.TH hifiasm 1 "22 August 2021" "hifiasm-0.16.0 (r369)" "Bioinformatics tools" .SH NAME .PP hifiasm - haplotype-resolved de novo assembler for PacBio Hifi reads. .SH SYNOPSIS * Assemble HiFi reads: .RS 4 .B hifiasm .RB [ -o .IR prefix ] .RB [ -t .IR nThreads ] .RB [ -z .IR endTrimLen ] .R [options] .I input1.fq .RI [ input2.fq .R [...]] .RE * Trio binning assembly with yak dumps: .RS 4 .B yak count .B -o .I paternal.yak .B -b37 .RB [ -t .IR nThreads ] .RB [ -k .IR kmerLen ] .I paternal.fq.gz .br .B yak count .B -o .I maternal.yak .B -b37 .RB [ -t .IR nThreads ] .RB [ -k .IR kmerLen ] .I maternal.fq.gz .br .B hifiasm .RB [ -o .IR prefix ] .RB [ -t .IR nThreads ] .R [options] .B -1 .I paternal.yak .B -2 .I maternal.yak .I child.hifi.fq.gz .RE .SH DESCRIPTION .PP Hifiasm is an ultrafast haplotype-resolved de novo assembler for PacBio Hifi reads. Unlike most existing assemblers, hifiasm starts from uncollapsed genome. Thus, it is able to keep the haplotype information as much as possible. The input of hifiasm is the PacBio Hifi reads in fasta/fastq format, and its outputs consist of multiple types of assembly graphs in GFA format. .SH OPTIONS .SS General options .TP 10 .BI -o \ FILE Prefix of output files [hifiasm.asm]. For detailed description of all assembly graphs, please see the .B OUTPUTS section of this man-page. .TP 10 .BI -t \ INT Number of CPU threads used by hifiasm [1]. .TP .BI -h Show help information. .TP .BI --version Show version number. .SS Error correction options .TP 10 .BI -k \ INT K-mer length [51]. This option must be less than 64. .TP .BI -w \ INT Minimizer window size [51]. .TP .BI -f \ INT Number of bits for bloom filter; 0 to disable [37]. This bloom filter is used to filter out singleton k-mers when counting all k-mers. It takes .RI 2^( INT -3) bytes of memory. A proper setting saves memory. .BR -f37 is recommended for human assembly. For small genomes, use .BR -f0 to disable the initial bloom filter which takes 16GB memory at the beginning. For genomes much larger than human, applying .BR -f38 or even .BR -f39 is preferred to save memory on k-mer counting. .TP .BI -D \ INT Drop k-mers occurring .I >FLOAT*coverage times [5.0]. Hifiasm discards these high-frequency k-mers during error correction to reduce running time. The .I coverage is determined automatically by hifiasm based on k-mer plot, representing homozygous read coverage. Raising this option may improve the resolution of repetitive regions but takes longer time. .TP .BI -N \ INT Consider up to .I max(-D*coverage,-N) overlaps for each oriented read [100]. The .I coverage is determined automatically by hifiasm based on k-mer plot, representing homozygous read coverage. Raising this option may improve the resolution of repetitive regions but takes longer time. .TP .BI -r \ INT Rounds of haplotype-aware error corrections [3]. This option affects all outputs of hifiasm. Odd rounds of correction are preferred in practice. .TP .BI -z \ INT Length of adapters that should be removed [0]. This option remove .I INT bases from both ends of each read. Some old Hifi reads may consist of short adapters (e.g., 20bp adapter at one end). For such data, trimming short adapters would significantly improve the assembly quality. .TP .BI --max-kocc \ INT Employ k-mers occurring < .IR INT times to rescue repetitive overlaps [2000]. This option may improve the resolution of repeats. .TP .BI --hg-size \ INT (k/m/g) Estimated haploid genome size used for inferring read coverage [auto]. This option is used to get accurate homozygous read coverage during error correction. Common suffices are required, for example, 100m or 3g. .TP .BI --min-hist-cnt \ INT When analyzing the k-mer spectrum, ignore counts below .IR INT [5]. For very low coverage of HiFi data, set smaller value for this option. .SS Assembly options .TP .BI -a \ INT Rounds of assembly graph cleaning [4]. This option is used with .B -x and .BR -y . Note that unlike .BR -r , this option does not affect error corrected reads and all-to-all overlaps. .TP .BI -m \ INT Maximal probing distance for bubble popping when generating primary/alternate contig graphs [10000000]. Bubbles longer than .I INT bases will