1. bookVolume 67 (2018): Issue 1 (February 2018)
Journal Details
First Published
22 Feb 2016
Publication timeframe
1 time per year
access type Open Access

Microsatellite Analysis to Study Genetic Diversity in Khasi Pine (Pinus Kesiya Royle Ex. Gordon) Using Chloroplast SSR Markers

Published Online: 18 Oct 2018
Page range: 99 - 105
Journal Details
First Published
22 Feb 2016
Publication timeframe
1 time per year

Pinus kesiya (Khasi pine) is the principal pine species in northeast India having high commercial value. Chloroplast microsatellites (cpSSR) were used to study the genetic diversity and population genetic structure of 10 populations of P. kesiya covering entire natural range of distribution in India. A total of 33 primer pairs (cpSSRs) of P. thunberghii and P. sylvestris were tested in P. kesiya for their transferability, out of which 18 chlo­roplast primers showed positive amplification and 10 were found polymorphic. A total of 250 individuals from 10 different populations were genotyped using the selected 10 cpSSRs. When alleles at each of the 10 loci were jointly analysed a total of 36 size variants were discovered, which combined to desig­nate 90 haplotypes among 250 individuals. None of the haplo­type was found common among the populations as they were population specific. The cpSSR indicated that P. kesiya popula­tions have maintained a moderately high genetic diversity (HT=0.638) which is typical in most coniferous species. Howe­ver, the inter-population genetic diversity was higher than the intra population diversity and the genetic differentiation bet­ween populations was also found to be very high (FST=0.47). A Bayesian cluster analysis separated the populations into six clusters where most of the individuals were found in single population clusters with minor admixtures. The distribution of genetic diversity and sub structuring of P. kesiya reflect week pollen mediated gene flow due to geographic isolation and genetic drift. The study has revealed useful cpSSR markers for P. kesiya, which were lacking earlier and also added an insight into the state of Khasi pine forest in the region, which can be useful for the better management and future conservation programs.


AITKEN SN, YEAMAN S, HOLLIDAY JA, WANG T, CURTIS MCLANE S (2008) Adapta­tion, migration or extirpation: climate change outcomes for tree popula­tions. Evolutionary Applications, 1(1): 95-111. https://doi.org/10.1111/j.1752-4571.2007.00013.x10.1111/j.1752-4571.2007.00013.xOpen DOISearch in Google Scholar

AVISE JC (1994) Molecular Markers, Natural History and Evolution. Chapman and Hall, New York. https://doi.org/10.1007/978-1-4615-2381-910.1007/978-1-4615-2381-9Open DOISearch in Google Scholar

BAYER C, FAY MF, DE BRUIJN AY, SAVOLAINEN V, MORTON CM, KUBITZKI K, AL­VERSON WS, CHASE MW (1999) Support for an expanded family concept of Malvaceae within recircumscribed order Malvales: a combined analysis of plastid atpB and rbcL DNA sequences. Botanical Journal of the Linnean So­ciety, 129: 267-303. https://doi.org/10.1111/j.1095-8339.1999.tb00505.x10.1111/j.1095-8339.1999.tb00505.xOpen DOISearch in Google Scholar

CAI N, XU Y, WANG D, CHEN S, LI G (2017) Identification and characterization of microsatellite markers in Pinus kesiya var. langbianensis (Pinaceae). Applica­tions in Plant Sciences, 5(2): 1-4. https://doi.org/10.3732/apps.160012610.3732/apps.1600126Open DOISearch in Google Scholar

CATO SA, RICHARDSON TE (1996) Inter- and intraspecific polymorphism at chlo­roplast SSR loci and the inheritance of plastids in Pinus radiata D. Don. The­oretical and Applied Genetics, 93: 587-592. https://doi.org/10.1007/s001220050319 CHAUDHARY V, BHATTACHARYYA A (2002) Suitability of Pinus kesiya in Shillong, Meghalaya for tree-ring analysis, Current Science, 83(8): 1010-1015.10.1007/s001220050319CHAUDHARY(2002)PinuskesiyaMeghalayatree-83(8):1010-1015Open DOISearch in Google Scholar

