# Double space a file
perl -pe '$\="\n"'
perl -pe 'BEGIN { $\="\n" }'
perl -pe '$_ .= "\n"'
perl -pe 's/$/\n/'
perl -nE 'say'

# Double space a file, except the blank lines
perl -pe '$_ .= "\n" unless /^$/'
perl -pe '$_ .= "\n" if /\S/'

# Triple space a file
perl -pe '$\="\n\n"'
perl -pe '$_.="\n\n"'

# N-space a file
perl -pe '$_.="\n"x7'

# Add a blank line before every line
perl -pe 's//\n/'

# Remove all blank lines
perl -ne 'print unless /^$/'
perl -lne 'print if length'
perl -ne 'print if /\S/'

# Remove all consecutive blank lines, leaving just one
perl -00 -pe ''
perl -00pe0

# Compress/expand all blank lines into N consecutive ones
perl -00 -pe '$_.="\n"x4'

# Fold a file so that every set of 10 lines becomes one tab-separated line
perl -lpe '$\ = $. % 10 ? "\t" : "\n"'

LINE NUMBERING
--------------

# Number all lines in a file
perl -pe '$_ = "$. $_"'

# Number only non-empty lines in a file
perl -pe '$_ = ++$a." $_" if /./'

# Number and print only non-empty lines in a file (drop empty lines)
perl -ne 'print ++$a." $_" if /./'

# Number all lines but print line numbers only non-empty lines
perl -pe '$_ = "$. $_" if /./'

# Number only lines that match a pattern, print others unmodified
perl -pe '$_ = ++$a." $_" if /regex/'

# Number and print only lines that match a pattern
perl -ne 'print ++$a." $_" if /regex/'

# Number all lines, but print line numbers only for lines that match a pattern
perl -pe '$_ = "$. $_" if /regex/'

# Number all lines in a file using a custom format (emulate cat -n)
perl -ne 'printf "%-5d %s", $., $_'

# Print the total number of lines in a file (emulate wc -l)
perl -lne 'END { print $. }'
perl -le 'print $n=()=<>'
perl -le 'print scalar(()=<>)'
perl -le 'print scalar(@foo=<>)'
perl -ne '}{print $.'
perl -nE '}{say $.'

# Print the number of non-empty lines in a file
perl -le 'print scalar(grep{/./}<>)'
perl -le 'print ~~grep{/./}<>'
perl -le 'print~~grep/./,<>'
perl -E 'say~~grep/./,<>'

# Print the number of empty lines in a file
perl -lne '$a++ if /^$/; END {print $a+0}'
perl -le 'print scalar(grep{/^$/}<>)'
perl -le 'print ~~grep{/^$/}<>'
perl -E 'say~~grep{/^$/}<>'

# Print the number of lines in a file that match a pattern (emulate grep -c)
perl -lne '$a++ if /regex/; END {print $a+0}'
perl -nE '$a++ if /regex/; END {say $a+0}'

CALCULATIONS
------------

# Check if a number is a prime
perl -lne '(1x$_) !~ /^1?$|^(11+?)\1+$/ && print "$_ is prime"'

# Print the sum of all the fields on a line
perl -MList::Util=sum -alne 'print sum @F'

# Print the sum of all the fields on all lines
perl -MList::Util=sum -alne 'push @S,@F; END { print sum @S }'
perl -MList::Util=sum -alne '$s += sum @F; END { print $s }'

# Shuffle all fields on a line
perl -MList::Util=shuffle -alne 'print "@{[shuffle @F]}"'
perl -MList::Util=shuffle -alne 'print join " ", shuffle @F'

# Find the minimum element on a line
perl -MList::Util=min -alne 'print min @F'

# Find the minimum element over all the lines
perl -MList::Util=min -alne '@M = (@M, @F); END { print min @M }'
perl -MList::Util=min -alne '$min = min @F; $rmin = $min unless defined $rmin && $min > $rmin; END { print $rmin }'

# Find the maximum element on a line
perl -MList::Util=max -alne 'print max @F'

# Find the maximum element over all the lines
perl -MList::Util=max -alne '@M = (@M, @F); END { print max @M }'

# Replace each field with its absolute value
perl -alne 'print "@{[map { abs } @F]}"'

# