DOI: 10.5061/dryad.pzgmsbd1v
title: Data from: DNA barcoding analyses and taxonomy reveal two new species of genus <em>Inocybe</em> from pine forests of Pakistan

description: Here we describe two new species of Inocybe from the Pine forests of
 Pakistan: I. hazarensis and I. shimlaensis. Morphological and molecular
 data show that these species have not been described before, hence they
 need to be described as new. Both species are smooth-spored and pruinose
 only in the apical part of the stipe. Inocybe hazarensis is characterized
 by a rather small size, brown to dark brown, dense radial fibrils to
 rimose to glabrous pileus with prominent umbo, finely fibrillose at apex
 only becoming pruinose to strigose, along the rest of the length of stipe,
 slightly bulbous stipe base, narrow basidiospores spores and smaller
 clavate, oblong, ovoid, narrowly utriform cheilocystidia. Inocybe
 shimlaensis is characterized by a brown to golden pileus, a low and broad
 umbo, radially fibrillose, rimose to granulose, with brown appressed
 squalmulose, a stipe with only the apex pruinose, and a submarginate base.
 Anatomically, it has small (6.8 × 4.4 μm) basidiospores and smaller
 conical to fusiform cheilocystidia. Phylogenetic estimation based on DNA
 sequences from the internal transcribed spacer (ITS) region and large
 subunit (LSU) of the nuclear ribosomal DNA (rDNA) genes is congruent with
 the morphological characters that help to delimit these as new species of
 Inocybe.
description: # Data from: DNA barcoding analyses and taxonomy reveal two new species of
 genus *Inocybe* from pine forests of Pakistan Dataset DOI:
 [10.5061/dryad.pzgmsbd1v](https://doi.org/10.5061/dryad.pzgmsbd1v) ##
 Description of the data and file structure Microscopic characters are
 based on free-hand sections from fresh and dried specimens mounted in 5%
 (w/v) aqueous potassium hydroxide (KOH) solution. Tissues from lamellae,
 pileipellis, and stipitipellis were mounted in phloxine (1%) for better
 contrast and examined using a Meiji Techno MX4300H compound microscope. A
 total of thirty basidiospores, basidia, cystidia and hyphae were measured
 from each collection. For basidiospores, the abbreviation “n/m/p”
 indicates n basidiospores measured from m fruit bodies of p collections.
 Dimensions for basidiospores are given using length × width (L × W) and
 extreme values are given in parentheses. The range contains a minimum of
 90% of the values. Measurements include arithmetic mean of spore length
 and width for all spores measured, Q means spore length divided by spore
 width and avQ indicates average Q of all spores ± standard deviation.
 Total Genomic DNA was extracted from basidioma gills following a modified
 CTAB extraction method (Bruns 1995). We targeted two DNA regions: (1) the
 internal transcribed spacer regions of nuclear ribosomal DNA (ITS); (2)
 partial large subunit of nuclear ribosomal DNA (LSU). The following primer
 pairs were used for the DNA amplifications: ITS1F–ITS4 for ITS (White et
 al. 1990; Gardes & Brun 1993), LR0R–LR5 for LSU (Vilgalys and Hester
 1990). Polymerase chain reactions (PCR) were performed in 25 μl volume
 reactions. Visualisation of PCR products were accomplished using SYBR
 Green and 1.5% agarose gels with TAE buffer for gel electrophoresis.
 Successful amplicons were purified by enzymatic purification using
 Exonuclease I and Shrimp Alkaline Phosphatase enzymes (Werle et al. 1994).
 After sequencing of Purified products, sequence chromatograms were
 trimmed, edited and assembled using Sequencher 4.1 (GeneCodes, Ann Arbor,
 MI). Once sequences were assembled and edited, they were deposited in
 GenBank ([http://www.ncbi.nlm.nih.gov](http://www.ncbi.nlm.nih.gov)). ###
 Files and variables #### File: ITS_LSU.phy **Description:**  Alignement
 file for Maximum Likelihood Phylogram of *Inocybe hazarensis* &
 *Inocybe shimlaensis*, inferred from combine ITS & LSU dataset with
 *Inosperma geminum* (OR823936 & OR823937) used as out-group. Bootstrap
 support values (≥ 50 %) are indicated above the nodes. The sequence
 generated in this study are represented in boldface. ## Code/software
 BioEdit was used for the analysis.