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. 2020 Oct 6;15(10):e0239275. doi: 10.1371/journal.pone.0239275

The R2R3-MYB gene family in banana (Musa acuminata): Genome-wide identification, classification and expression patterns

Boas Pucker 1, Ashutosh Pandey 1,2, Bernd Weisshaar 1, Ralf Stracke 1,*
Editor: Matthew J Hegarty3
PMCID: PMC7537896  PMID: 33021974

Abstract

The R2R3-MYB genes comprise one of the largest transcription factor gene families in plants, playing regulatory roles in plant-specific developmental processes, defense responses and metabolite accumulation. To date MYB family genes have not yet been comprehensively identified in the major staple fruit crop banana. In this study, we present a comprehensive, genome-wide analysis of the MYB genes from Musa acuminata DH-Pahang (A genome). A total of 285 R2R3-MYB genes as well as genes encoding three other classes of MYB proteins containing multiple MYB repeats were identified and characterised with respect to structure and chromosomal organisation. Organ- and development-specific expression patterns were determined from RNA-Seq data. For 280 M. acuminata MYB genes for which expression was found in at least one of the analysed samples, a variety of expression patterns were detected. The M. acuminata R2R3-MYB genes were functionally categorised, leading to the identification of seven clades containing only M. acuminata R2R3-MYBs. The encoded proteins may have specialised functions that were acquired or expanded in Musa during genome evolution. This functional classification and expression analysis of the MYB gene family in banana establishes a solid foundation for future comprehensive functional analysis of MaMYBs and can be utilized in banana improvement programmes.

Introduction

Banana (Musa spp.), including dessert and cooking types, is a staple fruit crop for a major world population, especially in developing countries. The crop is grown in more than 100 countries throughout the tropics and sub-tropics, mainly in the African, Asia-Pacific, and Latin American and Caribbean regions [1]. Bananas provide an excellent source of energy and are rich in certain minerals and in vitamins A, C and B6. Furthermore, this perennial, monocotyledonous plant provides an important source of fibre, sugar, starch and cellulose (used for paper, textiles). Bananas have also been considered as a useful tool to deliver edible vaccines [2]. Certain agronomic traits, such as stress and pest resistance as well as fruit quality, are thus of considerable interest. Banana improvement through breeding exercises has been challenging for various reasons. Therefore, genetic engineering-based optimisations hold great promise for crop improvement. For this purpose, candidate gene targets need to be identified. The release of a high quality banana genome sequence [3] provides an useful resource to understand functional genomics of important agronomic traits and to identify candidate genes to be utilized in banana improvement programmes.

Almost all biological processes in eukaryotic cells or organisms are influenced by transcriptional control of gene expression. Thus, the regulatory level is a good starting point for genetic engineering. Regulatory proteins are involved in transcriptional control, alone or complexed with other proteins, by activating or repressing (or both) the recruitment of RNA polymerase to promoters of specific genes [4]. These proteins are called transcription factors. As expected from their substantial regulatory complexity, transcription factors are numerous and diverse [5]. By binding to specific DNA sequence motifs and regulating gene expression, transcription factors control various regulatory and signaling networks involved in the development, growth and stress response in an organism.

One of the widest distributed transcription factor families in all eukaryotes is the MYB (myeloblastosis) protein family. In the plant kingdom, MYB proteins constitute one of the largest transcription factor families. MYB proteins are defined by a highly conserved MYB DNA-binding domain, mostly located at the N-terminus of the protein. The MYB domain generally consists of up to four imperfect amino acid sequence repeats (R) of about 50–53 amino acids, each forming three alpha–helices [summarised in 6]. The second and third helices of each repeat build a helix–turn–helix (HTH) structure with three regularly spaced tryptophan (or hydrophobic) residues, forming a hydrophobic core [7]. The third helix of each repeat was identified as the DNA recognition helix that makes direct contact with DNA [8]. During DNA contact, two MYB repeats are closely packed in the major groove, so that the two recognition helices bind cooperatively to the specific DNA recognition sequence motif. In contrast to vertebrates genomes, which only encode MYB transcription factors with three repeats, plants have different MYB domain organisations, comprising one to four repeats [6, 9]. R2R3-MYBs, which are MYB proteins with two repeats (named according to repeat numbering in vertebrate MYBs), are particularly expanded in plant genomes. Copy numbers range from 45 unique R2R3-MYBs in Ginkgo biloba [10] to 360 in Mexican cotton (Gossypium hirsutum) [11]. The expansion of the R2R3-MYB family was coupled with widening in the functional diversity of R2R3-MYBs, considered to regulate mainly plant-specific processes including secondary metabolism, stress responses and development [6]. As expected, R2R3-MYBs are involved in regulating several biological traits, for example wine quality, fruit color, cotton fibre length, pollinator preferences and nodulation in legumes.

In this study, we have used genomic resources to systematically identify members of the M. acuminata (A genome) R2R3-MYB gene family. We used knowledge from other plant species, including the model plant A. thaliana, leading to a functional classification of the banana R2R3-MYB genes based on the MYB phylogeny. Furthermore, RNA-Seq data was used to analyse expression in different M. acuminata organs and developmental stages and to compare expression patterns of closely grouped co-orthologs. The identification and functional characterization of the R2R3-MYB gene family from banana will provide an insight into the regulatory aspects of different biochemical and physiological processes, as those operating during fruit ripening as well as response to various environmental stresses. Our findings offer the first step towards further investigations on the biological and molecular functions of MYB transcription factors with the selection of genes responsible for economically important traits in Musa, which can be utilized in banana improvement programmes.

Material and methods

Search for MYB protein coding genes in the M. acuminata genome

A consensus R2R3-MYB DNA binding domain sequence [12] (S1 Table) was used as protein query in tBLASTn [13] searches on the M. acuminata DH-Pahang genome sequence (version 2) (https://banana-genome-hub.southgreen.fr/sites/banana-genome-hub.southgreen.fr/files/data/fasta/version2/musa_acuminata_v2_pseudochromosome.fna) in an initial search for MYB protein coding genes. To confirm the obtained amino acid sequences, the putative MYB sequences were manually analysed for the presence of an intact MYB domain. All M. acuminata MYB candidates from the initial BLAST were inspected to ensure that the putative gene models encode two or more (multiple) MYB repeats. The identified gene models were analysed to map them individually to unique loci in the genome and redundant sequences were discarded from the data set to obtain unique MaMYB genes. The identified MaMYB genes were matched to the automatically annotated genes from the Banana Genome Hub database [14]. The open reading frames of the identified MaMYB were manually inspected and, if available, verified by mapping of RNA-Seq data to the genomic sequence. Resulting MaMYB annotation is provided in the supporting information: Multi-FASTA files with MaMYB CDS sequences (S1 File) and MaMYB peptide sequences (S2 File), general feature format (GFF) file (S3 File) describing the MaMYB genes for use in genome viewers/browers.

Genomic distribution of MaMYB genes

The MaMYB genes were located on the corresponding chromosomes by the MapGene2chromosome web v2 (MG2C) software tool (http://mg2c.iask.in/mg2c_v2.0/) according to their position information of a physical map, available from the DH-Pahang genome annotation (V2).

Phylogenetic analyses

Protein sequences of 133 A. thaliana MYBs were obtained from TAIR (http://www.arabidopsis.org/). We also considered other multiple MYB-repeat proteins from A. thaliana in the phylogenetic analysis to determine orthologs in the M. acuminata genome: five AtMYB3R, AtMYB4R1 and AtCDC5. Additionally, 43 well-known, functionally characterised landmark plant R2R3-MYB protein sequences were collected from GenBank at the National Center for Biotechnology Information (NCBI) (http://www.ncbi.nlm.nih.gov/).

Phylogenetic trees were constructed from ClustalΩ [15] aligned MYB domain sequences (293 MaMYBs, 132 AtMYBs and 43 plant landmark MYBs) using MEGA7 [16] with default settings. A mojority rule Maximum Likelihood (ML) consensus tree inferred from 1000 bootstrap replications was calculated. M. acuminata MYB proteins were classified according to their relationships with corresponding A. thaliana and landmark MYB proteins.

Expression analysis from RNA-Seq data

RNA-Seq data sets were retrieved from the Sequence Read Archive (https://www.ncbi.nlm.nih.gov/sra) via fastq-dump v2.9.6 (https://github.com/ncbi/sra-tools) (S2 Table). STAR v2.5.1b [17] was applied for the mapping of reads to the Pahang v2 reference genome sequence [18] using previously described parameters [19]. featureCounts [20] was applied for quantification of the mapped reads per gene based on the Pahang v2 annotation. Previously developed Python scripts were deployed for the calculation of normalized expression values (https://github.com/bpucker/bananaMYB) [19].

Results and discussion

The first annotated reference genome sequence of M. acuminata (A genome) became available in 2012 [3]. It was obtained from a double haploid (DH) plant of the Pahang cultivar, derived through haploid pollen and spontaneous chromosome doubling from the wild subspecies Musa acuminata ssp. malaccensis [3]. This wild subspecies was involved in the domestication of the vast majority of cultivated bananas and its genetic signature is commonly found in dessert and cooking bananas (ProMusa, http://www.promusa.org). An improved version (DH-Pahang version 2) of the genome assembly and annotation was presented in 2016 comprising 450.7 Mb (86% of the estimated size) from which 89.5% are assigned to one of the 11 chromosomes, predicted to contain 35,276 protein encoding genes [18]. This M. acuminata DH-Pahang version 2 genome sequence provides the platform of this study.

Identification and genomic distribution of M. acuminata MYB genes

A consensus R2R3-MYB DNA binding domain sequence (deduced from Arabidopsis, grape and sugarbeet R2R3-MYBs, S1 Table) was used as protein query in tBLASTn searches on the DH-Pahang version 2 genome sequence to comprehensively identify MYB protein coding genes in M. acuminata. The resulting putative MYB sequences were proven to map to unique loci in the genome and were confirmed to contain an intact MYB domain. This ensured that the gene models contained two or more (multiple) MYB repeats. We identified a set of 285 R2R3-MYB proteins and nine multiple repeat MYB proteins distantly related to the typical R2R3-MYB proteins: six R1R2R3-MYB (MYB3R) proteins, two MYB4R proteins and one CDC5-like protein from the M. acuminata genome sequence (Table 1).

