Molecular genetic testing of char by hypervariable genome loci

The purpose of the work is to confirm the species affiliation of the char living in Lake Melkoe. Objectives of the study: catching charrs in the Lake Melkoe, taking biomaterial for PCR-test, determination of DNA fragments characteristic of the species. The catch of char was carried out on Lake Melkoe basin of the Pyasina River in September-October 2022 with fixed vein networks with a mesh of 30-55 mm, 30 m long and 2 m high at a depth of 9 to 14 m. The exposure time is 12 hours. Water temperature is 6-7 °C. The biomaterial (muscles of the ridge) in the amount of seven samples (in duplicate) was fixed in 95 % ethanol, the reaction was carried out in the laboratory of Krasnoyarskagroplem (joint-stock company). Before DNA isolation, muscle tissue was soaked in a buffer solution of tris hydrochloride. To isolate DNA from biomaterial samples, a set of reagents Ekstran-2 manufactured by NPF Syntol (limited liability company), Russia, was used. Microsatellite inverted primers (CTC) were used to obtain ISSR DNA fragments (CTC)6G - CTC-CTC-CTC-CTC-CTC-CTC-G; (AGC)6G - AGC-AGC-AGC-AGC-AGC-AGC-G; (GAG)6C - GAG-GAG-GAG-GAG-GAG-GAG-C; (ACC)6G - ACC-ACC-ACC-ACC-ACC-ACC-G. PCR was performed in the hot start mode - 3 min/95 °C. Then 35 cycles in the mode: 1 min/94 °C - denaturation; primer annealing - 1 min/60 °C; DNA synthesis - 1 min/72 °C; completion - 5 min/72 °C. The results of molecular genetic analysis indicate that the length distribution and frequency of occurrence of DNA fragments are characteristic features of the char genome. DNA fragments from 200 to 1050 bp long are specific for Dryagin char. The results obtained will be used to determine the species systematic affiliation of the char when introduced into the aquaculture of the Krasnoyarsk Region.