Abstract
• Premise of the study: We developed microsatellite markers in the legume Lens culinaris from publicly available databases to enrich the limited marker resource available for the crop.
• Methods and Results: Eighty-two primer sets were identified using expressed sequence sets of L. culinaris available in the National Center for Biotechnology Information (NCBI) database and were characterized in six species of Lens. Among them, 20 simple sequence repeat (SSR) primers produced no amplification product, 43 produced monomorphic products, and 19 were polymorphic. The primers amplified mono-, di-, tri-, tetra-, penta-, and hexanucleotide repeats with one to four alleles. These SSR loci successfully amplified in five related wild species, with a total of 61 primer pairs in L. nigricans and L. odemensis (98.39%), 59 in L. tomentosus (95.1%), and 60 in L. ervoides and L. orientalis (96.7%), respectively.
• Conclusions: The microsatellite markers discovered in this study will be useful in genetic mapping, marker-assisted breeding, and characterization of germplasm.
Keywords: EST-SSRs, Fabaceae, Lens culinaris, microsatellites
Lentil (Lens culinaris Medik. subsp. culinaris) is a self-pollinated crop (2n = 2x = 14) belonging to the Vicieae tribe in the Fabaceae family. Lentil is a rich source of protein and micronutrients and is grown mainly in the Indian subcontinent, Middle East, North Africa, southern Europe, North and South America, Australia, and West Asia. Lentil has varied uses for consumption as a main dish, salads, or infant foods, and mixed with cereals to make bread and cakes. Although lentil is a highly nutritious food legume, its conservation and breeding potential is largely limited by a lack of molecular markers available for the crop.
Microsatellites are well-known genetic markers because of their codominant inheritance, polymorphism, and abundant coverage. They have become the markers of choice for many crops for studying genetic relatedness, diversity analysis, and constructing framework genetic maps. A limited number of microsatellite markers (approx. 100) have been published in L. culinaris to date. Hamwieh et al. (2005) developed 35 simple sequence repeat (SSR) markers, but these have reported no amplification or limited polymorphism, creating a major bottleneck to gene tagging and mapping studies in this crop. Kaur et al. (2011) developed 51 SSRs in L. culinaris, but these are less polymorphic and not sufficient to be used for genetic studies or for marker-assisted selection. An effective strategy for enrichment of microsatellite markers is the screening of expressed sequence tags, thereby reducing the time and cost for microsatellite development. The objective of this study was to develop new microsatellite markers using this strategy and characterize them in 18 L. culinaris accessions. The amplification success of these markers was also investigated in five wild Lens Mill. species (L. nigricans (M. Bieb.) Godron, L. odemensis Ladiz., L. tomentosus Ladiz., L. ervoides (Brign.) Grande, L. orientalis Popow) for potential genetic application or improvement of cultivated lentil (L. culinaris).
METHODS AND RESULTS
A total of 9513 L. culinaris expressed sequence tags (ESTs) were downloaded from the dbEST/GenBank database as of 15 January 2012 (http://www.ncbi.nlm.nih.gov). The ESTs were trimmed for poly(A) tails, and vector sequence contamination were removed using the SeqClean program (Masoudi-Nejad et al., 2006). The trimmed EST sequences were assembled into unigenes with the Cap3 program (Huang and Madan, 1999) to reduce redundancy. The unigenes containing 951 contigs and 3092 singletons produced a total of 251 putative SSRs using the software Troll (Martins et al., 2009). The fragments with inappropriate flanking sequences or with less than 500 bp were excluded and 82 SSRs were designed using Primer3 software (Rozen and Skaletsky, 2000).