Table 1. List of annotated MYB genes in the Musa acuminata (DH Pahang) genome sequence.

pseudochr. position BHLH coding peptide MYB   
gene ID synonym from to str. motif exons length type functional assignment
Ma01_g00440 314253 315377 - 4 297 R2R3 stress response, hormone signaling
Ma01_g02850 MusaMYB31 [21] 1866501 1867245 + + 3 200 R2R3 repressors PP, sinapate, lignin
Ma01_g04470 3005032 3006551 + 4 230 R2R3 ASR, flower morphogenesis, stilbene
Ma01_g10440 7520585 7526901 + 3 556 R2R3 anther development, stress response
Ma01_g10750 7722708 7723839 - 3 273 R2R3 photomorphogenesis
Ma01_g11890 8618982 8620082 + 2 234 R2R3 secondary cell wall, lignin
Ma01_g12250 8870030 8871190 - 3 334 R2R3
Ma01_g14370 10499811 10500874 + 4 195 R2R3 defense, stress response
Ma01_g15800 11477487 11478908 + 4 197 R2R3 secondary cell wall, lignin
Ma01_g16960 12409268 12410289 + 3 293 R2R3
Ma01_g17260 12621891 12623358 - 5 360 R2R3
Ma01_g17450 12782371 12783358 + 3 270 R2R3 defense, stress response
Ma01_g18470 13704465 13705557 - 2 330 R2R3 suberin
Ma01_g19610 MYB32 [22] 15271319 15272110 - + 2 241 R2R3 repressors PP, sinapate, lignin
Ma01_g19960 15935305 15936912 - 5 282 R2R3 stress response, hormone signaling
Ma01_g21340 20977298 20978421 - 3 196 R2R3 stamen development
Ma02_g00280 2702046 2703056 + 3 281 R2R3 defense, stress response
Ma02_g00290 4086487 4089065 + 3 320 R2R3 flavonols, phlobaphene
Ma02_g03780 15236983 15238054 + 3 307 R2R3 mucillage, lignin, stomatal closure
Ma02_g04860 16268865 16270029 + 3 325 R2R3
Ma02_g05880 17024294 17029382 - 7 489 3R cell cycle control
Ma02_g06190 17225149 17226252 + 3 301 R2R3 axillary meristem, root growth
Ma02_g06670 17595551 17596584 + 2 317 R2R3 suberin
Ma02_g09720 MYB46 [22] 19581503 19582518 + 2 297 R2R3 secondary wall, lignin
Ma02_g09870 19645550 19646834 - 3 381 R2R3
Ma02_g13370 21804465 21806215 + 3 280 R2R3 photomorphogenesis
Ma02_g15770 23338515 23340804 - 3 444 R2R3 anther development, stress response
Ma02_g16570 23869343 23870389 + 3 297 R2R3 root development
Ma02_g17950 MYB48 [22] 24668376 24669235 - 4 205 R2R3
Ma02_g19650 25862380 25864514 + + 3 258 R2R3
Ma02_g19770 MYB63 [22] 25962707 25963911 - 4 303 R2R3 PP, lignin
Ma02_g20270 26305481 26306910 + 4 365 R2R3
Ma02_g21230 26925219 26926055 + 2 244 R2R3 general flavonoid, trichome
Ma02_g21760 27304216 27317415 - 11 1076 3R cell cycle control
Ma02_g22540 27850957 27852253 + 3 229 R2R3 repressors PP, sinapate, lignin
Ma02_g23870 28706512 28707545 + 4 253 R2R3 defense, stress response
Ma02_g24520 29095026 29096192 - 3 292 R2R3 defense, stress response
Ma03_g01260 944774 947014 + 4 124 R2R3 defense, stress response
Ma03_g06410 4438262 4439441 - 4 287 R2R3 root development
Ma03_g07620 5357989 5358809 - 3 205 R2R3
Ma03_g07840 * 5566137 5567139 - + 3 282 R2R3 proanthocyanidins
Ma03_g07850 * 5573753 5574630 + + 3 238 R2R3 proanthocyanidins
Ma03_g08300 5982225 5985925 - 5 287 R2R3 defense, stress response
Ma03_g08930 6570843 6573019 + 3 360 R2R3 embryogenesis, seed maturation
Ma03_g09310 6861304 6862367 + 3 289 R2R3 defense, stress response
Ma03_g09340 6900833 6902218 + 4 319 R2R3 cell wall, lignin, seed oil, axillary meristem
Ma03_g09840 7294462 7296093 + 3 320 R2R3 anther-, tapetum development
Ma03_g11910 9246422 9247790 + 4 360 R2R3
Ma03_g12480 9621017 9623699 - 4 175 R2R3 defense, stress response
Ma03_g12720 9782781 9783893 - 4 290 R2R3 stress response, hormone signaling
Ma03_g14020 11197863 11198880 - 2 313 R2R3 suberin
Ma03_g18410 23945840 23948315 + 3 168 R2R3 cell wall, lignin, seed oil, axillary meristem
Ma03_g19810 25073787 25074642 + 2 259 R2R3 cell wall, lignin, seed oil, axillary meristem
Ma03_g20390 25556498 25558664 - 3 382 R2R3 embryogenesis, seed maturation
Ma03_g21920 26789598 26790783 + + 5 254 R2R3 flavonoid repressor
Ma03_g21970 26827699 26828817 + 3 303 R2R3
Ma03_g23170 27799667 27800542 - + 4 217 R2R3 repressors PP, sinapate, lignin
Ma03_g25780 29742629 29743879 + 4 344 R2R3 mucillage, lignin, stomatal closure
Ma03_g28720 31825604 31827664 + + 3 274 R2R3 proanthocyanidins
Ma03_g29070 32109761 32110872 - 3 325 R2R3
Ma03_g29510 32402508 32403434 - 2 288 R2R3 stress response, hormone signaling
Ma03_g29770 32613605 32614662 + 3 307 R2R3
Ma04_g00460 405252 406293 + 4 265 R2R3 defense, stress response
Ma04_g01010 896369 897222 - 3 226 R2R3
Ma04_g05460 4086523 4089539 + 4 254 R2R3 photomorphogenesis
Ma04_g06410 4737396 4738484 + 3 306 R2R3
Ma04_g09430 6706438 6707529 + 3 302 R2R3 stress response, hormone signaling
Ma04_g11930 8517172 8518916 + 3 228 R2R3 photomorphogenesis
Ma04_g12940 9788899 9789804 + 2 266 R2R3 stress response, hormone signaling
Ma04_g13260 10044518 10045599 + 3 308 R2R3
Ma04_g16770 16592910 16594109 - 3 348 R2R3 suberin
Ma04_g18740 20887894 20888897 + 4 146 R2R3 ASR, flower morphogenesis, stilbene
Ma04_g19500 22140585 22141813 + 2 347 R2R3 suberin
Ma04_g20120 22833201 22834619 + 4 322 R2R3 anther development, stress response
Ma04_g22200 24575839 24576649 - 3 195 R2R3 repressors PP, sinapate, lignin
Ma04_g22930 25120275 25121367 - 4 254 R2R3 axillary meristem, root growth
Ma04_g23220 25400512 25401385 - 3 243 R2R3 PP, lignin
Ma04_g24670 26639014 26639915 + 3 243 R2R3
Ma04_g26220 27753189 27754723 + 4 283 R2R3
Ma04_g26550 MYB85 [22] 27973322 27974181 - 2 249 R2R3 secondary cell wall, lignin
Ma04_g26660 28040859 28041969 - 4 128 R2R3 defense, stress response
Ma04_g26810 28140628 28141740 - 4 269 R2R3 stress response, hormone signaling
Ma04_g28300 29374644 29376644 - + 3 232 R2R3
Ma04_g28510 29563855 29564811 + 4 219 R2R3 ASR, flower morphogenesis, stilbene
Ma04_g30160 30890260 30891484 - 2 368 R2R3 anther development, stress response
Ma04_g31800 32024735 32025857 + 3 279 R2R3 axillary meristem, root growth
Ma04_g31880 32081718 32084107 - 4 142 R2R3 photomorphogenesis
Ma04_g32240 32306362 32307003 + 2 187 R2R3 repressors PP, sinapate, lignin
Ma04_g33920 MYB72 [22] 33328252 33329340 + 3 300 R2R3 PP, lignin
Ma04_g34300 33575039 33576369 + 3 382 R2R3 anther development, stress response
Ma04_g34660 33734735 33735587 + 3 226 R2R3 axillary meristem, root growth
Ma04_g35350 34164151 34165124 - 3 272 R2R3 PP, lignin
Ma04_g35730 34361423 34362164 + 3 204 R2R3 PP, lignin
Ma04_g35890 34456334 34457640 - 3 279 R2R3 photomorphogenesis
Ma04_g38740 36139124 36140139 + 3 279 R2R3 axillary meristem, root growth
Ma05_g01100 651244 652022 + 3 206 R2R3 repressors PP, sinapate, lignin
Ma05_g01880 1153222 1154262 - 3 281 R2R3 axillary meristem, root growth
Ma05_g03340 2404480 2405576 + 3 303 R2R3 PP, lignin
Ma05_g03690 2718554 2719768 - 4 240 R2R3 ASR, flower morphogenesis, stilbene
Ma05_g05670 4316981 4317987 - 4 246 R2R3 secondary cell wall, lignin
Ma05_g06310 4705834 4707231 - + 3 375 R2R3 general flavonoid, trichome
Ma05_g07140 5206984 5207790 - 1 269 R2R3 stress tolerance
Ma05_g07450 5427233 5429855 - 5 295 R2R3 defense, stress response
Ma05_g08960 6598702 6600983 + 5 192 R2R3
Ma05_g10430 7526380 7527556 + 3 279 R2R3 axillary meristem, root growth
Ma05_g12030 8749432 8751067 + 5 255 R2R3 flower meristem identity
Ma05_g14510 10595192 10596181 - 4 229 R2R3 ASR, flower morphogenesis, stilbene
Ma05_g17720 21202263 21203407 - 3 234 R2R3
Ma05_g18420 23864552 23865432 + 3 241 R2R3 repressors PP, sinapate, lignin
Ma05_g18710 24606320 24608917 - 5 288 R2R3 defense, stress response
Ma05_g19630 28142012 28143079 - 3 295 R2R3 root development
Ma05_g20320 31975647 31976748 + 3 307 R2R3 suberin
Ma05_g20740 32405906 32411144 + 6 375 R2R3
Ma05_g20940 32642222 32646244 + 3 268 R2R3
Ma05_g23480 35547417 35548386 + 4 182 R2R3 defense, stress response
Ma05_g23640 35794926 35797163 - 3 321 R2R3 flavonols, phlobaphene
Ma05_g24200 36482627 36483576 + 3 207 R2R3 axillary meristem, root growth
Ma05_g24840 36981509 36982626 + 3 323 R2R3 anther-, tapetum development
Ma05_g25150 37166050 37167522 + 3 316 R2R3 axillary meristem, root growth
Ma05_g25490 37423048 37423913 - 3 226 R2R3 secondary cell wall, lignin
Ma05_g25630 37499891 37501199 - 6 290 R2R3 trichome branching, petal morphogenesis
Ma05_g25680 37532411 37533682 + 3 302 R2R3 cell wall, lignin, seed oil, axillary meristem
Ma05_g28370 39396363 39402499 - + 5 210 R2R3 repressors PP, sinapate, lignin
Ma05_g30120 40637530 40638485 + 3 252 R2R3 cell wall, lignin, seed oil, axillary meristem
Ma05_g30720 40993457 40994353 + 3 237 R2R3 cell cycle regulation
Ma05_g31160 41195712 41202456 - 4 397 R2R3 stress tolerance
Ma05_g31440 41356727 41357758 + 1 344 R2R3 leaf-, shoot-, germ morphogenesis
Ma06_g00910 744618 747868 + 5 394 R2R3 cell wall, lignin, seed oil, axillary meristem
Ma06_g03570 2604334 2605906 + 3 333 R2R3 axillary meristem, root growth
Ma06_g04210 3055555 3059353 - + 4 361 R2R3 proanthocyanidins
Ma06_g04240 3072400 3074637 - 7 345 R2R3 trichome branching, petal morphogenesis
Ma06_g04270 3095165 3096766 + 4 354 R2R3 cell wall, lignin, seed oil, axillary meristem
Ma06_g04370 3152489 3153586 + 4 250 R2R3 defense, stress response
Ma06_g05680 4227458 4228553 + 3 305 R2R3 root development
Ma06_g05960 4396784 4397869 - + 3 275 R2R3 anthocyanins
Ma06_g06660 4801530 4808116 - 7 517 3R cell cycle control
Ma06_g08100 5748553 5749986 + 3 276 R2R3 embryogenesis, seed maturation
Ma06_g08440 5976699 5977672 - 3 243 R2R3
Ma06_g08910 6239164 6240262 + + 5 199 R2R3 repressors PP, sinapate, lignin
Ma06_g11140 MaMYB3 [23] 7827980 7828834 + + 4 200 R2R3 starch degradation, flavonoid repressor
Ma06_g11270 7905384 7906681 + 3 308 R2R3 defense, stress response
Ma06_g12110 8408418 8411546 - 12 469 R2R3 guard cell division, root gravitropism
Ma06_g12160 8449881 8450871 - 4 256 R2R3 defense, stress response
Ma06_g14470 9914817 9917018 - + 5 286 R2R3 repressors PP, sinapate, lignin
Ma06_g16350 11053657 11054699 + + 3 290 R2R3 general flavonoid, trichome
Ma06_g16920 11470816 11471624 + 3 220 R2R3 cell cycle regulation
Ma06_g17440 11851435 11852673 - 4 275 R2R3 cell wall, lignin, seed oil, axillary meristem
Ma06_g19030 13020150 13021197 + 2 263 R2R3 cell wall, lignin, seed oil, axillary meristem
Ma06_g27210 29243310 29244238 - 4 237 R2R3 repressors PP, sinapate, lignin
Ma06_g29060 30553280 30554423 + 3 195 R2R3 stamen development
Ma06_g31020 32237208 32238426 + 3 351 R2R3 defense, stress response
