Have a personal or library account? Click to login
Genetic characterisation of centuries-old oak and linden trees using SSR markers Cover

Genetic characterisation of centuries-old oak and linden trees using SSR markers

Folia Forestalia Polonica
Volume 64 (2022): Issue 1 (March 2022)
By: Svitlana Bilous,  Larysa Prysiazhniuk,  Snizhana Chernii,  Serghii Melnyk,  Yurii Marchuk and  Artur Likhanov  
Open Access
|Mar 2022

Figures & Tables

Figure 1

Results of PCR testing of samples of common oak with marker: A – ssrQrZAG 11: 1 – Yuzefin oak; 2 – Shevchenko oak; 3 – Rylskyi oak; 4 – centuries-old oak in the botanical garden of the NULES; 5 – Vytautas oak; 6 – centuries-old oak on the territory of NULES Ukraine; B – ssrQrZAG 65: 1 – Rylskyi oak; 2 – centuries-old oak in the NULES botanical garden; 3 – Vytautas oakNote: M – molecular weight marker 100-bp DNA ladder O’GeneRuler (Thermo Scientific), NULES – National University of Life and Environmental Sciences, PCR – polymerase chain reaction.
Results of PCR testing of samples of common oak with marker: A – ssrQrZAG 11: 1 – Yuzefin oak; 2 – Shevchenko oak; 3 – Rylskyi oak; 4 – centuries-old oak in the botanical garden of the NULES; 5 – Vytautas oak; 6 – centuries-old oak on the territory of NULES Ukraine; B – ssrQrZAG 65: 1 – Rylskyi oak; 2 – centuries-old oak in the NULES botanical garden; 3 – Vytautas oakNote: M – molecular weight marker 100-bp DNA ladder O’GeneRuler (Thermo Scientific), NULES – National University of Life and Environmental Sciences, PCR – polymerase chain reaction.

Figure 2

Cluster distribution of Quercus robur samples by SSR markersNote: Q1 – Yuzefin oak; Q2 – Shevchenko oak; Q3 – Rylskyi oak; Q4 – centuries-old oak in the NULES botanical garden; Q5 – Vytautas oak; Q 6 and Q7 - centuries-old oak near the territory of NULES Ukraine; NULES – National University of Life and Environmental Sciences, SSR – simple sequence repeat.
Cluster distribution of Quercus robur samples by SSR markersNote: Q1 – Yuzefin oak; Q2 – Shevchenko oak; Q3 – Rylskyi oak; Q4 – centuries-old oak in the NULES botanical garden; Q5 – Vytautas oak; Q 6 and Q7 - centuries-old oak near the territory of NULES Ukraine; NULES – National University of Life and Environmental Sciences, SSR – simple sequence repeat.

Figure 3

Cluster distribution of Tilia cordata samples by SSR markersNote: L.1 T.H. Shevchenko Tilia cordata, L.2 – P. Mohyla Tilia cordata, L.3 Tilia cordata of St. Theodosius of Pechersk, L.4 a centuries-old Tilia cordata of the Feofaniia Monument Park, L.5 a centuries-old Tilia cordata (the village of Irsha), L.6 a centuries-old Tilia cordata in the Holosiivskyi forest.
Cluster distribution of Tilia cordata samples by SSR markersNote: L.1 T.H. Shevchenko Tilia cordata, L.2 – P. Mohyla Tilia cordata, L.3 Tilia cordata of St. Theodosius of Pechersk, L.4 a centuries-old Tilia cordata of the Feofaniia Monument Park, L.5 a centuries-old Tilia cordata (the village of Irsha), L.6 a centuries-old Tilia cordata in the Holosiivskyi forest.