CHAUHAN P (2011) Molecular genetic analysis of Chir pine (Pinus roxburghiiSar­gh.) through microsatellite markers. Ph.D. Thesis submitted to Forest Re­search Institute (Deemed) University.Search in Google Scholar

CHAUHAN P, GINWAL HS, RAWAT A, BARTHWAL S (2010) Cross-species amplifica­tion and characterization of chloroplast and nuclear microsatellite markers in Himalayan Chir Pine (Pinus roxburghii Sarg).Molecular Ecology Resourc­es,11: 219-222.Search in Google Scholar

CLARK CM, WENTWORTH TR, O’MALLEY DM(2000) Genetic discontinuity re­vealed by chloroplastmicrosatellites in Eastern North AmericanAbies (Pina­ceae). American Journal of Botany,87: 774-782. https://doi.org/10.2307/265688510.2307/2656885Open DOISearch in Google Scholar

CORANDER J, MARTTINEN P (2006) Bayesian identification of admixture events using multilocus molecular markers. Molecular Ecology, 15: 2833-2843. https://doi.org/10.1111/j.1365-294x.2006.02994.x10.1111/j.1365-294x.2006.02994.xOpen DOISearch in Google Scholar

CORANDER J, WALDMANN P, SILLANPAA MJ (2003) Bayesian analysis of genetic differentiation between populations. Genetics, 163: 367-374.10.1093/genetics/163.1.367Search in Google Scholar

CORANDER J, WALDMANN P, MARTTINEN P, SILLANPAA MJ (2004) BAPS 2: en­hanced possibilities for the analysis of genetic population structure. Bioin­formatics,20: 2363-2369. https://doi.org/10.1093/bioinformatics/bth250 Bayesian analysis of population structure manual v. 6.0. http://www.helsinki.fi/bsg/software/BAPS/ Assessed 9 August 2018.10.1093/bioinformatics/bth250Search in Google Scholar

DELGADO P, PINERO D, CHAOS A, PEREZ-NASSER N, ALVAREZ-BUYLLA ER (1999) High population differentiation and genetic variation in the endangered Mexican pine Pinus rzedowskii (Pinaceae). American Journal of Botany, 86(5): 669-676. https://doi.org/10.2307/265657610.2307/2656576Open DOISearch in Google Scholar

DIEKMANN K, HODKINSON TR, BARTH S (2012) New chloroplast microsatellite markers suitable for assessing genetic diversity of Lolium perenne and oth­er related grass species. Annals of Botany, 110: 1327-1339. https://doi.org/10.1093/aob/mcs04410.1093/aob/mcs044Open DOISearch in Google Scholar

DOYLE JJ, DOYLE JL (1990) A rapid total DNA preparation procedure for fresh plant tissue. Focus, 12: 13-15. ECHT CS, DEVERNO LL, ANZIDEI M, VENDRAMIN GG (1998) Chloroplast microsat­ellite reveals population genetic diversity in red pine, Pinus resinosa Ait. Molecular Ecology, 7: 307-316. https://doi.org/10.1046/j.1365-294x.1998.00350.x10.1046/j.1365-294x.1998.00350.xOpen DOISearch in Google Scholar

ELIADES N-GH, GAILING O, LEINEMANN L, FADY B, FINKELDEY R (2011) High ge­netic diversity and significant population structure in Cedrus brevifolia Henry, a narrow endemic Mediterranean tree from Cyprus. Plant Systemat­ics and Evolution, 294: 185-198. https://doi.org/10.1007/s00606-011-0453-z10.1007/s00606-011-0453-zOpen DOISearch in Google Scholar

ELLEGREN H (2004) Microsatellites: simple sequences with complex evolution. Nature Reviews Genetics, 5: 435-445. https://doi.org/10.1038/nrg134810.1038/nrg1348Open DOISearch in Google Scholar

EXCOFFIER L, LAVAL G, SCHNEIDER S (2005) Arlequin ver. 3.0: An integrated soft­ware package for population genetics data analysis. Evolutionary Bioinfor­matics Online, 1: 47-50. https://doi.org/10.1177/11769343050010000310.1177/117693430500100003Search in Google Scholar