Ma06_g32530 33447718 33449121 + 3 315 R2R3 axillary meristem, root growth
Ma06_g33100 33858326 33871293 + 12 838 4R SNAP complex
Ma06_g33190 33914360 33915233 + 3 244 R2R3
Ma06_g33430 34073015 34074005 + 3 273 R2R3 axillary meristem, root growth
Ma06_g33920 34371289 34372730 + 3 346 R2R3 cell wall, lignin, seed oil, axillary meristem
Ma06_g35430 35255099 35260321 + 3 225 R2R3 stress tolerance
Ma06_g35620 35399156 35400628 + 4 400 R2R3 embryogenesis, seed maturation
Ma06_g37660 36662931 36664093 + 3 299 R2R3 anther-, trichome development
Ma06_g38880 37507220 37508388 - 3 311 R2R3 anther-, trichome development
Ma07_g00270 249167 250371 + 2 361 R2R3
Ma07_g02470 1968784 1973645 - 3 600 R2R3 anther development, stress response
Ma07_g05660 4114936 4116029 + 3 272 R2R3 defense, stress response
Ma07_g05780 4200621 4201699 + 2 251 R2R3 secondary cell wall, lignin
Ma07_g08110 6059949 6061103 + 5 256 R2R3 axillary meristem, root growth
Ma07_g10330 7698721 7708970 + 7 568 3R cell cycle control
Ma07_g10340 7710266 7711261 - 1 332 R2R3 leaf-, shoot-, germ morphogenesis
Ma07_g11110 8262980 8264078 - 3 187 R2R3 cell wall, lignin, seed oil, axillary meristem
Ma07_g12330 9212653 9213850 + 4 272 R2R3 secondary cell wall, lignin
Ma07_g13590 10213901 10216977 - + 3 307 R2R3 general flavonoid, trichome
Ma07_g17600 20759044 20760488 + 5 343 R2R3
Ma07_g19350 27376443 27377703 - 5 270 R2R3 defense, stress response
Ma07_g19470 27474578 27475521 + 4 241 R2R3 cell cycle regulation
Ma07_g19700 27655399 27656957 - 2 470 R2R3 cell wall, lignin, seed oil, axillary meristem
Ma07_g19720 27682692 27684725 + 6 357 R2R3 trichome branching, petal morphogenesis
Ma07_g19880 * 27793556 27794919 + + 4 264 R2R3 proanthocyanidins
Ma07_g19890 * 27798326 27799369 - + 3 269 R2R3 proanthocyanidins
Ma07_g20020 27925622 27926695 + 4 259 R2R3 secondary cell wall, lignin
Ma07_g20990 28973984 28975550 - 3 341 R2R3 axillary meristem, root growth
Ma07_g22540 30450574 30451644 - 4 269 R2R3 defense, stress response
Ma07_g23060 30809797 30823608 + 3 551 R2R3 anther development, stress response
Ma07_g23180 30932263 30933303 + 3 289 R2R3 defense, stress response
Ma07_g23230 * 30951709 30952887 + 3 274 R2R3 cell wall, lignin, seed oil, axillary meristem
Ma07_g23240 * 30951829 30952875 + 3 230 R2R3 cell wall, lignin, seed oil, axillary meristem
Ma07_g26530 33294011 33295448 - 4 292 R2R3
Ma08_g01300 1210795 1211973 - 5 263 R2R3 defense, stress response
Ma08_g01760 1466390 1467139 + 1 250 R2R3 stress tolerance
Ma08_g02100 1707379 1708990 + 3 421 R2R3 embryogenesis, seed maturation
Ma08_g02450 1904860 1905752 + 3 241 R2R3
Ma08_g03420 2489605 2490759 + 3 319 R2R3 anther-, trichome development
Ma08_g10260 7475942 7478109 - 3 371 R2R3 flavonols, phlobaphene
Ma08_g10600 7758649 7759595 - 3 256 R2R3 defense, stress response
Ma08_g11120 8204512 8205519 + 3 278 R2R3 photomorphogenesis
Ma08_g12510 9469693 9470558 + 3 231 R2R3 PP, lignin
Ma08_g13070 10389299 10390584 - 3 313 R2R3 axillary meristem, root growth
Ma08_g14720 14652484 14653886 + 4 370 R2R3
Ma08_g15820 16034645 16037684 - + 3 260 R2R3
Ma08_g15960 16626283 16635800 - 3 557 R2R3
Ma08_g16760 20724484 20725190 - + 3 194 R2R3 flavonoid repressor
Ma08_g17860 27301472 27302427 - 3 249 R2R3 defense, stress response
Ma08_g18540 32070622 32071809 - 4 276 R2R3 defense, stress response
Ma08_g23390 36792861 36793894 + 3 290 R2R3 proanthocyanidins
Ma08_g25570 38350630 38352260 + 3 307 R2R3 axillary meristem, root growth
Ma08_g25960 MYBS3 [24] 38629624 38630727 + 4 296 R2R3 stress response, hormone signaling
Ma08_g26720 39203311 39211401 - 6 541 R2R3
Ma08_g30360 41652620 41653601 + 3 269 R2R3 defense, stress response
Ma08_g31720 MYB83 [22] 42548860 42549849 + 3 279 R2R3 secondary wall, lignin
Ma08_g32760 43364702 43366041 + 4 200 R2R3 repressors PP, sinapate, lignin
Ma08_g34230 44310614 44311366 - + 3 203 R2R3 repressors PP, sinapate, lignin
Ma08_g34710 44706501 44707468 + 3 245 R2R3
Ma09_g03310 2240622 2241647 - 1 342 R2R3 leaf-, shoot-, germ morphogenesis
Ma09_g03740 2481273 2483877 + 2 322 R2R3 stress tolerance
Ma09_g04930 3159224 3160371 - 3 302 R2R3 cell wall, lignin, seed oil, axillary meristem
Ma09_g06730 4303577 4304525 + 3 204 R2R3 stamen development
Ma09_g08140 5353783 5354623 + 2 256 R2R3 cell wall, lignin, seed oil, axillary meristem
Ma09_g08260 5451934 5452919 - + 3 272 R2R3 repressors PP, sinapate, lignin
Ma09_g09400 6192942 6193897 - 3 263 R2R3
Ma09_g09720 6398143 6410415 - 13 798 4R SNAP complex
Ma09_g10800 7342974 7344121 - 3 327 R2R3 anther-, trichome development
Ma09_g11770 8000924 8002091 - 4 262 R2R3 defense, stress response
Ma09_g13170 8908685 8910010 - 3 310 R2R3 axillary meristem, root growth
Ma09_g14260 9743683 9745180 - 4 187 R2R3 root development
Ma09_g15050 10356705 10357805 - 4 248 R2R3 secondary cell wall, lignin
Ma09_g15130 10456847 10458264 + 4 211 R2R3 cell wall, lignin, seed oil, axillary meristem
Ma09_g15440 10764272 10765275 - 4 257 R2R3 proanthocyanidins
Ma09_g15940 11297314 11298296 - 3 260 R2R3 defense, stress response
Ma09_g16940 12438026 12439426 + 4 279 R2R3 defense, stress response
Ma09_g16980 12476657 12478055 + 4 304 R2R3 cell wall, lignin, seed oil, axillary meristem
Ma09_g20280 28998021 28999222 + 3 273 R2R3 axillary meristem, root growth
Ma09_g22730 34631677 34632781 - 3 315 R2R3
Ma09_g23100 35024484 35029665 + 4 288 R2R3 stress response, hormone signaling
Ma09_g23770 35511400 35518711 + 5 1120 2R CDC5
Ma09_g24640 MYB31 [21] 36291996 36292655 - 3 173 R2R3 repressors PP, sinapate, lignin
Ma09_g25010 36615326 36616400 + 3 301 R2R3 defense, stress response
Ma09_g25590 36999501 37000692 - 3 323 R2R3 anther-, trichome development
Ma09_g27990 38852584 38853631 - + 3 260 R2R3 anthocyanins
Ma09_g28970 39592459 39593487 + 1 343 R2R3 leaf-, shoot-, germ morphogenesis
Ma09_g29010 39617399 39618193 - 3 214 R2R3
Ma09_g29660 MYB52 [22] 40061204 40062015 - 3 210 R2R3 cell wall, lignin, seed oil, axillary meristem
Ma10_g01730 5102446 5104125 - 5 286 R2R3 defense, stress response
Ma10_g01750 5159022 5160613 + 5 288 R2R3 defense, stress response
Ma10_g04420 15095718 15097543 - 5 284 R2R3
Ma10_g04920 15562156 15563431 - 3 321 R2R3 cell wall, lignin, seed oil, axillary meristem
Ma10_g05260 15989100 15989828 + 1 243 R2R3 stress tolerance
Ma10_g05680 17016063 17017842 + 3 499 R2R3 embryogenesis, seed maturation
Ma10_g06140 17599985 17600773 + 3 212 R2R3
Ma10_g09100 23301527 23306607 - 4 297 R2R3 anther-, trichome development
Ma10_g09370 23565764 23566747 - 3 268 R2R3 axillary meristem, root growth
Ma10_g10820 24568790 24569501 + 3 176 R2R3 repressors PP, sinapate, lignin
Ma10_g11100 24705755 24706780 - 3 260 R2R3 cell wall, lignin, seed oil, axillary meristem
Ma10_g13000 25942170 25943424 + 4 304 R2R3 cell wall, lignin, seed oil, axillary meristem
Ma10_g13640 26378393 26379280 - 3 236 R2R3 cell cycle regulation
Ma10_g14150 26690247 26691396 - + 4 298 R2R3 general flavonoid, trichome
Ma10_g14950 27211124 27223165 + 16 869 3R cell cycle control
Ma10_g16050 27917472 27918339 + + 3 217 R2R3 repressors PP, sinapate, lignin
Ma10_g17650 28964249 28965405 + + 3 278 R2R3 anthocyanins
Ma10_g18840 29621056 29624189 + 12 467 R2R3 guard cell division, root gravitropism
Ma10_g19130 29806495 29807615 - 3 297 R2R3 root development
Ma10_g19820 30234061 30235019 - 4 175 R2R3 defense, stress response
Ma10_g19970 30309772 30312309 - + 4 206 R2R3 flavonoid repressor
Ma10_g24510 33076512 33077586 - 4 305 R2R3 root development
Ma10_g25660 33694709 33695874 + 3 314 R2R3 root development
Ma10_g26540 34187707 34195688 + 7 567 3R cell cycle control
Ma10_g26660 34247924 34248785 + 3 226 R2R3
Ma10_g29230 35877509 35878859 - 5 315 R2R3 defense, stress response
Ma10_g29290 35933319 35936352 - 3 562 R2R3 anther development, stress response
Ma10_g29660 36170205 36171708 - 7 289 R2R3 trichome branching, petal morphogenesis
Ma10_g29900 36334814 36335832 + 3 285 R2R3 secondary cell wall, lignin
Ma11_g00330 235287 236166 - 3 235 R2R3
Ma11_g00350 255492 257081 + 6 375 R2R3
Ma11_g02310 1659794 1660627 - 3 213 R2R3 PP, lignin
Ma11_g03860 2954888 2956813 - 4 294 R2R3 flower meristem identity
Ma11_g04680 3650672 3651636 + 3 264 R2R3 defense, stress response
Ma11_g06880 5505145 5505826 - 3 167 R2R3 repressors PP, sinapate, lignin
Ma11_g07330 5826847 5827930 + 3 261 R2R3 axillary meristem, root growth
Ma11_g07530 6014287 6015836 - 3 357 R2R3 suberin
Ma11_g08730 6941094 6942543 - 4 275 R2R3 cell wall, lignin, seed oil, axillary meristem
Ma11_g10680 10271192 10273848 - 6 275 R2R3 defense, stress response
Ma11_g10710 10304514 10305532 + 3 275 R2R3
Ma11_g11300 12744937 12748426 + 6 251 R2R3 defense, stress response
Ma11_g11940 15469544 15470676 - 3 309 R2R3 defense, stress response
Ma11_g14670 20385602 20386542 - 3 262 R2R3 defense, stress response
Ma11_g15740 21412108 21413062 - 4 230 R2R3 PP, lignin
Ma11_g16150 21711896 21713795 - 5 233 R2R3
Ma11_g16430 21944276 21945343 + 3 287 R2R3 PP, lignin
Ma11_g19220 24164576 24166138 - 4 344 R2R3 defense, stress response
Ma11_g21160 25411345 25412681 - 6 303 R2R3
Ma11_g21730 25754364 25755420 - 2 326 R2R3 suberin
Ma11_g21820 25814103 25816846 - + 3 235 R2R3
Ma11_g23010 26544741 26545608 - 3 241 R2R3 cell cycle regulation
Ma11_g23420 26767696 26769125 + 3 183 R2R3 cell wall, lignin, seed oil, axillary meristem
Ma00_g01590 9744573 9745661 + 2 283 R2R3 stress response, hormone signaling
Ma00_g04340 36355056 36356428 + 2 283 R2R3 secondary wall, lignin
Ma00_g04960 43098834 43099818 - 3 267 R2R3 defense, stress response
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The genes are ordered by DH Pahang version 2 [18] pseudochromosomes, from north to south. The annotation-version specific gene code describing the chromosomal assignment and position on pseudochromosomes is given in the first column. "Ma00" indicates genes located in sequences without chromosomal assignment. Functional assignment is based on the Maximum Likelihood tree presented in Fig 2. Multi-FASTA files with MaMYB CDS and peptide sequences together with a general feature format (GFF) file describing the annotated MaMYB genes is provided in the supplementaries. str.: strand

