Dear Dilki Adikari how are you doing today?

Well…extract genetic material from sperm is kind difficult sometimes. Even for DNA is common to have some problems. The genetic material in the sperm is really condensed and this maybe compromise the extraction. I believe that you are using 200ul instead of 200ml of semen sample. As you are using extraction kit, just make sure to follow the manufacture’s instruction carefully.

Another option to the kit, you can use TRIzol method… this can be more efficient to extract RNA from sperm, but even in TRIzol could yield lower RNA quantity from sperm, but I think not so small as you have been getting. You also can dilute the RNA pellet in small amount of Ultra pure water to concentrate your sample a little bit, just make sure to get enough for cDNA synthesis.

200ul of sperm may be too much for the reaction occur well. Try to use 100ul or 50ul… For instance, 10ul of semen depending on the species you have billions of spermatozoa. So, the enzymatic digestion may be not going well because there too many cells and lower quantity of reagent. Well… I’ll hope it helped. Good luck with your research.

Tlio Teruo Yoshinaga's response was very acceptable to me. If you don't take the essential precautions, sperm RNA extraction can be quite complicated and difficult. The Trizol technique is a good strategy, but you must ensure that your RNA is DNA-free. Additionally, for any extraction procedure, you must assure that the number of sperm cells used in your extraction protocol accordingly you have to use the lysis buffer. Indeed, a sample volume of 200 ul for extraction is excessive.

I've included a link to the protocol that I think you'll find useful.

Article An Efficient Approach for RNA Extraction from Boar Sperm and...

Good Luck for your Research!

Careful sample handling is essential for RNA work. I have good success with using liquid nitrogen and grinding in a mortal and pestle to disrupt the sample. Make sure you aren't over-loading the column if it's a column kit. Sometimes it's better to use a smaller amount of starting material. See what folks use for other types of sperm (human, mouse, etc.).