Figure 4

Relationship between genetic distances of Quercus robur samples according to geographical coordinates and SSR markers (SSR – simple sequence repeat)
Relationship between genetic distances of Quercus robur samples according to geographical coordinates and SSR markers (SSR – simple sequence repeat)

Figure 5

Relationship between genetic distances of Tilia cordata samples according to geographical coordinates and SSR markers (SSR – simple sequence repeat)
Relationship between genetic distances of Tilia cordata samples according to geographical coordinates and SSR markers (SSR – simple sequence repeat)

Material characteristics

Centuries-old trees nomenclatureLocationAge [years]Geographical coordinates
Quercus robur
Q1Yuzefinskyy oakHlynne, Rivne regionnearly 100051°55′33″N 27°37′86″E
Q2Oak T. ShevchenkoKyivmore than 60050°49′59″N 30°45′16″E
Q3M. Rylskyi oakKyivnearly 60050°38′68″N 30°51′01″E
Q4Centuries-old oak in the NULES botanical gardenKyivmore than 20050°38′16″N 30°50′28″E
Q5Oak VitovtaKyivmore than 40050°38′42″N 30°49′73″E
Q6Centuries-old oak 1 on the territory of NULES UkraineKyivmore than 40050°38′38″N 30°50′53″E
Q7Centuries-old oak 2 on the territory of NULES UkraineKyivmore than 40050°38′37″N 30°50′61″E
Tilia cordata
L1Linden T. ShevchenkoSedniv, Chernihiv regionmore than 60051°63′76″N 31°56′95″E
L2Linden P. MohylaKyivmore than 60050°45′75″N 30°51′71″E
L3Centuries-old linden tree of the Feofaniia Monument ParkKyivmore than 40050°43′25″N 50°43′25″E
L4Linden tree of St. Feodosiya Pechers′kohoKyivmore than 70050°34′00″N 30°48′’68″E
L5Centuries-old linden tree in Holosiivskyi forestKyivmore than 20050°72′18″ N 29°44′92″E
L6Centuries-old linden treev. Irsha, Radomyshl district, Zhytomyr regionmore than 20050°38′29″ N 30°50′51″E

Sizes of obtained alleles for Quercus robur with SSR markers

SSRAllele size, bp
Q1*Q2Q3Q4Q5Q6Q7
ssrQrZAG 7132120, 140132152152140146
ssrQrZAG 11300332268, 300290290268, 300290
ssrQrZAG 25116206206206194158, 206112, 158
ssrQrZAG 30204, 226212, 230182192212192, 212204
ssrQrZAG 31146, 182146, 182140, 158174182158146, 182
ssrQrZAG 44116, 156124, 180144144, 180144156, 216156
ssrQrZAG 65272, 320264, 312312312, 340296, 320396388

Characteristics of SSR markers for assessment of Quercus robur polymorphism

SSRNucleotide sequences of primers, 5′→3′MotiveThe expected size of the amplicons, bp
ssrQrZAG 7F: CAACTTGGTGTTCGGATCAA(TC)17150
R: GTGCATTTCTTTTATAGCATTCAC
ssrQrZAG 11F: CCTTGAACTCGAAGGTGTCCTT(TC)22273
R: GTAGGTCAAAACCATTGGTTGACT
ssrQrZAG 25F: GATATGAAAGATTCTTATTCCATCC(GA)32135
R: GTTAGAACCAATGTACCAAAGTCC
ssrQrZAG 30F: TGCTCCGTCATAATCTTGCTCTGA(GA)26211
R: GCAATCCTATCATGCACATGCACAT
ssrQrZAG 31F: CTTAGTTTGGTTGGGAAGAT(GA)31190
R: GCAACCAAACAAATGAAAT
ssrQrZAG 44F: ACCCTTGTAGTCATGTTCGTTG(GA)29 (TG)31145
R: GAAATCTCACCTGCTCCCTATC
ssrQrZAG 65F: CAGTGGTGTCAACTCCTCCCAG(TC)21(TA)10270
R: GTCAGGTGACCATTCAAACCTAGAA