*: paralogs, ASR: abiotic stress response, PP: phenylpropanoid.

The number of R2R3-MYB genes is one of the highest among the species that have been studied to date, ranging from 45 in Ginkgo biloba [10] over 157 in Zea mays [25] and 249 in Brassica napus [26] to 360 in Gossypium hirsutum [11]. This is probably due to three whole-genome duplications (γ 100 Myr ago and α, β 65 Myr ago) that occurred during Musa genome evolution [3, 27]. The number of atypical multiple repeat MYB genes identified in M. acuminata is in the same range as those reported for most other plant species, up to six MYB3R and up to two MYB4R and CDC5-like genes.

The 285 MaR2R3-MYB genes identified constitute approximately 0.81% of the 35,276 predicted protein-coding M. acuminata genes and 9.0% of the 3,155 putative M. acuminata transcription factor genes [18]. These were subjected to further analyses. The identified MaR2R3-MYB genes were named following the nomenclature of the locus tags provided in the DH-Pahang version 2 genome annotation (Table 1). A keyword search in the NCBI database (http://www.ncbi.nlm.nih.gov/) revealed no evidence for M. acuminata MYB genes not present in Table 1.

Six publications dealing with M. acuminata MYB genes were identified: one study describes the elevated expression of nine MaMYB genes in transgenic banana plants overexpressing the NAC domain transcription factor MusaVND1 (vascular related NAC domain) indicating a role of these MaMYBs in the regulation of secondary wall deposition [22]. MYBS3 (Ma08_25960) was found to be differentially expressed between cold-sensitive and cold-tolerant bananas [24]. Another publication described the M. acuminata R2R3-MYB gene Ma05_03690 being upregulated in the early response to the endoparasitic root-knot nematode Meloidogyne incognita in roots [28]. MusaMYB31 (Ma01_02850) was identified as a negative regulator of lignin biosynthesis and the general phenylpropanoid biosynthesis pathway [21]. MaMYB3 (Ma06_11140) was found to repress starch degradation in fruit ripening [23] and MaMYB4 (Ma01_19610) was recently described to control fatty acid desaturation [29].

On the basis of the DH-Pahang version 2 annotation, 291 of the 294 MaMYB genes could be assigned to the eleven chromosomes. The chromosomal distribution of MaMYB genes on the pseudochromosomes is shown in Fig 1 and revealed that M. acuminata MYB genes are distributed across all chromosomes.

Fig 1. Distribution of MaMYB genes on the eleven M. acuminata chromosomes.

Fig 1

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Gene structure analysis revealed that most MaR2R3-MYB genes (155 of 285; 54.4%) follow the previously reported rule of having two introns and three exons, and display the highly conserved splicing arrangement that has also been reported for other plant species [25, 30, 31]. A total of 23 (8.1%) MaR2R3-MYB genes have two exons and seven (2.5%) were single exon genes. 67 (23.5%) MaR2R3-MYB genes have four exons, 21 (7.4%) five exons, eight (2.8%) six exons and two of each with seven and twelve exons, respectively. The complex exon-intron structure of Ma06_12110 and Ma10_18840 is conserved in their A. thaliana orthologs AtMYB88 and AtMYB124/FOUR LIPS (FLP) containing ten and eleven exons, respectively. This supports their close evolutionary relationship, but also suggests the conservation of this intron pattern in evolution since the monocot-dicot split 140–150 Myr ago [32].

