Toward Identification of Black Lemma and Pericarp Gene Blp1 in Barley Combining Bulked Segregant Analysis and Specific-Locus Amplified Fragment Sequencing

Abstract

Black barley is caused by phytomelanin synthesized in lemma and/or pericarp and the trait is controlled by one dominant gene Blp1. The gene is mapped on chromosome 1H by molecular markers, but it is yet to be isolated. Specific-locus amplified fragment sequencing (SLAF-seq) is an effective method for large-scale de novo single nucleotide polymorphism (SNP) discovery and genotyping. In the present study, SLAF-seq with bulked segregant analysis (BSA) was employed to obtain sufficient markers to fine mapping Blp1 gene in an F2 population derived from Hatiexi No.1 × Zhe5819. Based on SNP screening criteria, a total of 77,542 polymorphic SNPs met the requirements for association analysis. Combining two association analysis methods, the overlapped region with a size of 32.41 Mb on chromosome 1H was obtained as the candidate region of Blp1 gene. According to SLAF-seq data, markers were developed in the target region and were used for mapping the Blp1 gene. Linkage analysis showed that Blp1 co-segregated with HZSNP34 and HZSNP36, and was delimited by two markers (HZSNP35 and HZSNP39) spanning 8.1 cM in 172 homozygous yellow grain F2 plants of Hatiexi No.1 × Zhe5819. More polymorphic markers were screened in the reduced target region and were used to genotype the population. As a result, Blp1 was delimited within a 1.66 Mb on chromosome 1H by the upstream marker HZSNP63 and the downstream marker HZSNP59. Our results demonstrated the utility of SLAF-seq-BSA approach to identify the candidate region and discover polymorphic markers at the specific targeted genomic region.

Study on Hybrid Characteristics of Medicinally Used Cultivated Codonopsis Species Using Ribosomal Internal Transcribed Spacer (ITS) Sequencing

“…A large number of molecular marker techniques, such as simple sequence repeat (SSR), single nucleotide polymorphism (SNP), and internal transcribed spacer (ITS), have been used in plant classification, identification, and analysis of genetic characteristics [ 6 , 7 , 8 , 9 , 10 , 11 , 12 , 13 , 14 , 15 ]. Among them, ITS sequence of nuclear ribosomal DNA (nrDNA) has been proved to be a helpful non-coding marker to infer hybridization events [ 5 , 16 , 17 , 18 , 19 , 20 , 21 , 22 ].…”

Physiological characterization and gene mapping of a novel cuticular wax-related mutant in barley

“…However, the 9 intervals were as long as 15.10 Mb, which was larger than the candidate region of 0.19 MB and 0.24 MB in cucumber and rice, respectively [23,24]. Similarly, the black lemma and pericarp gene in barley is also initial mapped to a region of 32.41 MB using the SLAF-BSA method [26], which could be attributed to the genomes size and sequence structure. The barley genome is ~ 5.3 Gb and contains many repeat sequences [17].…”

“…The combination of SLAF-seq and BSA has been proved to be effective for identifying major QTLs or candidate gene. This combination has been successfully used in rice [23], cucumber [24], barley [25,26], wheat [27], tomato [28], and pepper [29].…”

Molecular Mechanism of Functional Ingredients in Barley to Combat Human Chronic Diseases

“…The flavanone-3-hydroxylase locus completely cosegregated with the barley Ant17 position on the chromosome arm 2HL [47]. One dominant gene Blp1 for synthesized phytomelanin of the lemma and pericarp in black barley cosegregated with a 1.66 Mb between HZSNP34 and HZSNP36 on chromosome 1H [48].…”