Study On Metaphase Conditions Exploration In Mitosis For Lymphocyte And Y Chromosome Microdissection Research

Cashmere goat is a variety that can be used for its Wool,meat and leather. It has high economic value. After long-term natural selection and artificial breeding of separated, this important livestock Growes in the specific ecological environment. Due to the current ecological environment deterioration, Cashmere goat growth condition gradually get worse. This condition led to the lack of quality breeding sheep and the decrease of quality of wool, so strengthing the quality of the Cashmere goat is imminent. Male Cashmere goat has played a very important role in reproduction and its wool production is high. So cultivating excellent Cashmere goat is imperative. But male breeding goat depend on the Y chromosome. The Y chromosome sequencing of cashmere goat hasn’t be reported in the world.To get high purity Y chromosome of cashmere goat is crucial. This study was conducted to obtain high purity Y chromosome successfully with a laser micro cutting system1. The explore of lymphocyte metaphase condition(1)Lymphocytes was cultured for48h without any stimulation. The proportion of G2/M phase was only0.06%by Flow cytometry.(2)Lymphocytes were stimulated to proliferate with PHA and ConA, The results indicate that the stimulation effect of ConA worked better than PHA,.The optimal concentration of PHA is60ug/ml and ConA is5ug/ml.(3) Colchicines is added at different culture time to make lymphocytes stalling in the G2/M phase, the results show that Colchicines is added at45h,thus, The best condition is52.8%by Flow cytometry(4)When the time of colchicine to treat lymphocytes prolonged to5h, The proportion G2/M phase reached to58.38%. The time of colchicines extend appropriatelywhich can make more cells focusing on G2/M phase2. Chromosome microdissectionChromosome karyotype is made after lymphocytes culturing.Laser Chromosome microdissection system is used to cut Y chromosome. Through the exploration of lymphocyte G2/M phase, lymphocyte were cultured for45h and used colchicine to treat5h. At this time, Chromosome karyotype was made with membrane slides, chromosome karyotype more than other condition, membrane slides was used in Chromosome microdissection. Chromosome karyotype were found under Zeiss microscope and determined the target Y chromosome. Then laser chromosome microdissection system was used for cutting the whole Y chromosome...