Chromosomes are drawn to scale. The positions of centromeres (grey ovals) are roughly estimated from repeat distribution data. The chromosomal positions of the MaMYB genes (given in DH Pahang version 2 annotation ID) are indicated. R2R3-MYB genes are given in black letters, MYB genes with more than two MYB repeats are given in blue letters. The number of MYB genes on each chromosome is given below the respective chromosome.

Phylogenetic analysis of the M. acuminata MYB family

With the aim to explore the putative function of the predicted M. acuminata MYBs, we assigned them to plant MYB proteins with known function. For this, we chose primarily data from A. thaliana, which is the source of most functional MYB characterisations. From comparable studies, MYB function appears conserved across MYB clades, suggesting that closely related MYBs recognise similar/same target genes and possess cooperative, overlapping or redundant functions.

To unravel the relationships, we constructed a phylogenetic tree with 468 MYB domain amino acid sequences of MYB proteins. We used 293 MaMYBs (omitting the CDC5-like MaMYB), the complete A. thaliana MYB family (132 members, including 126 R2R3-MYB, five MYB3R and one MYB4R) and 43 functionally well characterised landmark R2R3-MYBs from other plant species. The phylogenetic tree topology allowed us to classify the analysed MYBs into one MYB3R clade, one MYB4R clade and 42 R2R3-MYB protein clades (Fig 2).

Fig 2. Phylogenetic Maximum Likelihood (ML) tree.

Fig 2

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Most R2R3-MYB clades (31 of 42) include variable numbers of MYB proteins from Arabidopsis and banana, indicating that the appearance of most MYB genes in these two species predates the monocot-dicot split as observed in other studies [33]. Several of these clades also contain landmark MYBs from other plant species. The two clades 4 and 28 only contain Arabidopsis R2R3-MYB members, while seven clades (9, 13, 19, 21, 26, 29 and 32, displayed with white background in Fig 2) only contain banana R2R3-MYB members. Additionally, two clades, 24 with monocot anthocyanin biosynthesis regulators and 27 with the strawberry landmark flavonoid biosynthesis repressor FaMYB1 [34], do not contain any A. thaliana MYB (Fig 2).

ML consensus tree inferred from 1000 bootstraps with 468 MYB domain amino acid sequences of MYB proteins from Musa accuminata (Ma), Arabidopsis thaliana (At) and landmark MYBs from other plant species built with MEGA7. Clades are labeled with different colors and functional annotations are given. The numbers at the branches give bootstrap support values from 1000 replications. SCW, secondary cell wall.

The lineage specificity of some MYB clades could indicate that these clades may have been lost or gained in a single order or species during plant evolution, as indicated by other studies [35–37]. For example, clade 28 lacked M. acuminata orthologs, but includes the A. thaliana R2R3-MYBs AtMYB0/GLABRA1 and AtMYB66/WEREWOLF, which have been identified as being involved in the formation of trichomes and root hairs from epidermal cells [38, 39]. Similar observations have been made in maize (monocot) and sugarbeet (eudicot, caryophyllales) [30, 31], both not containing clade 28 orthologs, while grape (eudicot, rosid) and poplar (eudicot, rosid) do [12, 40]. It has been hypothesized that GLABRA1-like MYB genes have been acquired in rosids after the rosid-asterids split [41]. The absence of MaMYBs in clade 28 is consistent with this hypothesis, since monocots branched off before the separation of asterids and rosids in eudicots. Clade 4 also lacks banana R2R3-MYBs. This clade contains the glucosinolate biosynthesis regulators AtMYB28, AtMYB34 and AtMYB51 [42, 43]. The absence of MaMYBs in this clade is concordant with the fact that glucosinolates are only present in the Brassicaceae family. This clade is thought to have originated from a duplication event before the divergence of the genus Arabidopsis from Brassica [44].

The seven clades containing only MaMYBs were manually inspected by applying BLAST searches at the NCBI protein database in order to identify high similarity to functionally characterized landmark plant R2R3-MYBs. In no case could a landmark MYB be identified. Consequently, these clades could be described as a lineage-specific expansion in M. acuminata, reflecting a species-, genus- or order-specific evolutionary change. These MaMYB proteins may have specialised functions that were acquired or expanded in M. acuminata during genome evolution. Further research will be needed to decipher the biological roles of these MaMYB genes.

R2R3-MYBs may interact with basic helix-turn-helix (bHLH)-type transcription factors, together with WD-repeat (WDR) proteins, forming a trimeric MBW complex. These R2R3-MYBs are defined by a bHLH-binding consensus motif [D/E]Lx2[R/K]x3Lx6Lx3R [45] found in all bHLH-interacting R2R3-MYBs. A search in the MaMYB proteins for the mentioned bHLH-interaction motif identified 30 MaMYBs containing this motif (Table 1) and thus putatively interacting with bHLH proteins. 26 of these 30 MaMYBs were all functionally assigned to clades containing (potentially) known bHLH-interacting R2R3-MYBs: nine in clade 22 (repressors phenylpropanoid, sinapate-, lignin regulators), six in clade 25 (proanthocyanidin regulators), four in clade 23 (general flavonoid, trichome regulators), four in clade 27 (flavonoid repressors) and three in clade 24 (monocot anthocyanin regulators). Four MaMYBs, Ma02_19650, Ma04_28300, Ma08_15820 and Ma11_21820 were all found in the M. acuminata-specific clade 26. These MaMYBs may interact with bHLH-type transcription factors. The close relation to flavonoid biosynthesis regulators suggests that they may act similarly to R2R3-MYB proanthocyanidin regulators. Further research is needed to determine if they regulate a Musa-specific biosynthesis pathway.

Expression profiles for M. acuminata MYB genes in different organs and developmental stages

Since it is not unusual for large transcription factor families in higher organisms to have redundant functions, a particular transcription factor needs to be studied and characterised in the context of the whole family. In this regard, the gene expression pattern can provide a hint to gene function. In this study we used publicly-available RNA-Seq reads from the Sequence Read Archive (S2 Table) to analyse the expression of the 293 MaMYB genes in different organs and developmental stages: embryonic cell suspension, seedling, root and young, adult and old leaf, pulp stages S1-S4 and peel stages S1-S4. Filtered RNA-Seq reads were aligned to the reference genome sequence and the number of mapped reads per annotated transcript were quantified and compared across the analysed samples to calculate normalised RNA-Seq read values which are given in Table 2.

Table 2. The expression profiles of MaR2R3-MYB genes in different organs and developmental stages based on RNA-Seq data.