not be popped. For detailed description of these graphs, please see the .B OUTPUTS section of this man-page. .TP .BI -p \ INT Maximal probing distance for bubble popping when generating haplotype-resolved processed unitig graph without small bubbles [0]. Bubbles longer than .I INT bases will not be popped. Small bubbles might be caused by somatic mutations or noise in data. Please note that hifiasm automatically pops small bubbles based on coverage, which can be tweaked by .BR --hom-cov . For detailed description of this graph, please see the .B OUTPUTS section of this man-page. .TP .BI -n \ INT A unitig is considered small if it is composed of less than .I INT reads [3]. Hifiasm may try to remove small unitigs at various steps. .TP .BI -x \ FLOAT1, -y \ FLOAT2 Max and min overlap drop ratio [0.8, 0.2]. This option is used with .BR -a . Given a node .I N in the assembly graph, let max(N) be the length of the longest overlap of .I N. Hifiasm iteratively drops overlaps of .I N if their length / max(N) are below a threshold controlled by .B -x and .BR -y . Hifiasm applies .B -a rounds of short overlap removal with an increasing threshold between .I FLOAT1 and .I FLOAT2. .TP .BI -i Ignore error corrected reads and overlaps saved in .IR prefix .*.bin files so that hifiasm will start again from scratch. Apart from assembly graphs, hifiasm also outputs three binary files that save all overlap information during assembly step. With these files, hifiasm can avoid the time-consuming all-to-all overlap calculation step, and do the assembly directly and quickly. This might be helpful when users want to get an optimized assembly by multiple rounds of experiments with different parameters. .TP .BI -u Disable post-join step for contigs which may improve N50. The post-join step of hifiasm improves contig N50 but may introduce misassemblies. .TP .BI --hom-cov \ INT Homozygous read coverage inferred automatically in default. This option affects different types of outputs, including Hi-C phased assembly and HiFi-only assembly. .TP .BI --pri-range \ INT1[,INT2] Min and max coverage cutoff of primary contigs. Keep contigs with coverage in this range at p_ctg.gfa. Inferred automatically in default. If .I INT2 is not specified, it is set to infinity. Set -1 to disable. .TP .BI --lowQ \ INT Output contig regions with >=INT% inconsistency to the bed file with suffix .B lowQ.bed [70]. Set 0 to disable. .TP .BI --b-cov \ INT Break contigs at potential misassemblies with INT-fold coverage [-1]. Work with .B --m-rate. Set -1 to disable. .TP .BI --m-rate \ FLOAT Break contigs with <=FLOAT*coverage exact overlaps [0.75]. Only work with .B --b-cov and .B --h-cov. .TP .BI --primary Output a primary assembly and an alternate assembly. Hifiasm outputs two balanced assemblies and a primary assembly in default. Enable this option or .B -l0 outputs a primary assembly and an alternate assembly. .SS Trio-partition options .TP 10 .BI -1 \ FILE K-mer dump generated by .B yak count from the paternal/haplotype1 reads [] .TP .BI -2 \ FILE K-mer dump generated by .B yak count from the maternal/haplotype2 reads [] .TP .BI -3 \ FILE List of paternal/haplotype1 read names [] .TP .BI -4 \ FILE List of maternal/haplotype2 read names [] .TP .BI -c \ INT Lower bound of the binned k-mer's frequency [2]. When doing trio binning, a k-mer is said to be differentiating if it occurs >= .B -d times in one sample but occurs < .B -c times in the other sample. .TP .BI -d \ INT Upper bound of the binned k-mer's frequency [5]. When doing trio binning, a k-mer is said to be differentiating if it occurs >= .B -d times in one sample but occurs < .B -c times in the other sample. .TP .BI --t-occ \ INT Forcedly remove unitig including > .I INT unexpected haplotype-specific reads without considering graph topology [60]. .SS Purge-dups options .TP 10 .BI -l \ INT Level of purge-dup. 0 to disable purge-dup, 1 to only purge contained haplotigs, 2 to purge all types of haplotigs, 3 to purge all types of haplotigs in most aggressive way. In default, [3] for non-trio assembly, [0] for trio assembly. For trio assembly, only level 0 and level 1 are allowed. .TP .BI -s \ FLOAT Similarity