The newly identified microsatellite markers were screened on 32 individuals representing six Lens species including 18 accessions from cultivated species and 14 wild accessions comprising five species. Voucher specimens were sourced from the International Center for Agricultural Research in the Dry Areas (ICARDA), Aleppo, Syria, and the lentil breeding program at the Indian Agricultural Research Institute (IARI), New Delhi, India (Appendix 1). The genomic DNA from each individual was isolated using the cetyltrimethylammonium bromide (CTAB) method as described by Murray and Thompson (1980). PCR mixtures of 20 μL consisted of 2.0 μL 10× buffer (100 mM Tris-HCl, 500 mM KCl, 15 mM MgCl2, 0.01% gelatin), 200 μM each dNTP, 0.5 μM each of forward and reverse primers, 1 U Taq DNA polymerase (PCR reagents and primers procured from Sigma-Aldrich, St. Louis, Missouri, USA), and ∼40 ng DNA and were performed in a Veriti Thermal Cycler (Applied Biosystems, Life Technologies, Singapore). The PCR protocol consisted of one denaturation cycle at 94°C for 4 min followed by 30 cycles of 94°C for 1 min, annealing at 59–62°C (depending upon the primer) for 30 s, extension at 72°C for 1 min, and a final extension at 72°C for 8 min. The amplification fragments were separated on 3% MetaPhor Agarose gels (Lonza, Rockland, Maine, USA) and visualized by ethidium bromide staining. The band size was obtained in comparison to a 100-bp DNA ladder (MBI, Fermentas, Vilnius, Lithuania). Among these 82 SSRs, 62 amplified successfully in most of the species, providing an amplification success rate of 75.6%, and 19 of them showed more than two clear scorable bands in the Lens species (Table 1). Primer sequences and related information for 43 monomorphic EST-SSR primers are available as Appendix 2. The conservation of SSRs across species has been validated by sequencing one amplicon from each species. The EST-SSR markers amplified one to four alleles among the six species. The expected heterozygosity was determined on the basis of the number of genotypes amplified per species and ranged from 0 to 0.875 (Table 2). The putative functions of SSR-associated unigenes were determined by using BLASTX (Altschul et al., 1997) against the nonredundant GenBank database.
Table 1.
Characteristics of the 19 polymorphic EST-SSRs identified in Lens culinaris.
| No. | EST-SSR locus | GenBank EST no. (Probe DB_id) | Primer sequences (5′–3′) | Repeat motif | Ta (°C) | Allele size range (bp) | Putative function (organism) | BLASTX E-value |
| 1 | PLC5 | GT626272 (16537804) | F: CATTGCAGCTTATTCTCACAGC | (CAATGG)5 | 60 | 320–360 | Auxin-induced protein 5NG4-like (Glycine max) | 2E-41 |
| R: TGACCCATCCTCATCCTTAAAT | ||||||||
| 2 | PLC10 | GT62175 (16581945) | F: TGCAACAAAGGACACTAGAGGTT | (AT)6 | 59 | 279–328 | Predicted aspartic proteinase nepenthesin-1 like protein (Glycine max) | 1E-157 |
| R: ATTTCTTTCTCCCTAACCAGCC | ||||||||
| 3 | PLC16 | GT627608 (16581946) | F: CGTTTGATCTTCTAAGCCCCTA | (T)10 | 59 | 255–270 | Uncharacterized protein (Glycine max) | 2E-10 |