gene embryogenic seedling root leaf young_leaf adult_leaf old_leaf pulp pulpS1 pulpS2 pulpS3 pulpS4 peel peelS1 peelS2 peelS3 peelS4
Ma00_g01590 50 35 6 151 77 175 348 6 5 5 8 5 8 2 19 10 2
Ma00_g04340 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0
Ma00_g04960 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0
Ma01_g00440 181 49 38 129 53 175 276 32 31 58 34 7 30 9 63 35 11
Ma01_g02850 9 2 14 27 21 31 38 20 24 23 23 10 11 7 9 17 12
Ma01_g04470 1 0 12 4 1 6 2 9 13 9 7 8 6 6 3 7 7
Ma01_g10440 3 16 4 5 7 5 5 4 5 5 3 3 3 3 2 3 4
Ma01_g10750 0 0 2 2 0 0 4 1 1 1 1 2 4 5 4 4 5
Ma01_g11890 0 0 3 3 10 1 0 0 0 0 0 0 0 0 0 0 0
Ma01_g12250 1 0 4 5 17 2 0 3 4 3 2 2 4 4 4 3 3
Ma01_g14370 0 17 11 0 0 0 0 5 7 7 8 0 8 3 7 18 4
Ma01_g15800 1 0 4 1 0 0 0 4 2 5 7 3 6 8 5 2 9
Ma01_g16960 0 0 1 0 1 0 0 0 0 0 0 0 0 0 0 0 0
Ma01_g17260 1 0 11 1 0 0 0 3 3 3 2 3 4 5 3 4 5
Ma01_g17450 0 0 6 2 0 1 6 5 13 2 2 1 1 1 2 2 0
Ma01_g18470 0 0 0 0 0 0 0 0 1 0 0 0 1 2 1 0 0
Ma01_g19610 13 0 6 39 56 40 57 29 78 22 15 2 12 7 36 2 5
Ma01_g19960 35 1 6 97 94 148 145 5 7 6 5 4 19 5 58 10 4
Ma01_g21340 0 0 0 0 0 0 0 2 8 0 0 0 0 0 0 0 0
Ma02_g00280 1 1 21 4 1 1 5 6 8 10 4 2 5 5 4 5 7
Ma02_g00290 0 0 3 9 5 26 1 8 25 0 6 2 3 2 3 2 5
Ma02_g03780 0 0 29 18 26 0 0 13 14 9 18 12 30 39 31 17 33
Ma02_g04860 0 0 1 0 0 0 0 0 0 1 0 0 0 0 0 0 0
Ma02_g05880 4 2 2 3 2 5 3 2 4 2 2 1 2 1 1 1 3
Ma02_g06190 0 0 19 8 0 0 0 16 30 12 13 11 28 42 18 18 32
Ma02_g06670 1 1 4 1 0 0 0 4 3 7 4 4 5 6 2 6 5
Ma02_g09720 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 1
Ma02_g09870 29 1 23 37 99 33 11 19 15 28 27 6 11 11 10 9 13
Ma02_g13370 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0
Ma02_g15770 6 0 1 0 0 1 0 1 2 0 1 0 1 1 1 0 1
Ma02_g16570 0 0 0 0 1 0 1 0 0 0 0 0 0 0 0 0 0
Ma02_g17950 8 0 46 14 4 6 13 33 53 35 16 26 19 19 14 20 22
Ma02_g19650 0 0 3 3 5 1 0 7 10 5 3 11 9 15 8 6 5
Ma02_g19770 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0
Ma02_g20270 3 0 3 16 57 4 0 1 1 2 2 1 4 3 4 3 7
Ma02_g21230 2 2 1 5 14 3 1 3 8 3 0 0 1 0 3 2 0
Ma02_g21760 0 3 7 2 2 0 0 8 11 7 8 7 8 11 5 5 12
Ma02_g22540 1 0 7 4 2 6 7 1 1 1 2 0 1 1 1 1 1
Ma02_g23870 3 0 0 7 4 20 3 2 8 0 0 0 0 0 0 0 0
Ma02_g24520 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0
Ma03_g01260 11 0 6 16 0 0 61 9 31 3 1 2 2 1 3 4 1
Ma03_g06410 0 0 11 1 0 0 0 1 0 3 0 1 1 2 0 2 1
Ma03_g07620 0 0 3 0 0 0 0 0 0 0 0 0 0 0 0 0 0
Ma03_g07840* 0 0 2 1 2 0 0 10 24 9 3 5 2 1 3 1 2
Ma03_g07850* 0 0 1 1 1 0 0 1 2 1 0 1 2 2 2 3 1
Ma03_g08300 0 1 3 0 0 0 1 1 5 0 0 0 1 0 0 6 0
Ma03_g08930 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0
Ma03_g09310 0 0 0 12 42 7 1 0 0 0 0 0 0 0 0 0 0
Ma03_g09340 2 0 7 1 1 0 1 5 8 4 5 2 3 4 2 3 5
Ma03_g09840 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0
Ma03_g11910 5 8 25 35 76 25 6 12 10 13 12 13 24 26 30 18 22
Ma03_g12480 0 0 4 0 0 0 0 1 1 1 0 0 0 0 0 1 0
Ma03_g12720 16 10 4 45 59 49 70 2 2 2 2 3 18 2 32 35 3
Ma03_g14020 0 0 1 0 0 0 0 2 1 4 1 3 2 3 1 3 2
Ma03_g18410 0 0 0 1 2 0 0 4 15 0 0 0 0 0 0 0 0
Ma03_g19810 0 0 2 0 0 0 0 0 0 0 0 0 0 0 0 0 0
Ma03_g20390 5 1 1 2 2 0 4 2 1 3 2 2 0 1 0 0 1
Ma03_g21920 5 0 2 17 13 32 22 4 12 2 2 0 2 0 6 3 0
Ma03_g21970 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0
Ma03_g23170 0 0 3 4 8 2 0 2 2 3 1 2 4 6 2 3 4
Ma03_g25780 3 2 60 18 14 3 0 37 28 81 18 22 40 57 29 26 48
Ma03_g28720 3 3 1 2 0 1 5 1 3 1 0 1 0 0 0 0 0
Ma03_g29070 0 0 1 2 4 2 0 1 1 0 3 0 2 1 2 1 2
Ma03_g29510 14 4 5 44 49 65 55 9 25 5 5 2 6 3 7 11 3
Ma03_g29770 1 0 3 1 1 0 0 2 3 4 0 2 1 2 1 1 2
Ma04_g00460 2 0 1 9 14 20 2 0 0 0 0 0 1 0 2 1 0
Ma04_g01010 0 0 7 0 0 0 1 1 3 1 1 0 1 1 1 0 1
Ma04_g05460 0 0 0 0 0 0 0 0 0 0 1 0 1 0 1 0 1
Ma04_g06410 0 1 1 0 1 0 0 0 1 0 0 0 0 0 0 0 0
Ma04_g09430 4 1 9 12 17 19 2 4 4 2 6 5 8 11 7 3 11
Ma04_g11930 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0
Ma04_g12940 4 0 2 3 6 2 4 2 3 1 2 0 1 0 1 0 0
Ma04_g13260 0 2 2 1 1 1 2 2 3 2 3 1 0 0 0 0 0
Ma04_g16770 0 1 1 1 0 1 0 1 0 1 0 3 1 1 0 1 0
Ma04_g18740 11 1 7 2 1 1 5 2 0 0 6 0 1 0 1 2 0
Ma04_g19500 0 0 4 1 0 0 0 2 4 3 0 1 1 1 1 1 0
Ma04_g20120 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0
Ma04_g22200 0 0 3 0 0 0 0 3 2 8 1 3 1 1 0 1 0
Ma04_g22930 0 0 4 1 0 0 0 2 2 3 0 2 4 6 2 3 4
Ma04_g23220 0 0 5 3 0 0 1 1 3 1 1 1 4 3 6 1 6
Ma04_g24670 0 0 33 6 0 0 0 24 32 24 19 19 17 22 9 16 19
Ma04_g26220 0 1 1 2 1 1 0 1 3 1 1 0 4 2 11 2 1
Ma04_g26550 0 0 5 1 1 0 0 2 3 3 1 1 1 1 1 1 3
Ma04_g26660 0 2 22 1 0 1 3 12 27 11 5 4 1 0 2 1 2
Ma04_g26810 41 78 15 153 258 145 185 39 35 74 32 17 33 13 54 50 14
Ma04_g28300 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0
Ma04_g28510 8 1 3 11 2 22 10 9 13 8 14 2 3 2 5 3 3
Ma04_g30160 1 0 0 0 1 1 0 2 5 1 0 0 0 0 0 0 0
Ma04_g31800 0 0 0 1 0 0 3 0 1 1 0 0 2 0 1 6 0
Ma04_g31880 0 0 2 0 0 0 0 1 0 1 1 0 1 1 2 0 1
Ma04_g32240 0 0 1 0 0 0 0 2 2 4 2 0 0 1 0 0 1
Ma04_g33920 3 7 2 2 4 1 1 0 1 1 0 0 1 1 1 3 1
Ma04_g34300 1 0 0 1 2 1 0 2 4 1 1 2 1 1 0 0 1
Ma04_g34660 0 1 0 0 0 0 0 1 0 0 0 3 0 0 0 0 0
Ma04_g35350 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0
Ma04_g35730 0 0 4 0 0 0 0 0 1 0 0 0 0 0 0 0 0
Ma04_g35890 1 0 0 0 0 0 0 1 2 1 0 0 0 1 0 1 0
Ma04_g38740 0 0 1 0 0 0 0 0 0 0 0 0 1 1 1 0 1
Ma05_g01100 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0
Ma05_g01880 0 0 2 1 0 0 0 2 3 2 0 2 2 1 1 2 2
Ma05_g03340 0 0 2 1 1 0 1 1 0 2 0 0 0 0 0 0 0
Ma05_g03690 10 0 1 14 2 25 28 1 0 1 1 1 2 0 8 1 0
Ma05_g05670 1 0 1 0 0 0 0 1 2 0 0 0 0 0 0 1 0
Ma05_g06310 4 1 1 4 4 4 8 1 3 1 1 0 0 0 1 0 0
Ma05_g07140 4 0 0 0 0 0 0 0 1 1 1 0 0 0 0 2 0
Ma05_g07450 4 0 0 2 4 3 1 1 2 1 1 0 0 0 0 0 0
Ma05_g08960 27 1 9 34 47 53 23 17 38 11 8 10 9 10 3 9 12
Ma05_g10430 0 0 2 1 0 0 0 2 1 1 2 3 3 6 2 1 4
Ma05_g12030 4 1 2 1 1 1 2 3 8 2 1 2 2 2 2 1 1
Ma05_g14510 1 0 4 48 0 1 190 1 0 1 2 0 1 1 1 3 0
Ma05_g17720 0 0 1 0 0 0 0 0 1 0 0 0 4 6 4 2 4
Ma05_g18420 0 0 0 0 0 1 0 1 4 0 0 0 0 0 0 0 0
Ma05_g18710 4 0 3 3 4 4 5 3 5 2 3 0 2 1 4 2 1
Ma05_g19630 0 0 6 0 0 0 0 1 2 3 0 1 1 1 0 2 0
Ma05_g20320 1 0 2 10 5 14 19 0 0 1 0 0 1 1 0 1 0
Ma05_g20740 0 4 1 2 9 0 0 2 5 1 2 0 1 0 2 1 0
Ma05_g20940 1 0 5 11 0 0 44 1 1 0 1 0 0 1 0 0 1
Ma05_g23480 0 0 0 0 0 0 0 2 3 5 0 0 0 0 0 0 1
Ma05_g23640 0 0 1 1 0 3 0 2 7 0 2 0 2 1 3 0 2
Ma05_g24200 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0
Ma05_g24840 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0
Ma05_g25150 0 0 6 2 0 0 0 3 4 4 5 1 3 3 2 2 4
Ma05_g25490 0 1 2 2 1 3 1 1 2 2 1 1 1 2 1 1 2
Ma05_g25630 0 0 0 4 15 0 1 1 6 0 0 0 0 0 0 0 0
Ma05_g25680 1 0 1 2 2 1 3 7 24 2 3 1 0 0 1 1 0
Ma05_g28370 0 0 4 4 9 4 1 1 0 1 0 1 2 2 1 1 2
Ma05_g30120 0 3 16 1 0 1 0 4 7 4 2 2 2 2 1 2 2
Ma05_g30720 0 0 19 1 0 0 0 7 12 10 2 4 3 3 3 1 6
Ma05_g31160 9 3 5 6 7 7 7 5 7 4 4 3 2 2 3 2 4
Ma05_g31440 0 3 2 68 266 2 0 43 12 88 71 2 26 2 89 9 2
Ma06_g00910 0 7 1 5 1 1 15 0 0 0 0 0 1 1 1 3 0
Ma06_g03570 0 0 4 2 0 0 0 3 5 3 3 4 5 7 4 3 7
Ma06_g04210 0 0 5 2 1 0 0 5 5 2 7 4 3 4 3 2 5
Ma06_g04240 1 0 1 1 4 0 0 7 29 0 0 0 0 0 0 0 0
Ma06_g04270 11 1 1 1 0 1 2 3 6 3 2 1 0 0 1 0 0
Ma06_g04370 7 0 10 13 5 29 15 1 3 0 0 0 2 1 6 1 0
Ma06_g05680 0 0 7 4 0 0 0 20 15 34 15 16 8 9 2 11 10
Ma06_g05960 0 0 6 4 4 0 0 13 9 19 13 9 4 4 3 9 2
Ma06_g06660 3 2 2 3 2 3 6 2 3 1 2 2 1 1 1 1 1
Ma06_g08100 3 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0
Ma06_g08440 3 0 1 0 0 0 0 0 0 0 0 0 0 1 0 0 0
Ma06_g08910 4 3 5 28 68 23 16 8 6 11 9 5 10 4 20 14 3
Ma06_g11140 4 0 1 14 6 33 16 2 4 2 0 1 1 0 4 1 0
