title: ISSR markers for gender identification and genetic diagnosis of Hippophae rhamnoides ssp. turkestanica growing at high altitudes in Ladakh region (Jammu and Kashmir).
abstract: Hippophae rhamnoides L. ssp. turkestanica (Elaeagnaceae) is a predominantly dioecious and wind-pollinated medicinal plant species. The mature fruits of the species possess antioxidative, anti-inflammatory, antimicrobial, anticancerous, and antistimulatory properties that are believed to improve the immune system. The identification of male and female plants in H. rhamnoides ssp. turkestanica is quite difficult until flowering which usually takes 3-4 years or more. A sex-linked marker can be helpful in establishing the orchards through identification of genders at an early stage of development. Therefore, we studied the genetic diversity of populations in Ladakh with the aim to identify a gender-specific marker using ISSR markers. Fifty-eight ISSR primers were used to characterize the genome of H. rhamnoides ssp. turkestanica, of which eight primers generated 12 sex-specific fragments specific to one or more populations. The ISSR primer (P-45) produced a fragment which faithfully segregates all the males from the female plants across all the three valleys surveyed. This male-specific locus was converted into a SCAR. Forward and reverse primers designed from this fragment amplified a 750-bp sequence in males only, thus specifying it as an informative male-specific sex-linked marker. This SCAR marker was further validated for its capability to differentiate gender on an additional collection of plants, representing three geographically isolated valleys (Nubra, Suru, and Indus) from Ladakh region of India. The results confirmed sex-linked specificity of the marker suggesting that this conserved sequence at the Y chromosome is well preserved through the populations in Ladakh region. At present, there are no reliable markers which can differentiate male from female plants across all the three valleys of Ladakh region at an early stage of plant development. It is therefore envisaged that the developed SCAR marker shall provide a reliable molecular tool for early identification of the sex in this commercial crop. The genetic diversity of populations as surveyed by ISSR primers revealed 85.71 % polymorphism at the population level. The dendrogram obtained divided the genotypes into three different clusters, and the distribution of male and female genotypes in all the clusters was random. The Nei's genetic similarity index was in the range of 0.63-0.96.
authors.affiliation: Department of Botany, University of Delhi, Delhi, India.
authors.forename: Kamal
authors.initials: K
authors.lastname: Das
authors.affiliation: Department of Botany, University of Delhi, Delhi, India.
authors.forename: Showkat Hussain
authors.initials: SH
authors.lastname: Ganie
authors.affiliation: Department of Botany, University of Delhi, Delhi, India.
authors.forename: Yash
authors.initials: Y
authors.lastname: Mangla
authors.affiliation: Centre for Biodiversity Studies, School of Biosciences and Biotechnology, Baba Ghulam Shah Badshah University, Rajouri, J&K, India.
authors.forename: Tanvir-Ul-Hassan
authors.initials: TU
authors.lastname: Dar
authors.affiliation: Department of Biotechnology, Amity University, Noida, Uttar Pradesh, India.
authors.forename: Manju
authors.initials: M
authors.lastname: Chaudhary
authors.affiliation: Department of Biotechnology, Amity University, Noida, Uttar Pradesh, India.
authors.forename: Rakesh Kumar
authors.initials: RK
authors.lastname: Thakur
authors.affiliation: Department of Botany, University of Delhi, Delhi, India.
authors.forename: Rajesh
authors.initials: R
authors.lastname: Tandon
authors.affiliation: Department of Biotechnology, Amity University, Noida, Uttar Pradesh, India.
authors.forename: S N
authors.initials: SN
authors.lastname: Raina
authors.affiliation: Department of Botany, University of Delhi, Delhi, India. shailendragoel@gmail.com.
authors.forename: Shailendra
authors.initials: S
authors.lastname: Goel
mesh_terms: D000531:Altitude; D001483:Base Sequence; D018744:DNA, Plant / Q000235:genetics; D005819:Genetic Markers; D005828:Genetics, Population; D005843:Geography; D031281:Hippophae / Q000235:genetics* / Q000254:growth & development*; D007194:India; D018895:Microsatellite Repeats / Q000235:genetics*; D010802:Phylogeny; D011110:Polymorphism, Genetic; D025341:Principal Component Analysis; D015203:Reproducibility of Results
chemical_list: D018744:DNA, Plant; D005819:Genetic Markers
keywords: Gender-specific marker; Genetic diversity; Hippophae rhamnoides; ISSR; SCAR