| R: AAGGGAAAGGATGTTTGACTTG | ||||||||
| 4 | PLC17 | GT624932 (16581947) | F: AAGCTGAAGGAAATCAAAGTGG | (TCTTT)3 | 59 | 315–335 | Peptidyl prolyl cis-trans isomerase (Arabidopsis thaliana) | 1E-118 |
| R: TCAACACACTCCATGTTTAGAGC | ||||||||
| 5 | PLC21 | GT626993 (16581949) | F: AACTCGCATCCTCTTCACAACT | (TTC)6 | 60 | 264–286 | Glutathione peroxidase (Medicago truncatula) | 5E-112 |
| R: GGACCTTTCCCTTGTAGTCACC | ||||||||
| 6 | PLC22 | GT626865 (16581950) | F: TACACTGAAGGAGATGCACTGG | (T)11 | 60 | 279–290 | DNA-directed RNA polymerase I, II, and III subunits (Medicago truncatula) | 1E-43 |
| R: TAACAACAAAACACAGCTTCGC | ||||||||
| 7 | PLC30 | GT625366 (16581951) | F: TTGGTCAGGTTCTCAATCCTCT | (T)10 | 61 | 243–257 | Uncharacterized protein (Glycine max) | 3E-39 |
| R: ACGGATGAACGCTTGTAAAGAA | ||||||||
| 8 | PLC35 | GT619232 (16581952) | F: TTGCTTCCTCCTCTTCTCACTC | (T)10 | 60 | 260–277 | GDP-l-galactose phosphorylase-1 like (Glycine max) | 7E-141 |
| R: AGCCTCAGTACCCTCCTCTTTT | ||||||||
| 9 | PLC38 | GT626497 (16581953) | F: CCTGGAGAAGTCTGTGGAAGAT | (TTTGT)3 | 59 | 309–334 | LEA protein (Medicago truncatula) | 4E-65 |
| R: AGCTCTAGCATTTTGCATGTGA | ||||||||
| 10 | PLC39 | GT624018 (16581954) | F: CAGAGAAATCCCCTGCTGAG | (AAG)5 | 62 | 158–178 | 5′-adenylylsulfate reductase 3 (Glycine max) | 1E-177 |
| R: CATGATTCCCATAGCCTTGC | ||||||||
| 11 | PLC42 | GT626865 (16581955) | F: AACCAATCATGGCTTCTGCT | (GA)8 | 60 | 183–210 | BZIP transcription factor ATB2 (Medicago truncatula) | 6E-78 |
| R: TTTCACCGTCTTTATGAACCA | ||||||||
| 12 | PLC46 | GT624901 (16581956) | F: CAAACTGGAAGATGCTGCTG | (CAATGG)3 | 61 | 192–220 | Auxin-induced protein 5NG4-like (Medicago truncatula) | 2E-71 |
| R: TGACCCATCCTCATCCTTAAA | ||||||||
| 13 | PLC51 | GT624642 (16581957) | F: CCATGATGAGCCTTGAATGA | (GAA)10 | 62 | 125–143 | Peroxidase (Medicago truncatula) | 0.0 |
| R: TCTTCAATCTCCAGGAACACTTT | ||||||||
| 14 | PLC60 | GT624076 (16581958) | F: TGCTTGGACCCTAAATTTGC | (TA)6 | 60 | 130–145 | Cysteine proteinase inhibitor (Medicago truncatula) | 4E-131 |
| R: AAGAAAAGGGCAACCACTGA | ||||||||
| 15 | PLC63 | GT619353 (16581959) | F: TTGATGGCTATGGGAGTGGT | (TTA)8 | 60 | 175–189 | Early nodulin-like protein (Medicago truncatula) | 1E-51 |
| R: TGGTCCCAACAAAATACCAA | ||||||||
| 16 | PLC70 | GT618700 (16581960) | F: CATCTCTTCGTGGCGTAAT | (GTT)9 | 60 | 179–195 | Albumin-2 (Pisum sativum) | 6E-159 |
| R: AGCAAACAACAGCACACATA | ||||||||
| 17 | PLC74 | GT624794 (16581961) | F: GATTTACCGATGGATCTTCA | (TTA)6 | 61 | 168–191 | Xylose isomerase (Medicago truncatula) | 5E-68 |
| R: CTAAGGGAGAGAAAGAAAAGG | ||||||||
| 18 | PLC81 | GT621832 (16581962) | F: GGGTAGAGTATTATTGAAGGTGG | (TA)6 | 61 | 182–209 | Chromo-domain–containing protein LHP1 (Medicago truncatula) | 8E-34 |
| R: AGAATCGCTAGTTTAGAGCAAG | ||||||||
| 19 | PLC82 | GT621329 (16581963) | F: CACCAATCTTCACTTCACTTTC | (GAA)4 | 60 | 178–200 | Legumin protein (garden pea) | 5E-121 |
| R: CAAGTACAAGGACTGACTAGGG |
Note: Ta = annealing temperature.
Table 2.
Total number of alleles (A) and expected heterozygosity (He) of EST-SSRs in six Lens species.
