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Оценка Генетического Разнообразия Популяций Tulipa Suaveolens Волгоградской

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Оценка Генетического Разнообразия Популяций Tulipa Suaveolens Волгоградской ОРИГИНАЛЬНАЯ СТАТЬЯ • ORIGINAL АRTICLE ОЦЕНКА ГЕНЕТИЧЕСКОГО РАЗНООБРАЗИЯ ПОПУЛЯЦИЙ TULIPA SUAVEOLENS ВОЛГОГРАДСКОЙ ОБЛАСТИ DOI: ./----- GENETIC DIVERSITY OF TULIPA SUAVEOLENS ROTH УДК ... POPULATIONS IN VOLGOGRAD PROVINCE Поступление/Received: .. Принято/Accepted: .. Т. А. КРИЦКАЯ, А. С. КАШИН T. A. KRITSKAYA, A. S. KASHIN Саратовский национальный исследовательский Saratov State University, государственный университет им. Н.Г. Чернышевского 83 Astrakhanskaya St., Saratov 410012, Russia; 410012 Россия, г. Саратов, ул. Астраханская, 83; [email protected] [email protected] Актуальность. Современныи естественныи ареал Tulipa Background. The current natural habitat of Tulipa suaveolens Roth стремительно сокращается. Для выбора suaveolens Roth rapidly decreases. In order to work out the стратегии сохранения этого вида необходима оценка strategy of the species’ preservation, evaluation of its intra- внутри- и межпопуляционного полиморфизма. Матери- and interpopulation polymorphism is required. Materials алы и методы. Для анализа 125 образцов 10 популяции and methods. Molecular-genetic ISSR markers were used T. suaveolens Волгоградскои области и 4 популяции Сара- to analyze 125 samples from 10 populations of T. suaveolens товскои области были выбраны молекулярно-генетиче- occurring in Volgograd Province and 4 populations from ские ISSR-маркеры. Результаты. ISSR-анализ позволил Saratov Province. Results. ISSR analysis revealed high выявить высокии уровень полиморфизма (73−89%) вну- intrapopulation polymorphism (73–89%) in T. suaveolens три популяции T. suaveolens Волгоградскои области. По populations form Volgograd Province. AMOVA attributed результатам AMOVA, большая часть изменчивости the largest proportion of variability (74.3%) to (74,3%) приходится на внутрипопуляционныи полимор- intrapopulation polymorphism. Interpopulation differences физм. Доля изменчивости, которая приходится на меж- account for 25.7%. Total subdivision of populations (FST) популяционные различия, составила 25,7%. Общая под- was 0.257; total gene low (Nm) between populations was разделенность популяции (FST) равна 0,257. Общии поток 0.723. According to Bayesian analysis and clustering with генов (Nm) между популяциями – 0,723. Согласно резуль- both UPGMA and Neighbor Joining methods, all the studied татам баи есовского анализа и кластеризации двумя раз- T. suaveolens samples from Volgograd Province make up личными методами (UPGMA и Neighbor Joining), все ис- a large genetic group: within that group none of the potential следованные особи T. suaveolens Волгоградскои области subgroups may be associated with a particular place of образуют одну большую генетическую группу, в которои collecting. The NewHybrids software was applied, and the ни одна из потенциальных подгрупп не может быть ассо- results pointed to the hybrid nature of most samples. циирована с тем или иным раи оном сбора образцов. Ре- Samples of three populations from Saratov Province made зультаты, полученные в программе NewHybrids, указы- up a separate genetic group; those samples fell under the вают на гибридную природу большеи части особеи . От- category of parent forms. Conclusion. Considering that дельную генетическую группу составили образцы трех genetic subdivision of T. suaveolens populations within the популяции Саратовскои области, которые были отнесе- administrative borders of Volgograd Province is ны в категорию родительскои формы. Заключение. Учи- insigniicant, while all the province’s natural parks and тывая то, что в пределах административных границ Вол- a number of protected natural areas undertake measures to гоградскои области популяции T. suaveolens слабо отли- preserve the species, the existing conservation strategy чаются друг от друга генетически и охраняются на тер- may be recognized as effective and suficient. ритории всех природных парков области, а также ряда особо охраняемых природных территории , введенные меры охраны следует считать достаточными для сохра- нения вида. Ключевые слова: Tulipa schrenkii, ISSR-анализ, програм- Key words: Tulipa schrenkii, ISSR analysis, NewHybrids ма NewHybrids, Волгоградская область. software, Volgograd Province. Введение пространен в южных и юго-восточных раи онах Восточнои Европы, включая Крым, а также на Кавказе, в Казахстане, В связи со стремительным сокращением ареалов рас- юге Западнои Сибири. Встречается по всеи Волгоградскои пространения многих дикорастущих видов растении про- области, более часто в южнои и юго-восточнои части. Наи- блема сохранения их биологического разнообразия явля- более крупные попул яции расположены в Палласовском, ется в настоящее время весьма актуальнои . Особенно это