FALK DA, RICHARDS CM, MONTALVO AM; KNAPP EE (2006) Chapter 2: Popula­tion and ecological genetics in restoration ecology. In Foundations of resto­ration ecology. Edited by D.A. Falk, M.A. Palmer, and J.B. Zedler. Island Press, Washington, D. C. pp. 14-44.Search in Google Scholar

FRANKHAM R, BRISCOE DA, BALLOU JD (2002) Introduction to conservation ge­netics. Cambridge University Press, New York, New York, USA. https://doi.org/10.1017/cbo978051180899910.1017/cbo9780511808999Open DOISearch in Google Scholar

GANEA S, RANADE SS, HALL D, ABRAHAMSSON S, GARCIA-GIL MR (2015) Devel­opment and transferability of two multiplex nSSR in Scots pine (Pinus syl­vestris L.). Journal of Forestry Research, 26(2): 361-368. https://doi.org/10.1007/s11676-015-0042-z10.1007/s11676-015-0042-zOpen DOISearch in Google Scholar

GΌMEZ A, GONZÁLEZ- MARTÍNEZ SC, COLLADA C, CLIMENT J, GIL L (2003) Com­plex population genetic structure in the endemic Canary Islandpine re­Search in Google Scholar

vealed using chloroplast microsatellite markers. Theoretical and Applied Genetics, 107:1123-1131. https://doi.org/10.1007/s00122-003-1320-210.1007/s00122-003-1320-2Open DOISearch in Google Scholar

GΌMEZ- GARAY A, MASSELLI S, BUENO MA (2010) Distribution of Genetic Diver­sity of Pinus ayacahuite (Ehrenberg) at the Communal Forest of Totonica­pan, Guatemala. Bioremediation, Biodiversity and Bioavailability, 4: 35-41.Search in Google Scholar

HAMRICK JL, GODT MJW (1992) Sherman-Broyles, S.L. Factors influencing levels of genetic diversity in woody plant species. New Forest, 6: 95-124. HANSEN OK, KJÆR ED, Vendramin GG (2005) Chloroplast microsatellite variation in Abies nordmanniana and simulation of causes for low differentiation among populations. Tree Genetics & Genomes, 1: 116-123. https://doi.org/10.1007/s11295-005-0016-y10.1007/s11295-005-0016-yOpen DOISearch in Google Scholar

HARTL DL, CLARK AG (1997) Principles of Population Genetics, 3rdedn. Sinauer Associates, Inc, Sunderland, MA.Search in Google Scholar

KELCHNER SA (2000) The evolution of non-coding chloroplast DNA and its ap­plication in plant systematics. Annals of Missouri Botanical Garden, 87:499- 527. https://doi.org/10.2307/2666142 LEDIG FT, CAPΌ- ARTEAGA MA,10.2307/2666142FTCAPΌ-Open DOISearch in Google Scholar

HODGSKISS PD, SBAY H, FLORES-LΌPEZ C, CON­KLE MT, BERMEJO-VELÁZQUEZ B (2001) Genetic diversity and the mating system of a rare Mexican Piñon, Pinus pinceana, and a comparison with Pi­nus maximartinezii (Pinaceae). American Journal of Botany, 88(11): 1977- 1987. https://doi.org/10.2307/355842510.2307/3558425Open DOISearch in Google Scholar

LΌPEZ- VINYALLONGA S, LΌPEZ- ALVARADO J, CONSTANTINIDIS TH, SUSANNA A, GARCIA- JACAS N (2011) Microsatellite cross- species amplification in the genus Centaurea (Compositae), Collectanea Botanica, 30: 17-27. https://doi.org/10.3989/collectbot.2011.v30.00210.3989/collectbot.2011.v30.002Open DOISearch in Google Scholar

MCDERMOTT JM, MCDONALD BA (1993) Gene flow in plant pathosystems. Annu. Rev. Phytopathol,31: 353-373. https://doi.org/10.1146/annurev.py.31.090193.00203310.1146/annurev.py.31.090193.002033Open DOISearch in Google Scholar

MILLER CN (1982) Current status of Paleozoic and Mesozoic confers. Review of Palaeobotany and Palynology, 37: 99-114 (Cited by Millar, 1999) https://doi.org/10.1016/0034-6667(82)90039-210.1016/0034-6667(82)90039-2Open DOISearch in Google Scholar