| EST-SSR locus | L. culinaris Medik. (n = 18) | L. nigricans (M. Bieb.) Godron (n = 3) | L. odemensis Ladiz. (n = 3) | L. tomentus Ladiz. (n = 2) | L. ervoides (Brign.) Grande (n = 4) | L. orientalis Popow (n = 2) | ||||||
| A | He | A | He | A | He | A | He | A | He | A | He | |
| PLC5 | 3 | 0.494 | 2 | 0.444 | 1 | 0.000 | 2 | 0.500 | 2 | 0.688 | 2 | 0.500 |
| PLC10 | 3 | 0.586 | 2 | 0.444 | 2 | 0.444 | 1 | 0.000 | 2 | 0.375 | 1 | 0.000 |
| PLC16 | 4 | 0.799 | 2 | 0.444 | 1 | 0.000 | 2 | 0.500 | 2 | 0.500 | 1 | 0.000 |
| PLC17 | 2 | 0.198 | 2 | 0.444 | 2 | 0.444 | 1 | 0.000 | 2 | 0.375 | 2 | 0.500 |
| PLC21 | 4 | 0.722 | 1 | 0.000 | 2 | 0.444 | 1 | 0.000 | 2 | 0.375 | 1 | 0.000 |
| PLC22 | 3 | 0.667 | 2 | 0.444 | 2 | 0.444 | 2 | 0.500 | 3 | 0.625 | 2 | 0.500 |
| PLC30 | 3 | 0.648 | 2 | 0.444 | 1 | 0.000 | 1 | 0.000 | 2 | 0.500 | 1 | 0.000 |
| PLC35 | 4 | 0.667 | 2 | 0.444 | 1 | 0.000 | 2 | 0.500 | 2 | 0.375 | 2 | 0.500 |
| PLC38 | 4 | 0.722 | 2 | 0.444 | 2 | 0.444 | 1 | 0.000 | 2 | 0.375 | 2 | 0.500 |
| PLC39 | 3 | 0.537 | 2 | 0.444 | 3 | 0.667 | 1 | 0.500 | 2 | 0.500 | 2 | 0.500 |
| PLC42 | 3 | 0.648 | 2 | 0.444 | 1 | 0.000 | 1 | 0.000 | 2 | 0.375 | 2 | 0.500 |
| PLC46 | 3 | 0.046 | 3 | 0.667 | 4 | 0.222 | 2 | 0.500 | 3 | 0.438 | 1 | 0.000 |
| PLC51 | 3 | 0.278 | 2 | 0.444 | 3 | 0.667 | 2 | 0.500 | 2 | 0.375 | 2 | 0.500 |
| PLC60 | 4 | 0.747 | 2 | 0.444 | 2 | 0.444 | 1 | 0.000 | 2 | 0.625 | 1 | 0.000 |
| PLC63 | 4 | 0.623 | 3 | 0.667 | 1 | 0.000 | 2 | 0.500 | 3 | 0.375 | 1 | 0.000 |
| PLC70 | — | — | 2 | 0.444 | 1 | 0.000 | — | — | 2 | 0.875 | 1 | 0.000 |
| PLC74 | 4 | 0.747 | 3 | 0.667 | 1 | 0.000 | 2 | 0.500 | 3 | 0.625 | 2 | 0.500 |
| PLC81 | 4 | 0.386 | 2 | 0.778 | 3 | 0.667 | 1 | 0.000 | 1 | 0.000 | 1 | 0.000 |
| PLC82 | 4 | 0.574 | 2 | 0.444 | 2 | 0.444 | 1 | 0.000 | 3 | 0.625 | 1 | 0.000 |
Note: — = no amplification; n = number of accessions used for each species.
CONCLUSIONS
The EST-SSR markers identified and characterized in this study have enriched the limited microsatellite marker resources in Lens species. The markers developed will be helpful in saturating Lens genetic maps and for tagging and mapping of genes and quantitative trait loci associated with important traits to be further used in marker-assisted breeding for enhancing productivity and quality. These markers would also be helpful in studying genetic diversity and detecting interspecies polymorphisms for marker-based introgression of genes from related species.
Appendix 1.
Voucher information for Lens species used for the EST-SSR polymorphism study.
