PHENOTYPIC AND MOLECULAR CHARACTERIZATION OF THE LAND RACES OF PEARS CULTIVATED IN NORTHERN PAKISTAN MOHAMMAD ISLAM DEPARTMENT OF BOTANY HAZARA UNIVERSITY MANSEHRA 2015 HAZARA UNIVERSITY MANSEHRA Department of Botany PHENOTYPIC AND MOLECULAR CHARACTERIZATION OF THE LAND RACES OF PEARS CULTIVATED IN NORTHERN PAKISTAN BY Mohammad Islam This research study has been conducted and reported as partial fulfillment for the requirements of PhD degree in Botany awarded by Hazara University Mansehra, Pakistan The Tuesday 15, October 2014 CONTENTS 2. 6. RNA Analyses ................................................................................................................. 29 2. 6.1 Amplification of RNA ................................................................................................ 29 2. 6. 2Electrophoresis and Visualization of DNA ............................................................. 32 2.6.3. 1X TAE Buffer (pH. 8.4) ........................................................................................... 32 2. 6. 4. Ethedium Bromide Solution .................................................................................... 32 2. 6. 4 Preparation of Gel Casting Tray .............................................................................. 32 2. 6. 5 Preparation of Gel Solution ...................................................................................... 32 2. 6. 6 Gel Casting……….................................................................................................... 33 2. 6. 7 Gel Electrophoresis .................................................................................................... 33 2. 6. 8 Visualization of Gel/DNA and Photography ....................................................... 33 2. 6. 9 Extraction of DNA from the Gel ............................................................................ 33 2. 6. 10 Confirmation of DNA purification ...................................................................... 35 2.7. Data Analyses .................................................................................................................... 35 2.8. The rRNA Sequencing ..................................................................................................... 37 2.9. Consensus Sequence Development ................................................................................ 38 Chapter 3 ....................................................................................................................................... 39 RESULTS ....................................................................................................................................... 39 3.1 NUMERICAL TRAIT ANALYSIS.................................................................................... 39 3.1.1PETIOLE LENGTH…… .............................................................................................. 39 3. 1. 2LEAF AREA………. .................................................................................................... 39 3. 1.3 PEDICEL LENGTH ................................................................................................... 40 3.1.4 FRUIT LENGTH……................................................................................................... 40 3.1.5 FRUIT WIDTH…….. ................................................................................................... 40 3.1.6 FRUIT WEIGHT…… ................................................................................................... 41 3.2 TAXONOMIC DESCRIPTION OF THE TAXA ............................................................. 50 A. ORIENTAL PEARS ............................................................................................................. 51 3. 2. 1 Pyrus pashia Buchanan-Hamilton ex D. Don, ........................................................ 51 3.2.2. Pyrus bretschneideri Rehder…………………………………………………………55 3.2.4 Pyrus calleryana Decaisne………………………………………………………........69 B. OCCIDENTAL PEARS ........................................................................................................ 61 3.2.6 Pyrus communis L….. .................................................................................................. 63 3.2.7 Pyrus sinkiangensis …………………………………………………………………...66 3.2.8 Pyrus hopeienses …….................................................................................................... 69 3.2.9 Pyrus armeniacaefolia…... ............................................................................................. 71 3.2.10 Pyrus serrulata……. .................................................................................................... 73 3.2.11 Pyrus ovoidea Rehder .................................................................................................. 75 3.2.13 Pyrus ussuriensis………... .......................................................................................... 79 3. 2. 14 Pyrus xerophila ……………………………………………………………………..81 3.3.1 DNA Extraction……................................................................................................... 82 3.3.2 Amplification ofRAPD primers ................................................................................. 84 3.3.3 Primers specificity… .................................................................................................. 90 3.3.4 Similarities among the land races.............................................................................. 92 3.3.5Phylogenetic Relationship of Pyrus Land races ....................................................... 95 3. 4. Nucleotide Sequencing .................................................................................................. 100 3. 4.1 Kacha Tora Tangai ………………………………………………………………...100 3.4.2 Zira Mamoothay…… ................................................................................................ 103 3. 4. 3. Khan Tango (Kt)….. ................................................................................................ 106 3. 4. 4. Ghata Zira Tangai.. ................................................................................................. 109 3. 4. 5. Nashpati…………. .................................................................................................. 111 3. 4. 6 Parawoo Tango…… ................................................................................................ 114 3. 4. 7 Pekhawry Tango…. ................................................................................................. 117 3.4. 8 Nak Tango………… ................................................................................................ 120 3. 4. 9 Spina Mamothy…… ................................................................................................ 123 3. 4. 10 Asmasy Tango….. .................................................................................................. 126 3. 4. 11 Mamosay Batal-8.. .................................................................................................. 129 3. 4. 12 Mamosay Bat_12… ................................................................................................ 132 3. 4. 13 Mamoosay Bat_14 ................................................................................................ 135 3. 4. 14 Mamoosay B15….. ................................................................................................. 138 3. 4. 15 Gultar Tango……. ................................................................................................. 141 3. 4. 16 Hary Tango Batal.. ................................................................................................. 144 3. 4. 17 Kado Batang (Kb). .................................................................................................. 147 3. 4. 18 Malyzay Tango…. .................................................................................................. 150 3. 4. 19 Mamosranga…… ................................................................................................. 153 3. 4. 20 Guraky Tango….................................................................................................... 156 3. 4. 21 Ghata Tora Tangai (Gtt) ....................................................................................... 159 3. 4. 22 Shaker Tango…… ................................................................................................. 162 3. 4. 23 ―Pak-24‖………… ................................................................................................. 165 3. 4. 24 Shaker Tango……. ................................................................................................ 168 Chapter 4 ..................................................................................................................................... 170 DISCUSSION ............................................................................................................................. 170 4.1. Numerical variations ...................................................................................................... 170 4.1.1 Petiole length…….. .................................................................................................. 170 4.1.2 Leaf area…………. .................................................................................................. 171 4.1.3 Pedicel length…….. ................................................................................................. 171 4.1.4 Fruit length ..............................................................................................................