and MS IV obtained in step (i);
- (k) fractionating the MS II fraction using reverse phase chromatography to obtain fractions SRP I, SRP II, and SRP III;
- (l) estimating gonadotropin releasing hormone (GnRH) activity in the fractions obtained in step (k);
- (m) fractionating fraction MQ II obtained in step (h) using reverse phase chromatography to obtain fractions QRP I, QRP II, and QRP III;
- (n) calculating gonadotropin releasing hormone (GnRH) activity in the fractions obtained in step (m);
- and
- (o) identifying the QRP II and SRP II fractions of steps (m) and (k) respectively as muGnRH I and muGnRH II respectively.
Dependencies are inferred from the claim text.