| Species | Country of origin/source | Voucher accession no. | Specimens collected/maintained |
| Lens culinaris Medik. | Turkey | IG-70208, IG-70211, IG-112, IG-115, IG-12 | ICARDA, Aleppo, Syria |
| Ethiopia | IG-208, IG-69502, IG-69513, IG-69517, IG-69522, IG-70174 | ICARDA, Aleppo, Syria | |
| India | L4149, PL08, L9-12, L830, L4603 | NBPGR and IARI, New Delhi, India | |
| Syria | FLIP2004-7L, ILL4605 | ICARDA, Aleppo, Syria | |
| L. nigricans (M. Bieb.) Godron | Syria | ILWL-111, ILWL-22, ILWL-445 | ICARDA, Aleppo, Syria |
| L. odemensis Ladiz. | Syria | ILWL-254, ILWL-35, ILWL-153 | ICARDA, Aleppo, Syria |
| L. tomentosus Ladiz. | Syria | ILWL-91, ILWL-93 | ICARDA, Aleppo, Syria |
| L. ervoides (Brign.) Grande | Syria | ILWL-126, ILWL-206, ILWL-139, ILWL-393 | ICARDA, Aleppo, Syria |
| L. orientalis Popow | Syria | ILWL-7, ILWL-81 | ICARDA, Aleppo, Syria |
Note: IARI = Indian Agricultural Research Institute; ICARDA = International Center for Agricultural Research in Dry Areas; NBPGR = National Bureau of Plant Genetic Resources.
Appendix 2.
Primer sequences and characteristics of the 43 monomorphic EST-SSR markers in Lens.a
| No. | EST-SSR locus | Primer sequences (5′–3′) | Repeat motif | Ta (°C) | Allele size (bp) | Putative function (organism) | BLASTX E-value |
| 1 | PLC2 | F: TTGACTGTTCTGGCGTTTTCTA | (T)19 | 56 | 330 | Peptidyl-prolyl cis-trans isomerase (Medicago truncatula) | 5E-101 |
| R: TGCACCATCTTTTGCCTACATA | |||||||
| 2 | PLC4 | F: CCTATCGGGAAACTACATGGAA | (GGCAGC)3 | 58 | 359 | Calnexin-like protein (Zea mays) | 1E-157 |
| R: TCTGCATTGGTCTTCTTCTCAA | |||||||
| 3 | PLC6 | F: ATCAAGTTAGGGACGATTGGAA | (GTA)6 | 56 | 185 | Hypothetical protein MTR_5g092090 (Medicago truncatula) | 2E-65 |
| R: TGGTTGTAGTCTTTTAGGGTTTGC | |||||||
| 4 | PLC7 | F: GCTTTTATGATCTTCTCGTGGT | (GAAT)4 | 56 | 184 | Chitinase domain-containing protein 1-like (Glycine max) | 6E-17 |
| R: CGAGGATTACTTTTCAATGGTC | |||||||
| 5 | PLC8 | F: CTCCTTCCATTTCTCTTTCTGC | (TTC)6 | 58 | 158 | Uncharacterized protein (Glycine max) | 1E-108 |
| R: TCCTGAACGACACCAACACTAC | |||||||
| 6 | PLC9 | F: ATGTGGATACGTCAGAAACCCT | (TATCTA)4 | 56 | 348 | Glycinin subunit G7 (Glycine max) | 8E-56 |
| R: TCGAGAACTGGGAGAGTCAAAT | |||||||
| 7 | PLC11 | F: GTTTGTTTGGTTTGACTGGGAT | (TTA)6 | 58 | 185 | Histone H1 (Pisum sativum) | 1E-35 |
| R: TTAGGAACGGTGTCGAGTACAA | |||||||
| 8 | PLC12 | F: GGAAGCAAGATGGAAGAAGTTG | (T)11 | 60 | 146 | Heat-shock protein (Medicago truncatula) | 2E-49 |
| R: GCGCCATTAGTGCAGAGTAAAT | |||||||
| 9 | PLC13 | F: TCACCATTTTGGGTTATCTTCC | (T)13 | 56 | 211 | Hydrophobic protein LTI6B-like (Cicer arietinum) | 1E-17 |
| R: AGCTTCACACTATCAATTCCACAC | |||||||