MIROV NT (1967) The genus Pinus. Ronald Press, New York, 602 pp. https://doi.org/10.2307/4004229 MORENO AC, MARCHELLI P, VENDRAMIN GG, GALLO LA (2011) Cross transfer­ability of SSRs to five species of Araucariaceae: useful tool for population genetic studies in Araucaria araucana. Forest Systems, 20(2): 303-314. https://doi.org/10.5424/fs/2011202-1144910.5424/fs/2011202-11449Search in Google Scholar

MORGANTE M, PFEIFFER A, COSTACURTA A, OLIVIERI AM (1996) Molecular tools for population and ecological genetics in coniferous trees. Phyton-Annales Rei Botanicae, 36: 129- 138. NEALE DB, SEDEROFF RR (1989) Paternal inheritance of chloroplast DNA and maternal inheritance of mitochondrial DNA in loblolly pine. Theoretical and Applied Genetics, 77: 212-216. https://doi.org/10.1007/bf0026618910.1007/bf00266189Open DOISearch in Google Scholar

NEI M (1987) Molecular Evolutionary Genetics. Columbia University Press, New York. https://doi.org/10.1016/0047-2484(89)90093-610.1016/0047-2484(89)90093-6Open DOISearch in Google Scholar

PERRY JP (1991) The Pines of Mexico and Central America. Timber Press, Port­land,OR. https://doi.org/10.2307/122252310.2307/1222523Open DOISearch in Google Scholar

PORTH I, EL-KASSABY YA (2014) Assessment of the genetic diversity in forest tree populations using molecular markers. Diversity, 6: 283-295. https://doi.org/10.3390/d602028310.3390/d6020283Open DOISearch in Google Scholar

PROVAN J, SORANZO N, WILSON NJ, MCNICOL JW, FORREST GI, COTRELL J, POW­ELL E (1998) Genepool variation in Caledonia and European Scots pine (Pi­nus sylvestris L.) revealed by chloroplast Simple Sequence Repeats. In: Pro­ceedings of Royal Society of London, Series B, 265: 1697-1705. https://doi.org/10.1098/rspb.1998.049110.1098/rspb.1998.0491Open DOISearch in Google Scholar

PROVAN J, SORANZO N, WILSON NJ, GOLDSTEIN DB, POWELL W (1999).A low mutation rate for chloroplast microsatellites. Genetics, 153: 943-947. QUELLER D, STRASSMANN J, HUGHES CR (1993)10.1093/genetics/153.2.943Search in Google Scholar

Microsatellites and kinship. Trends in Ecology and Evolution, 8:285-288. https://doi.org/10.1016/0169-5347(93)90256-o RIBEIRO MM, MARIETTE S, VENDRAMIN GG, SZMIDT AE, PLOMION C, KREMER A (2002) Comparison of genetic diversity estimates within and among popu­lations of maritime pine using chloroplast simple-sequence repeat and am­plified fragment length polymorphism data. Molecular Ecology,11: 869-877. https://doi.org/10.1046/j.1365-294x.2002.01490.x10.1046/j.1365-294X.2002.01490.xSearch in Google Scholar

SCHUG MD, SMITH SG, TOZIER-PEARCEA, MCEVEY SF (2007):The genetic struc­ture of Drosophila ananassae populations from Asia, Australia and Samoa. Genetics, 175: 1429-1440. https://doi.org/10.1534/genetics.106.06661310.1534/genetics.106.066613Open DOISearch in Google Scholar

SINCLAIR WT, MONCUR JD, ENNOS RA (1997) Multiple origins for Scots pine (Pi­nus sylvestris L.) in Scotland: evidence from mitochondria DNA variation. Heredity, 80: 233-240. https://doi.org/10.1038/sj.hdy.688287010.1038/sj.hdy.6882870Open DOISearch in Google Scholar

STANGE C, PREHN D, JOHNSON PA (1998) Isolation of Pinus radiata genomic DNA suitable for RAPD analysis. Plant Molecular Biology Reporter, 16: 1-8.10.1023/A:1007540901981Search in Google Scholar