Sizes of obtained alleles for Tilia cordata with SSR markers

SSRAllele size, bp
L1*L2L3L4L5L6
Tc927152, 168152152152, 168152152
Tc5154154, 174, 180154158154154, 180
Tc915154154, 168154154, 174182168
Tc920218, 232224, 240218, 232232232, 252252
Tc937152152152162162162
Tc963246246238238246246

PCR protocol for Quercus robur and Tilia cordata

Step nameQ. roburT. cordata
parameters [°C/time]cycle numberparameterscycle number
Initial denaturation95/3 min195/3 min1
Denaturation94 (89)*/30 s33*94/30 s35
Annealing50/30s54**/1 min
Extension72/30 s72/30 s
Final extension72/3 min172/3 min1

Characteristics of SSR markers for assessment of Tilia cordata polymorphism

SSRNucleotide sequences of primers, 5′→3′MotiveThe expected size of the amplicons, bp
Tc5F: TTTTCATACATTTAGAGACTTTTAGCA(AG)12150
R: TGCATGATTTGTATGTTTAGGG
Tc915F: ACATCGATTGTATTTCCCTTTAAC(CT)16165
R: GTTGTATTTTGCCCTTAACATTG
Tc920F: AAATGTCTTCAGAGTGACTAGATGG(GA)2(GT)15 (AG)4232
R: TGCCTCATTATTCTCCTAATTCTC
Tc927F: AGTCCTCCTGTCAAATGCTG(AG)10157
R: ATCACACTCGTTTATGACATCTTG
Tc937F: AGCCAACCAACTTTTACAATACAG(AG)13162
R: AGATAAAAGCACATAAATCGATGG
Tc963F: CTAACCCCATTCTCTTTAATTCTG(CT)11238
R: GCTTTCATTTCAGTTTTCCTCTAC

Composition of reaction mixture for PCR of Quercus robur and Tilia cordata

ComponentsFinal concentration
Q. roburT. cordata
PCR buffer*1×1×
MgCl21.5 μM2 μM
dNTP (dATP, dTTP, dGTP, dCTP)100 μM200 μM
Polymerase Taq1 U0.5 U
Primer (F)1 μM0.2 μM
Primer (R)1 μM0.2 μM
DNA50 ng50 ng

Characteristics of the obtained alleles by SSR markers for the samples of Quercus robur and Tilia cordata

SSRNumber of allelesAllele size rangeAlleles’ frequencyPIC
Q. robur
ssrQrZAG 75120–1520.07–0.290.77
ssrQrZAG 114268–3320.14–0.430.69
ssrQrZAG 255112–2060.07–0.500.68
ssrQrZAG 306182–2300.07–0.290.80
ssrQrZAG 315140–1820.07–0.360.76
ssrQrZAG 446116–2160.07–0.360.76
ssrQrZAG 658264–3960.07–0.290.84
T. cordata
Tc54154–1800.08–0.580.60
Tc9154154–1820.08–0.500.65
Tc9205218–2520.08–0.250.72
Tc9272152–1680.17–0.830.28
Tc9372152–1620.50.50
Tc9632238–2460.33–0670.44
DOI: https://doi.org/10.2478/ffp-2022-0006 | Journal eISSN: 2199-5907 | Journal ISSN: 0071-6677
Journal RSS Feed
Language: English
Page range: 58 - 68
Submitted on: Jul 5, 2021
|
Accepted on: Feb 8, 2022
|
Published on: Mar 11, 2022
Published by: Forest Research Institute
In partnership with: Paradigm Publishing Services
Publication frequency: 4 issues per year
Keywords:
centuries-old trees,
DNA analysis,
L.,
L
Related subjects:
Life sciences,
Plant science,
Medicine,
Veterinary medicine

© 2022 Svitlana Bilous, Larysa Prysiazhniuk, Snizhana Chernii, Serghii Melnyk, Yurii Marchuk, Artur Likhanov, published by Forest Research Institute
This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 License.

Previous articleVolume 64 (2022): Issue 1 (March 2022)