| 10 | PLC14 | F: TCTGGAAGAGGGTTTGTACCAT | (T)12 | 56 | 210 | Uncharacterized protein (Cicer arietinum) | 1E-66 |
| R: GCAGTTAGATCACAGCTACCAAAA | |||||||
| 11 | PLC15 | F: CCAGTAAAAGAGCTTGCATTCC | (A)10 | 58 | 345 | Vicilin precursor (Vicia faba) | 1E-179 |
| R: AGAAAAGAGTTGCAGAGAAGCG | |||||||
| 12 | PLC18 | F: GGACCATCAACTAGCACATGAC | (GAA)6 | 56 | 382 | Peroxidase (Medicago truncatula) | 1E-91 |
| R: TCACATCATCAACATGCTCAAC | |||||||
| 13 | PLC25 | F: GTTGCAGAAAATGTAACTGCGT | (A)12 | 56 | 396 | Uncharacterized mRNA (Glycine max) | 8E-44 |
| R: ACAATGAGAGGCCAGTGCTTA | |||||||
| 14 | PLC28 | F: CAAGGTTGGAAAAGACAAGAGG | (A)18 | 60 | 398 | 60S ribosomal protein L36 (Medicago truncatula) | 2E-68 |
| R: TTTGGAGCTAGACTTCGCATTT | |||||||
| 15 | PLC40 | F: CAACTCGCATCCTCTTCACA | (TTC)6 | 60 | 163 | Glutathione peroxidase (Medicago truncatula) | 2E-108 |
| R: CAAAGGGGTTGGAGTCGTAA | |||||||
| 16 | PLC41 | F: TTTGTTGATGTTGTTGGCGT | (T)12 | 60 | 164 | Arabinogalactan peptide 16-like (Cicer arietinum) | 1E-12 |
| R: CTCCTCCGCGTTCTACAAAC | |||||||
| 17 | PLC48 | F: TGTGGTACATGCACACCAAAT | (ACC)5 | 58 | 168 | Proline-rich protein (Medicago truncatula) | 1E-27 |
| R: GGTGGTAGCAGTGGTGGAGT | |||||||
| 18 | PLC49 | F: TTGTTTTGAGAACCTTCCCC | (T)28 | 58 | 200 | Hypothetical protein PRUPE_ppa010183mg (Prunus persica) | 3E-35 |
| R: TTTTGCAAGGGTATTTCTTTTTG | |||||||
| 19 | PLC50 | F: CGATTGGTCTTATATGGTTCTG | (ATGTA)4 | 60 | 172 | Peptide transporter PTR3-A (Medicago truncatula) | 9E-41 |
| R: AAGCTACCTGCATACTTGGTC | |||||||
| 20 | PLC52 | F: CGTTTGATCTTCTAAGCCCC | (T)10 | 58 | 198 | Uncharacterized protein (Glycine max) | 3E-10 |
| R: TCGGCACATTGTTGAAAAGA | |||||||
| 21 | PLC53 | F: TCGTGATAAAAACGGGGAAG | (GAA)5 | 56 | 200 | BRI1-KD interacting protein (Medicago truncatula) | 2E-94 |
| R: TATCTTTGCCACTGCCTCCT | |||||||
| 22 | PLC54 | F: GTAAACGAAGCTCAGAGCCG | (GGA)5 | 56 | 200 | Glycine-rich RNA-binding protein (Medicago truncatula) | 3E-48 |
| R: CATATCCACGATCCCTGCTT | |||||||
| 23 | PLC55 | F: AGACACCGGCATCAAATCAT | (A)10 | 60 | 173 | Acylamino-acid-releasing enzyme (Medicago truncatula) | 3E-111 |
| R: CATATTCAAATATTCAGTGTCATGTTC | |||||||
| 24 | PLC57 | F: GGAAGTGATTGTGGTTTTTAATCA | (A)17 | 60 | 182 | WD repeat-containing protein 26-like (Glycine max) | 8E-45 |
| R: ATTGCTCATTCCCACCAAAG | |||||||
| 25 | PLC58 | F: TGGAAGAAAGAGAAGGGCAA | (T)12 | 60 | 138 | Putative zinc finger protein (Arabidopsis thaliana) | 1E-102 |
| R: CACAGCTACCAAAAATCAGTTCC | |||||||
| 26 | PLC59 | F: TTGTTTAGCTGGTGTGGTTTTC | (A)18 | 56 | 180 | F-box protein SKP2B-like (Glycine max) | 1E-46 |
| R: CTACAGCACGTTTGCAAGGA | |||||||
| 27 | PLC61 | F: ACTAGGAAAGGAAAACGGCG | (TC)26 | 56 | 145 | No significant similarity | — |
| R: GAGTGACACGTGAATGGTGG | |||||||