TERRAB A, PAUN O, TALAVERA S, TREMETSBERGER K, ARISTA M, STUESSY TF (2006) Genetic diversity and population structure in naturalpopulations of Moroccan Atlas Cedar (Cedrus atlantica; Pinaceae) determined with cpSSR markers. American Journal of Botany, 93: 1274-1280. https://doi.org/10.3732/ajb.93.9.127410.3732/ajb.93.9.1274Open DOISearch in Google Scholar

TORO MA, CABALLERO A (2005) Characterization and conservation of genetic diversity in subdivided populations. Philosophical Transactions of the Royal Society, 360: 1367-1378. https://doi.org/10.1098/rstb.2005.168010.1098/rstb.2005.1680Open DOISearch in Google Scholar

VENDRAMIN GG, LELLILR ROSSI P, MORGANTE M (1996) A set of primers for the amplification of 20 chloroplast microsatellites in Pinaceae. Molecular Ecolo­gy, 5: 595-598. https://doi.org/10.1111/j.1365-294x.1996.tb00353.x10.1111/j.1365-294x.1996.tb00353.xOpen DOISearch in Google Scholar

VENDRAMIN GG, ANZIDEI M, MADAGHIELE A, BUCCI G (1998) Distribution of ge­netic diversity in Pinus pinaster Ait. as revealed by chloroplast microsatel­lites. Theoretical and Applied Genetic, 97: 456- 463. https://doi.org/10.1007/s00122005091710.1007/s001220050917Open DOISearch in Google Scholar

VENDRAMIN GG, ANZIDEI M, MADAGHIELE A, SPERISEN C, BUCCI G (2000) Chlo­roplast microsatellite analysis reveals the presence of population subdivi­sion in Norway spruce (Picea abies K.). Genome, 43: 68-78. https://doi.org/10.1139/gen-43-1-6810.1139/gen-43-1-68Open DOISearch in Google Scholar

VILLALOBOS- ARÁMBULA AR, PÉRÉZ DE LA ROSA JA, ARIAS A, RAJORA OP (2014) Cross-species transferability of eastern white pine (Pinus strobus) nuclear microsatellite markers to five Mexican white pines. Genetics and Molecular Research,13: 7571-7576. https://doi.org/10.4238/2014.september.12.2410.4238/2014.september.12.24Open DOISearch in Google Scholar

WANG D, SHI J, CARLSON SR, CREGAN PB, WARD RW, Diers BW (2003) Alow cost, high throughput Polyacrylamide gel electrophoresis system forgenotying with microsatellite DNA markers. Crop Science, 43: 1828-1832. https://doi.org/10.2135/cropsci2003.182810.2135/cropsci2003.1828Open DOISearch in Google Scholar

WATANO Y, IMAZU M, SHIMIZU T (1996) Spatial distribution of cpDNA and mtD­NA haplotypes in a hybrid zone between Pinus pumila and P. parviflora var. pentaphylla. 2ndedn. Taylor &Francis Group, CRS Press, New York. https://doi.org/10.1007/bf0234455510.1007/bf02344555Open DOISearch in Google Scholar

WRIGHT S (1951) The genetical structure of populations. Annals of Eugenics,15: 323-354. https://doi.org/10.1111/j.1469-1809.1949.tb02451.x XIANG-XIANG F, JI-SEN S (2005) Identification of seeds of Pinus species by Micro­satellite Markers. Journal of Forestry Research, 16(4): 281-284. https://doi.org/10.1007/bf0285818910.1007/BF02858189Search in Google Scholar

XU Y, ZHANG R, TIAN B, BAI Q, WANG D, CAI N, HE C, KANG X, DUAN A (2013) De­velopment of novel microsatellite markers for Pinus yunnanensis and their cross amplification in congeneric species. Conservation Genetic Re­source,5(4):1113-1114. https://doi.org/10.1007/s12686-013-9964-y10.1007/s12686-013-9964-yOpen DOISearch in Google Scholar

YEH FC, YANG RC, BOYLE TBJ (1999) PopGene Version 1.31: Microsoft windows based Freeware for Population Genetic Analysis. University of Albert, Ed­monton. http://www.ualbert.ca/fyeh.Search in Google Scholar

Recommended articles from Trend MD

Plan your remote conference with Sciendo