Чернышковском, Калачевском, Серафимовичском, Быков- касается близких родственников культурных растении , ском и др. р-нах, одна из самых обширных – близ оз. Булух- которые могут служить донорами ценных генов и призна- та (более 100 км2) (Popov et al., 2017). Однако высокая чи- ков в селекционном процессе. сленность популяции далеко не всегда коррелирует Одним из таких видов является Tulipa suaveolens Roth с уровнем ее генетического разнообразия (Trifonova et al., (= T. schrenkii Regel, T. gesneriana L.) – высоко декоративныи 2017). Поэтому для успешного сохранения генофонда луковичныи поликарпик семеи ства Liliaceae, родоначаль- T. suaveolens Волгоградскои области необходима оценка ник садовых тюльпанов (Mordak, 1990; Zonnenveld, 2009). внутри- и межпопуляционного полиморфизма. Вид занесен в Красные книги России, Украины, Казахстана Такая оценка уже проводилась для популяции (как T. schrenkii) и Азербаи джана (как T. gesneriana). Рас- T. suaveolens Саратовскои области и Республики Крым , 180 (4), 2019 . • . • 180 (4), 2019 • (Kashin et al., 2016; Kritskaya et al., 2018; Kritskaya, Kashin, ДНК выделяли с использованием набора NucleoSpin® 2018) с помощью маркеров ISSR (Inter Simple Sequence Plant II (MACHEREY-NAGEL, Германия) согласно протоко- Repeats). Полученные результаты позволили подтвердить лу производителя. ПЦР проводили в амплификаторе рациональность распределения особо охраняемых при- Mastercycler gradient (Eppendorf, Germany) с 10 ISSR-праи - родных территории в указанных регионах, в полнои мере мерами, синтезированными НПК «Синтол» (Москва), вы- охватывающих генетическое разнообразие T. suaveolens. бранными нами ранее для T. suaveolens (Kashin et al., Несмотря на то что в Волгоградскои области активно 2016). ПЦР проводили в объеме 20 μl. Реакционная смесь проводятся исследования генетического разнообразия содержала 4 μl готовои реакционнои смеси MaGMix (по популяции редких и исчезающих растении с помощью мо- 200 μM каждого dNTP, 1.5 mM MgCl2, 1.5 ед. SmarTaqDNA- лекулярно-генетических методов (Khadeeva et al., 2011; полимеразы и буфер; Dialat Ltd., Москва, Россия), 15 μl де- Khadeeva et al., 2012; Trifonova et al., 2017), подобного рода ионизированнои воды, 3.4 pmol каждого праи мера и 1 μl исследования регионального масштаба до настоящего исходнои ДНК. Программа ПЦР включала следующие времени не затрагивали этот уязвимыи вид. этапы: изначальная денатурация в течение 5 мин при Цель данной работы – оценить генетическое разно- 95°C, затем 35 циклов по 30 сек при 95°C, 30 сек при 44°C образие и определить популяционно-генетическую струк- и 2 мин при 72°C, с финальнои элонгациеи в течение туру T. suaveolens в Волгоградскои области с помощью 10 мин при 72°C. ISSR-маркеров. Полученную матрицу предварительно анализиро- вали в программе PAST (Hammer et al., 2001) методом Материалы и методы попарного невзвешенного среднего (UPGMA) с исполь- зованием коэффициента Жаккара и в программе Сбор материала проводили в десяти природных попу- SplitsTree 4 (Huson, Bryant, 2006) методом Neighbor ляциях T. suaveolens Волгоградскои области и четырех Joining (NJ). Анализ популяционнои структуры прово- популяциях прилегающеи территории Саратовскои об- дили методом Баи еса в программе Structure 2.3 ласти. Всего собрано 125 образцов, представляющих (Pritchard et al., 2000; Evanno et al., 2005; Jakobsson, 14 популяции (табл. 1). Материалом служили листья, вы- Rosenberg 2007). Анализ проводился с использованием сушенные силикагелем. модели генетического смешения (admixture). Предва- Таблица 1. Перечень образцов Tulipa suaveolens Roth, использованных в исследовании Table 1. The list of Tulipa suaveolens Roth samples involved in the study №Место сбора / Collecting site nN Np N/Np Волгоградская обл., Котельниковскии р-н, окр. хут. Захаров 1 5731000.73 Volgograd Province, Kotelnikovsky District, Zakharov farm Волгоградская обл., Светлоярскии р-н, окр. ст. Тингута 2 570910.77 Volgograd Province, Svetloyarsky District, Tinguta station Волгоградская обл., Ленинскии р-н, окр. г. Ленинск 3 10 100 112 0.89 Volgograd Province, Leninsky District, Leninsk town Волгоградская обл., Иловлинскии р-н, окр. хут. Хмелевскои 4 10 111 127 0.87 Volgograd Province, Ilovlinsky District, Khmelevskoy farm Волгоградская обл., Алексеевскии р-н, окр. хут. Нестеровскии 5 10 87 103 0.85 Volgograd Province, Alekseevsky District, Nesterovsky farm Волгоградская обл., Михаи ловскии р-н, окр. г. Михаи ловка 6 10 101 117 0.86 Volgograd Province, Mikhailovsky District, Mikhailovka town Волгоградская обл., Быковскии р-н, окр. с. Верхнии Балыклеи 7 10 90 104 0.87 Volgograd Province, Bykovsky District, Verkhny Balykley village Волгоградская обл., Палласовскии р-н, берег оз. Эльтон 8 574930.80 Volgograd Province, Pallasovsky District, Elton Lake shore Волгоградская обл., Даниловскии р-н, 7 км С-З с. Орехово 9 10 104 120 0.87 Volgograd Province, Danilovsky District, 7 km SW Orekhovo
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