| 28 | PLC62 | F: GCAAAGAACAAGAATAACGTGG | (AAAC)4 | 56 | 126 | Beta-1,3-galactosyltransferase 2-like isoform 1 (Glycine max) | 1E-91 |
| R: CAAACCGAAGAATAAGAGAGGG | |||||||
| 29 | PLC64 | F: CAAACTCTTCACCGACACGC | (TCTTC)5 | 60 | 181 | Bcr-associated protein (BAP) putative (Ricinus communis) | 4E-83 |
| R: AACGAGGGTTAGGATGAGAAGC | |||||||
| 30 | PLC65 | F: TGTTGCAATGCTTTTAGCCT | (A)11 | 56 | 165 | 40S ribosomal protein SA (Medicago truncatula) | 3E-110 |
| R: CAGAAGCTTTTCGGTGTTCC | |||||||
| 31 | PLC66 | F: ATTTGGAGCAAAGATGCAGG | (A)10 | 56 | 200 | d-tyrosyl-tRNATyr deacylase-like (Glycine max) | 1E-69 |
| R: GGATCGACCTCCAATCAAGA | |||||||
| 32 | PLC67 | F: GCATAATCAGTTTGTTTTTGCG | (A)23 | 58 | 190 | Cyclin-dependent kinases regulatory subunit 1-like (Cicer arietinum) | 3E-45 |
| R: TTCTGCAAAAGCTTCTGGGT | |||||||
| 33 | PLC68 | F: AAAAAGAGGCCATCATGTTCA | (A)18 | 56 | 156 | Ferritin (Pisum sativum) | 1E-48 |
| R: CAGCAGTGACGGCAATTTTA | |||||||
| 34 | PLC69 | F: CGCTCTACCAACAGCATAA | (CT)19 | 56 | 195 | No significant similarity | — |
| R: GAGGTCTCTTTTGTTCTTCACT | |||||||
| 34 | PLC71 | F: AGTGAGCAAGGAATAAAACG | (AG)38 | 58 | 276 | Legumin J acidic chain (Pisum sativum) | 0.0 |
| R: GAGTAGCAAGGAAAGTGAAAAC | |||||||
| 36 | PLC72 | F: TATGATGAAAGCCAGGACA | (TAT)8 | 58 | 142 | Aminocyclopropane-1-carboxylate oxidase (Pisum sativum) | 9E-180 |
| R: GACTGCACAATCTTAAACACC | |||||||
| 37 | PLC73 | F: GAAAGGAAAGGTTTTAGCTG | (AG)13 | 60 | 198 | 40S ribosomal protein S18 (Medicago truncatula) | 8E-91 |
| R: CTTTGATTGAGGTAAGAGCA | |||||||
| 38 | PLC75 | F: TCGTTCCATATCTGTGTTCA | (AATC)3 | 56 | 195 | Xylose isomerase (Medicago truncatula) | 5E-68 |
| R: GTAGCGAGATTCATACCTATCC | |||||||
| 39 | PLC76 | F: AGGAAGGTGGAGTTACGG | (CT)52 | 56 | 160 | Cyclin-like F-box (Medicago truncatula) | 6E-164 |
| R: AAACCTAGAAGTAAAGGGGAAG | |||||||
| 40 | PLC77 | F: GGAAAGAGCCAAGAAGTTG | (CAATGG)5 | 56 | 230 | Auxin-induced protein 5NG4-like (Glycine max) | 8E-70 |
| R: ACCCATCCTCATCCTTAAAT | |||||||
| 41 | PLC78 | F: CTATGACTGCTCAAACTCAAGA | (GAT)6 | 56 | 150 | Nascent polypeptide-associated complex subunit alpha-like (Medicago truncatula) | 1E-115 |
| R: CCTTCTACATCATCATCTTCCT | |||||||
| 42 | PLC79 | F: AATTTCTGGTGTTTCTGGTG | (GAT)7 | 58 | 165 | Translational elongation factor 1 subunit beta (Pisum sativum) | 2E-94 |
| R: TCTTCTCTTCCTCAGTCTCTTC | |||||||
| 43 | PLC80 | F: GCTAACAAACAACACCATGA | (GAA)10 | 58 | 150 | Peroxidase (Arabidopsis thaliana) | 3E-176 |
| R: GCATCTAAGTTCTTCAATCTCC |
Note : T a = annealing temperature.
Only polymorphic primers were submitted to GenBank, therefore GenBank IDs for monomorphic markers are not available.
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