GSM8246586: Ambystoma mexicanum, retina, rep1; Ambystoma mexicanum; RNA-Seq
Eyeballs were excised and preserved in ice-cold, oxygenated Hanks Balanced Salt Solution (HBSS). Using fine scissors, the cornea was removed with a circular incision along the ora serrata. The lens and vitreous body were then extracted as thoroughly as possible. The detachment of the retina from the scleral cup was performed using ophthalmic forceps, followed by the careful removal of any retinal pigment epithelium (RPE) adhering to the retina. The isolated retina was washed once with fresh HBSS and immediately frozen in liquid nitrogen. For subsequent dissociation, the samples were transported on dry ice. The dissociated retinal tissue was washed with phosphate-buffered saline (PBS) and cut into smaller pieces, followed by the addition of 100 μL of precooled 0.1 × Lysis Buffer, thorough mixing, and incubation on ice for 5 min. The sample was then washed with 1 mL of precooled wash buffer, and cell debris and larger tissues were removed using 40-μm MACS Smart Strainer filtration. The sample was then centrifuged at 4 °C and 500 ×g for 5 min, after which the supernatant without nuclear particles was discarded. The sample underwent three wash cycles with wash buffer. The pellet was resuspended on ice in cooled, diluted nuclei buffer. Nuclear concentration was determined using a Countess II FL automatic cell counter, ensuring the nuclear suspension concentration was within the appropriate range. The human retinal cells underwent library construction using a 10X Chromium Single-Cell 3' Library Kit v3 (PN-1000078) following the guidelines provided by the manufacturer.The other 24 retina cells underwent library construction using DNBelab C4.