threshold for duplicate haplotigs that should be purged [0.75 for .BR -l1/-l2 , 0.55 for .BR -l3 ]. .TP .BI -O \ FLOAT Min number of overlapped reads for duplicate haplotigs that should be purged [1]. .TP .BI --purge-max \ INT Coverage upper bound of Purge-dups, which is inferred automatically in default. If the coverage of a contig is higher than this bound, don't apply Purge-dups. Larger value makes assembly more contiguous but may collapse repeats or segmental duplications. .TP .BI --n-hap \ INT Assumption of haplotype number. If it is set to >2, the quality of primary assembly for polyploid genomes might be improved. .SS Hi-C-partition options [experimental, not stable] .TP .BI --h1 \ FILEs File names of input Hi-C R1 [r1_1.fq,r1_2.fq,...]. .TP .BI --h2 \ FILEs File names of input Hi-C R2 [r2_1.fq,r2_2.fq,...]. .TP .BI --n-weight \ INT Rounds of reweighting Hi-C links [3]. Increasing this may improves phasing results but takes longer time. .TP .BI --n-perturb \ INT Rounds of perturbation [10000]. Increasing this may improves phasing results but takes longer time. .TP .BI --f-perturb \ FLOAT Fraction to flip for perturbation [0.1]. Increasing this may improves phasing results but takes longer time. .TP .BI --l-msjoin \ INT Detect misjoined unitigs of >= .I INT in size; 0 to disable [500000]. .TP .BI --seed \ INT RNG seed [11]. .SH OUTPUTS .PP In general, hifiasm generates the following assembly graphs in the GFA format: .RS 2 .TP 2 * .IR prefix .r_utg.gfa: haplotype-resolved raw unitig graph. This graph keeps all haplotype information. .TP * .IR prefix .p_utg.gfa: haplotype-resolved processed unitig graph without small bubbles. Small bubbles might be caused by somatic mutations or noise in data, which are not the real haplotype information. Hifiasm automatically pops such small bubbles based on coverage. The option .BR --hom-cov affects the result. In addition, the option .BR -p forcedly pops bubbles. .TP * .IR prefix .p_ctg.gfa: assembly graph of primary contigs. This graph includes a complete assembly with long stretches of phased blocks. .TP * .IR prefix .a_ctg.gfa: assembly graph of alternate contigs. This graph consists of all contigs that are discarded in primary contig graph. .TP * .IR prefix .*hap*.p_ctg.gfa: phased contig graph. This graph keeps the phased assembly. .RE .PP Hifiasm outputs .B *.r_utg.gfa and .B *.p_utg.gfa in any cases. Specifically, hifiasm outputs the following assembly graphs with trio-binning options: .RS 2 .TP 2 * .IR prefix .dip.hap1.p_ctg.gfa: fully phased paternal/haplotype1 contig graph keeping the phased paternal/haplotype1 assembly. .TP * .IR prefix .dip.hap2.p_ctg.gfa: fully phased maternal/haplotype2 contig graph keeping the phased maternal/haplotype2 assembly. .RE .PP With Hi-C partition options, hifiasm outputs: .RS 2 .TP 2 * .IR prefix .hic.p_ctg.gfa: assembly graph of primary contigs. .TP * .IR prefix .hic.hap1.p_ctg.gfa: fully phased contig graph where each contig is fully phased. .TP * .IR prefix .hic.hap2.p_ctg.gfa: fully phased contig graph where each contig is fully phased. .TP * .IR prefix .hic.a_ctg.gfa (optional with .BR --primary): assembly graph of alternate contigs. .RE .PP Hifiasm keeps Hi-C alignment results and Hi-C index in two bin files: .B *hic.lk.bin and .B *hic.tlb.bin. Rerunning hifiasm with different Hi-C reads needs to delete these bin files or use .BR -i . .RE .PP Hifiasm generates the following assembly graphs only with HiFi reads: .RS 2 .TP 2 * .IR prefix .bp.p_ctg.gfa: assembly graph of primary contigs. .TP * .IR prefix .bp.hap1.p_ctg.gfa: partially phased contig graph of haplotype1. .TP * .IR prefix .bp.hap2.p_ctg.gfa: partially phased contig graph of haplotype2. .RE .PP If the option .BR -l0 or .BR --primary is specified, hifiasm outputs: .RS 2 .TP 2 * .IR prefix .p_ctg.gfa: assembly graph of primary contigs. .TP * .IR prefix .a_ctg.gfa: assembly graph of alternate contigs. .RE .PP For each graph, hifiasm also outputs a simplified version without sequences for the ease of visualization. Hifiasm keeps corrected reads and overlaps in three binary files such as it can regenerate assembly graphs from the binary files without redoing error correction.