Striking variation in chromosome structure within Musa acuminata and its diploid cultivars

The majority of cultivated bananas originated from inter- and intra(sub)specific crosses between two wild diploid species, Musa acuminata and Musa balbisiana. Hybridization and polyploidization events during the evolution of bananas led to the formation of clonally propagated cultivars characterized by a high level of genome heterozygosity and reduced fertility. The combination of low fertility of edible clones and differences in the chromosome structure among M. acuminata subspecies greatly hampers the breeding of improved banana cultivars. Using comparative oligo painting we investigated large chromosomal rearrangements in a set of wild M. acuminata subspecies and cultivars that originated by natural crosses. Additionally, we analyzed chromosome structure of F1 progeny that resulted from crosses between Mchare bananas and wild M. acuminata ‘Calcutta 4’ genotype. Analysis of chromosome structure within M. acuminata revealed the presence of a large number of chromosomal rearrangements s..., Analysis of proportion of individual parental subgenomes in the F1 hybrid clones was done using vcfHunter pipeline (https://github.com/SouthGreenPlatform/vcfHunter) according to Baurens et al. (2019). Briefly, trimmed reads were aligned to reference genome sequence of M. acuminata ssp. malaccensis ‘DH Pahang’ v4 (Belser et al., 2021) by BWA-MEM v0.7.15 (Li 2013), followed by removing redundant reads using MarkDuplicate from Picard Tools v2.7.0, and locally realigned around indels using the IndelRealigner tool of GATK v3.3 package (McKenna et al., 2010). Bases with a mapping quality ≥10 were counted using the process_reseq_1.0.py python script (https://github.com/SouthGreenPlatform/vcfHunter). Variant calling and SNP filtering steps were performed according to Baurens et al. (2019) using the VcfPreFilter.1.0 python script (alleles supported by at least three reads and with a frequency 0.25 were kept as variant) and vcfFilter.1.0.py python script (&lt;6-fold coverage for the minor allele ..., , # SNP datasets (vcf files) used for in silico painting of Mchare x M. acuminata 'Calcutta 4' F1 hybrid clones [https://doi.org/10.5061/dryad.44j0zpcnq](https://doi.org/10.5061/dryad.44j0zpcnq) Genomic DNA was isolated with the NucleoSpin PlantII kit (Macherey-Nagel, Düren, Germany) according to the manufacturer’s recommendations and further sheared by Bioruptor Plus (Diagenode, Liege, Belgium) to achieve an insert size of about 500 bp. Libraries for sequencing were prepared from 2 μg of fragmented DNA using TruSeq® DNA PCR-free kit (Illumina) and sequenced on a NovaSeq 6000 (Illumina), producing 2 × 150-bp paired-end reads to achieve a minimal sequence depth of 25 ×. Raw data were trimmed for low-quality bases and adapter sequences and to the same length using fastp v.0.20.1 (Chen et al., 2018). Analysis of proportion of individual parental subgenomes in the F1 hybrid clones was done using vcfHunter pipeline ([https://github.com/SouthGreenPlatform/vcfHunter](https://github.com/South...大部分栽培香蕉起源于两个野生二倍体物种——小果野芭蕉（Musa acuminata）和野芭蕉（Musa balbisiana）的种间及种内（亚种间）杂交。在香蕉演化过程中，杂交与多倍化事件催生了一批以高度基因组杂合性和育性降低为典型特征的无性繁殖栽培品种。可食用无性系的低育性，加之小果野芭蕉亚种间染色体结构的差异，极大阻碍了优良香蕉栽培品种的育种工作。 本研究采用比较寡核苷酸绘画技术（comparative oligo painting），对一系列野生小果野芭蕉亚种及自然杂交起源的栽培品种开展大规模染色体重排分析。此外，我们还分析了Mchare香蕉与野生小果野芭蕉‘Calcutta 4’基因型杂交获得的F1后代的染色体结构。对小果野芭蕉的染色体结构分析显示，其存在大量染色体重排…… 我们使用vcfHunter流程（https://github.com/SouthGreenPlatform/vcfHunter），参照Baurens等人（2019）的方法，分析F1杂种克隆的亲本亚基因组占比。具体步骤如下：将经过质量修剪的测序reads比对至小果野芭蕉马来西亚亚种（M. acuminata ssp. malaccensis）‘DH Pahang’ v4参考基因组（Belser等人，2021），比对工具选用BWA-MEM v0.7.15（Li，2013）；随后使用Picard Tools v2.7.0中的MarkDuplicate工具去除冗余reads，并通过GATK v3.3软件包的IndelRealigner工具在插入缺失（indel）区域进行局部重比对。使用process_reseq_1.0.py Python脚本统计比对质量≥10的碱基位点。变异检测与单核苷酸多态性（Single Nucleotide Polymorphism，SNP）过滤步骤参照Baurens等人（2019），分别通过VcfPreFilter.1.0 Python脚本（保留至少3条reads支持且等位基因频率为0.25的变异位点）与vcfFilter.1.0.py Python脚本（次要等位基因覆盖度<6倍……）完成。 # 用于Mchare与小果野芭蕉‘Calcutta 4’ F1杂种克隆计算机寡核苷酸绘画的SNP数据集（vcf格式文件） 可从https://doi.org/10.5061/dryad.44j0zpcnq获取。 基因组DNA提取采用NucleoSpin PlantII试剂盒（Macherey-Nagel，德国迪伦），操作流程严格遵循制造商说明书；随后使用Bioruptor Plus（Diagenode，比利时列日）将DNA片段化至约500 bp的插入片段大小。使用TruSeq® DNA PCR-free试剂盒（Illumina）从2 μg片段化DNA中制备测序文库，并在NovaSeq 6000（Illumina）平台完成测序，生成2×150 bp双端reads，以确保最低测序深度达25×。原始数据通过fastp v.0.20.1（Chen等人，2018）进行低质量碱基、接头序列修剪及长度均一化处理。 我们再次使用vcfHunter流程（https://github.com/SouthGreenPlatform/vcfHunter）分析F1杂种克隆的亲本亚基因组占比……