Ma06_g11270 0 1 10 0 0 0 0 0 1 0 0 0 1 0 1 3 0
Ma06_g12110 0 1 1 1 0 1 1 1 2 1 2 1 1 1 1 1 2
Ma06_g12160 1 1 1 1 1 2 2 1 2 0 1 1 1 0 0 2 1
Ma06_g14470 0 0 3 1 1 1 2 2 1 2 2 2 1 0 1 3 0
Ma06_g16350 0 1 0 3 4 2 5 1 2 1 1 2 1 0 0 1 0
Ma06_g16920 3 1 40 38 30 13 13 60 108 51 34 46 54 72 36 45 61
Ma06_g17440 0 0 0 0 0 0 0 1 1 1 0 1 1 0 1 1 2
Ma06_g19030 0 0 2 0 0 0 1 3 8 1 1 1 1 1 0 1 0
Ma06_g27210 2 2 8 6 2 6 16 3 4 5 0 2 2 2 1 2 2
Ma06_g29060 9 2 0 0 0 0 1 2 7 1 0 0 0 0 0 0 0
Ma06_g31020 7 3 7 6 3 9 8 1 2 2 2 0 2 1 7 1 1
Ma06_g32530 7 0 0 2 1 4 2 0 0 0 0 0 0 0 0 0 1
Ma06_g33100 4 6 3 3 2 3 3 4 5 3 2 6 3 4 2 2 5
Ma06_g33190 0 0 0 1 1 1 1 0 0 0 0 0 0 0 0 0 0
Ma06_g33430 0 1 11 2 1 1 0 8 20 7 2 4 3 5 3 4 3
Ma06_g33920 0 7 10 4 3 0 1 5 5 4 3 6 10 13 8 9 9
Ma06_g35430 6 15 28 31 49 14 31 20 37 22 13 8 33 24 63 25 17
Ma06_g35620 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0
Ma06_g37660 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0
Ma06_g38880 1 0 0 1 2 0 0 0 0 0 0 0 0 0 0 0 0
Ma07_g00270 0 0 1 1 1 0 0 0 1 0 0 0 0 1 0 0 0
Ma07_g02470 5 1 4 5 5 5 7 5 6 7 2 3 3 3 2 2 4
Ma07_g05660 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 2 0
Ma07_g05780 0 1 1 3 6 2 1 2 3 2 2 1 2 1 1 2 2
Ma07_g08110 0 1 5 1 0 0 4 0 1 0 0 0 0 0 0 1 0
Ma07_g10330 2 4 4 1 1 1 2 2 1 2 1 2 2 2 1 1 3
Ma07_g10340 2 11 6 23 82 3 2 11 22 10 8 5 14 8 15 28 5
Ma07_g11110 0 0 0 3 5 5 1 0 0 0 0 0 0 0 0 0 0
Ma07_g12330 0 0 1 7 5 18 3 1 2 1 0 0 0 0 1 0 0
Ma07_g13590 0 0 1 6 4 16 3 0 1 0 0 0 0 0 0 0 0
Ma07_g17600 1 1 8 2 4 0 1 1 1 1 0 1 2 2 2 2 2
Ma07_g19350 2 0 1 22 18 44 23 4 15 0 0 1 9 1 26 11 0
Ma07_g19470 3 0 15 7 5 6 6 54 80 62 33 42 10 14 6 9 10
Ma07_g19700 0 0 0 0 0 0 0 1 4 0 0 0 0 0 0 0 0
Ma07_g19720 0 0 7 7 26 1 0 5 11 6 3 1 4 2 12 1 1
Ma07_g19880* 1 1 5 4 0 0 2 10 14 7 11 8 9 12 3 8 12
Ma07_g19890* 0 0 0 0 0 0 0 0 2 0 0 0 0 0 0 0 0
Ma07_g20020 2 0 1 0 0 1 1 0 0 1 0 0 0 0 1 1 0
Ma07_g20990 0 0 2 0 0 0 0 2 2 1 3 0 2 2 1 1 1
Ma07_g22540 5 1 0 8 13 12 7 1 3 0 0 0 1 0 3 1 0
Ma07_g23060 0 1 2 1 2 0 0 2 4 2 3 1 1 2 1 1 1
Ma07_g23180 3 1 1 3 8 1 0 4 14 1 1 0 3 4 2 1 5
Ma07_g23230* 0 171 9 2 2 1 3 34 6 10 12 106 9 1 2 33 2
Ma07_g23240* 16 23 7 9 9 12 11 9 7 10 12 6 5 6 3 3 7
Ma07_g26530 0 1 1 1 0 2 1 1 4 0 0 0 0 0 0 0 0
Ma08_g01300 6 0 3 6 12 3 7 0 0 0 0 0 1 0 0 2 0
Ma08_g01760 6 0 0 2 2 2 4 0 1 0 0 0 1 0 1 1 1
Ma08_g02100 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0
Ma08_g02450 1 3 5 2 2 1 3 7 7 10 9 1 5 3 13 2 3
Ma08_g03420 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0
Ma08_g10260 1 0 3 7 16 3 1 4 12 1 1 3 13 18 6 7 19
Ma08_g10600 0 0 0 0 0 0 1 0 0 1 1 0 0 0 0 1 0
Ma08_g11120 0 0 0 0 0 0 0 0 1 1 0 0 0 0 0 0 0
Ma08_g12510 0 0 3 1 0 0 0 2 3 2 2 0 2 2 1 1 5
Ma08_g13070 0 0 10 4 0 0 0 17 12 30 18 8 15 19 7 12 21
Ma08_g14720 0 1 5 5 14 1 0 2 4 1 3 2 6 8 6 2 7
Ma08_g15820 0 1 4 2 1 1 2 6 7 6 6 4 4 5 4 5 2
Ma08_g15960 2 6 3 1 2 1 1 4 5 1 2 5 2 2 1 1 2
Ma08_g16760 2 0 2 10 5 13 21 3 5 3 0 3 3 1 4 7 0
Ma08_g17860 0 0 1 2 0 0 8 0 0 0 0 0 2 0 0 7 0
Ma08_g18540 0 0 0 1 0 1 2 0 0 0 0 0 0 0 0 2 0
Ma08_g23390 0 0 0 0 0 0 0 2 6 0 0 0 0 0 1 0 0
Ma08_g25570 7 1 16 11 7 6 7 20 20 26 25 10 19 16 19 16 23
Ma08_g25960 9 11 9 21 22 32 15 13 18 17 11 5 9 9 9 5 13
Ma08_g26720 1 0 7 9 19 4 13 1 1 2 1 1 1 0 1 1 1
Ma08_g30360 0 0 5 0 0 0 0 1 4 0 0 0 1 0 0 3 0
Ma08_g31720 0 0 1 1 1 0 0 0 0 0 1 0 0 0 0 0 0
Ma08_g32760 1 3 11 4 4 1 1 5 5 4 9 3 9 8 16 5 7
Ma08_g34230 0 0 2 1 0 1 0 2 4 1 1 1 2 1 1 2 2
Ma08_g34710 0 0 15 1 0 0 0 16 17 20 8 20 5 5 4 6 7
Ma09_g03310 2 1 11 4 8 3 1 2 5 1 1 2 3 3 5 1 3
Ma09_g03740 8 1 3 4 5 5 4 4 6 3 3 4 2 2 2 2 2
Ma09_g04930 0 2 12 2 0 0 0 10 7 22 7 3 5 5 9 4 3
Ma09_g06730 1 0 0 0 0 0 0 1 1 1 0 0 0 0 0 0 0
Ma09_g08140 0 0 2 0 1 0 0 3 10 0 0 0 0 0 0 0 0
Ma09_g08260 15 1 9 6 8 4 3 9 9 13 10 4 8 9 8 6 8
Ma09_g09400 0 0 7 1 0 0 1 1 2 2 0 1 1 0 1 1 0
Ma09_g09720 1 0 3 1 0 1 0 2 2 2 2 1 1 1 0 1 1
Ma09_g10800 0 0 5 0 0 0 0 1 1 2 0 2 1 1 0 1 0
Ma09_g11770 2 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0
Ma09_g13170 0 0 5 3 0 0 0 6 7 4 10 4 9 12 4 5 13
Ma09_g14260 0 0 1 0 0 0 0 0 0 0 0 0 1 3 0 2 1
Ma09_g15050 1 0 0 0 0 0 0 1 0 0 1 1 1 1 0 0 0
Ma09_g15130 1 0 1 0 0 0 0 1 0 0 3 0 1 1 0 1 1
Ma09_g15440 0 0 1 2 5 1 0 5 9 3 2 4 1 1 2 2 1
Ma09_g15940 2 0 2 3 0 0 13 0 0 0 0 0 0 0 0 0 0
Ma09_g16940 3 0 0 4 5 8 2 0 1 0 0 0 1 2 1 0 1
Ma09_g16980 0 1 7 0 0 0 0 1 4 1 0 1 1 1 1 1 1
Ma09_g20280 0 0 62 8 0 0 0 44 31 73 45 27 26 30 21 29 23
Ma09_g22730 0 0 2 0 0 0 0 1 1 0 0 1 1 1 1 1 1
Ma09_g23100 32 2 3 36 45 65 32 10 9 21 9 1 5 2 12 4 3
Ma09_g24640 0 0 8 4 3 0 8 7 12 6 3 6 4 5 2 6 4
Ma09_g25010 4 0 2 4 3 4 7 1 1 1 2 0 2 1 2 1 2
Ma09_g25590 0 0 2 0 0 0 0 0 0 0 0 0 0 0 0 0 0
Ma09_g27990 0 0 1 1 2 0 0 1 2 2 0 1 2 2 1 1 3
Ma09_g28970 1 0 0 7 26 2 0 1 4 0 0 0 0 0 0 0 0
Ma09_g29010 0 2 32 6 0 1 3 25 26 32 15 25 15 16 7 17 22
Ma09_g29660 0 0 0 0 0 2 0 0 0 0 0 0 0 0 0 0 0
Ma10_g01730 0 0 0 1 4 1 0 0 0 0 0 0 0 0 0 0 0
Ma10_g01750 0 0 0 0 1 1 0 0 0 0 0 0 0 0 0 0 0
Ma10_g04420 0 0 1 4 10 2 2 1 2 1 1 0 1 1 3 0 0
Ma10_g04920 3 0 0 1 3 2 0 0 1 0 0 0 0 0 1 0 0
Ma10_g05260 0 0 0 1 1 1 3 0 0 0 0 0 0 0 0 0 0
Ma10_g05680 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0
Ma10_g06140 0 0 0 0 0 0 1 0 1 0 0 0 0 0 0 0 0
Ma10_g09100 0 0 0 1 4 0 0 0 0 0 0 0 0 0 0 0 0
Ma10_g09370 0 0 4 3 1 0 5 2 2 3 3 1 3 3 2 4 2
Ma10_g10820 0 0 2 0 0 0 0 4 14 1 0 1 1 2 1 1 1
Ma10_g11100 0 1 6 2 0 0 6 0 0 0 0 0 2 0 0 7 0
Ma10_g13000 3 0 5 10 2 21 16 1 1 1 0 2 1 1 1 1 1
Ma10_g13640 2 1 40 31 30 34 12 22 25 19 20 25 35 48 21 28 44
Ma10_g14150 0 1 1 2 2 3 4 1 3 1 0 1 1 0 1 1 0
Ma10_g14950 0 0 3 2 0 0 0 4 8 1 2 3 4 4 2 3 7
Ma10_g16050 5 1 10 28 74 23 13 9 14 16 4 3 7 4 16 3 4
Ma10_g17650 0 30 0 0 0 0 0 4 13 1 0 0 2 2 2 4 1
Ma10_g18840 3 1 5 2 1 2 2 4 6 3 6 2 3 5 2 2 5
Ma10_g19130 0 0 2 0 0 0 0 1 0 0 1 1 1 0 1 0 1
Ma10_g19820 1 0 2 6 0 0 22 0 0 0 0 0 5 0 12 9 0
Ma10_g19970 1 0 0 2 6 3 1 1 3 1 0 0 0 0 0 0 0
Ma10_g24510 9 4 2 7 16 6 4 3 4 2 4 2 1 1 1 1 2
Ma10_g25660 1 0 0 0 0 0 0 1 1 0 0 1 0 0 0 0 0
Ma10_g26540 5 7 6 4 3 4 4 6 8 4 4 6 4 5 2 3 6
Ma10_g26660 2 0 6 3 1 2 2 10 14 11 8 5 4 3 3 5 6
Ma10_g29230 6 0 1 4 6 9 3 0 1 1 0 0 1 0 2 0 0
Ma10_g29290 4 1 1 2 2 2 2 1 1 0 1 1 1 1 0 0 1
Ma10_g29660 2 0 1 15 60 0 0 1 3 0 0 0 0 0 0 0 0
Ma10_g29900 5 1 9 10 7 11 11 8 8 7 9 8 9 11 10 10 7
Ma11_g00330 0 0 4 0 0 0 0 0 1 0 0 1 0 0 0 0 0
Ma11_g00350 0 0 6 5 13 1 0 3 5 2 2 2 5 7 5 3 6
Ma11_g02310 0 0 2 1 0 0 0 2 1 1 4 0 2 2 1 1 4
Ma11_g03860 0 0 2 13 47 1 0 5 14 4 1 0 2 1 5 1 1
Ma11_g04680 20 2 0 32 6 57 64 0 1 1 0 0 4 0 7 10 0
Ma11_g06880 22 4 15 13 7 19 12 31 41 39 30 14 7 6 4 9 11
Ma11_g07330 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0
Ma11_g07530 0 0 5 2 1 1 1 6 3 14 4 2 3 5 1 4 2
Ma11_g08730 1 1 1 1 1 1 1 1 1 1 0 1 1 0 1 0 1
Ma11_g10680 1 2 5 2 1 0 3 12 21 20 6 3 6 7 5 8 6
Ma11_g10710 0 1 2 1 4 0 0 2 5 2 1 2 3 1 6 2 1
Ma11_g11300 2 0 0 0 0 0 1 0 0 0 0 0 0 0 0 1 0
Ma11_g11940 0 0 5 0 0 0 0 0 1 0 0 0 0 0 0 0 0
Ma11_g14670 0 1 2 0 0 0 0 0 0 0 0 0 2 0 2 5 0
Ma11_g15740 4 0 2 5 7 4 6 6 9 4 6 5 2 1 3 2 1
Ma11_g16150 1 2 3 10 37 2 1 4 7 4 6 1 3 3 3 2 4
Ma11_g16430 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0
Ma11_g19220 1 0 1 0 0 0 0 0 1 1 0 0 2 0 0 7 0
Ma11_g21160 1 0 9 3 0 1 12 2 4 0 4 0 0 1 0 0 0
Ma11_g21730 0 0 2 1 0 1 0 2 2 3 0 1 2 4 0 2 2
Ma11_g21820 0 0 0 2 2 4 0 0 0 0 0 0 0 0 1 0 0
Ma11_g23010 0 0 6 1 0 0 0 3 3 5 2 2 2 3 1 2 3
Ma11_g23420 1 0 2 0 0 0 0 1 2 0 0 0 0 0 0 0 0
Open in a new tab

The intensity of green background is correlated to expression level given in FPKM.

* indicate paralogs.

Our expression analyses revealed that M. acuminata MYBs have diverse expression patterns in different organs. Many of the MaMYBs exhibited low transcript abundance levels with expression in only one or a few organs. This is consistent with other transcription factor genes, typically found to be expressed in this manner due to functional specificity and diversity. The highest number of expressed MaMYB genes (221; 75.4%) is observed in roots, followed by pulp (209; 71.3%), leaf (203; 69.3%), peel (196; 66.9%) and embryonic cell suspension (129; 44%). The fewest MaMYB genes are expressed in seedlings (98; 33.4%) in the considered dataset. 40 MaMYB genes (13.7%) were expressed in all samples analysed (albeit with varying expression levels), which suggested that these MaMYBs play regulatory roles at multiple developmental stages in multiple tissues. 13 MaMYB genes (4.4%) lacked expression information in any of the analysed samples, possibly indicating that these genes are expressed in other organs (e.g. pseudostem, flower, bract), specific cells, at specific developmental stages, under special conditions or are pseudogenes. 280 MaMYBs (95.6%) are expressed in at least one analysed sample, although the transcript abundance of some genes was very low. Some MaMYB genes were expressed in all analysed RNA-Seq samples at similar levels (e.g. R2R3-MYBs Ma06_g33440 and Ma11_g06880) while others show variance in transcript abundance with low (no) levels in one or several organs and high levels in others (or vice versa). For example, Ma02_g16570, Ma03_g09310, Ma07_g11110, Ma10_g01730, Ma10_g05260 and Ma10_g09100 show organ-specific expression, as their transcripts were exclusively detected in leaves, which hints to leaf-specific functionality. Ma03_g07840 and Ma07_g19470 were found to be predominantly expressed in pulp, showing expression also in other analysed organs, but not in the seedling. Overall, these results suggests that the corresponding MaMYB regulators are limited to distinct organs, tissues, cells or conditions.

Some paralogous MaMYB genes clustered in the genome (Tables 1 and 2) showed different expression profiles, while other clustered paralogous MaMYB genes did not. For example, the clustered R2R3-MYB genes Ma07_g19880 and Ma07_g19890 (both in the proanthocyanidin-related cluster 25): while Ma07_g19880 is expressed in pulps, peels, roots, and leaves, Ma07_g19890 is nearly not expressed in the analysed organs. The expression pattern of Ma03_g07840 and Ma03_g07850 (also both in the proanthocyanidin-related cluster 25) is, in contrast, very similar, with low expression in root, leaf, pulp and peel, but no expression in embyonic cells and roots. These results could point to functional redundancy of the genes Ma03_g07840 and Ma03_g07850, while Ma07_g19880 and Ma07_g19890 could be (partly) involved in distinct, tissue-specific aspects of specialised mebabolite biosynthesis. The functional categorisation and its expression domains make Ma07_g19880 a good candidate to encode a proanthocyanidin biosynthesis regulator in developing banana fruits (pulp and peel), and thus an excellent target for genetic manipulation of proanthocyanidin content, known to influence senescence of banana fruit [46], to maintain the freshness of harvested banana fruit.

Conclusions

The present genome-wide identification, chromosomal organisation, functional classification and expression analyses of M. acuminata MYB genes provide a first step towards cloning and functional dissection to decode the role of MYB genes in this economically interesting species. Further, knowledge of selected MYB genes can be biotechnologically utilized for improvement of fruit quality, yield, disease resistance, tolerance to biotic and abiotic stresses, and the biosynthesis of pharmaceutical compounds.

Supporting information

S1 Table. MYB domain consensus sequence of R2R3-MYBs, used for tBLASTn searches.

(DOCX)

Click here for additional data file. (12.7KB, docx)
S2 Table. RNA-Seq raw data used for expression analysis.

(DOCX)

Click here for additional data file. (12.3KB, docx)
S1 File. Multi-FASTA file with MaMYB CDS sequences.

(FASTA)

Click here for additional data file. (260.9KB, fasta)
S2 File. Multi-FASTA file with MaMYB peptide sequences.

(FASTA)

Click here for additional data file. (90KB, fasta)
S3 File. MaMYB genes General Feature Format (GFF) file.

For use in genome viewers/browsers on the M. accuminata pseudochromosomes.

(GFF)

Click here for additional data file. (171.4KB, gff)

Acknowledgments

We are grateful to Melanie Kuhlmann for excellent technical assistance. We thank Nathanael Walker-Hale for language editing.

Data Availability

All relevant data are within the manuscript and its Supporting Information files.

Funding Statement

The research stay of AP at Weisshaar lab was supported by a grant from Alexander von Humbold Foundation (AvH) (AvH-IND/1184925). The publication costs were supported by Deutsche Forschungsgemeinschaft (DFG) and the Open Access Publication Fund of Bielefeld University. This is a general fund of Bielefeld University with DFG support. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

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Associated Data

This section collects any data citations, data availability statements, or supplementary materials included in this article.

Supplementary Materials

S1 Table. MYB domain consensus sequence of R2R3-MYBs, used for tBLASTn searches.

(DOCX)

Click here for additional data file. (12.7KB, docx)
S2 Table. RNA-Seq raw data used for expression analysis.

(DOCX)

Click here for additional data file. (12.3KB, docx)
S1 File. Multi-FASTA file with MaMYB CDS sequences.

(FASTA)

Click here for additional data file. (260.9KB, fasta)
S2 File. Multi-FASTA file with MaMYB peptide sequences.

(FASTA)

Click here for additional data file. (90KB, fasta)
S3 File. MaMYB genes General Feature Format (GFF) file.

For use in genome viewers/browsers on the M. accuminata pseudochromosomes.

(GFF)

Click here for additional data file. (171.4KB, gff)

Data Availability Statement

All relevant data are within the manuscript and its Supporting Information files.

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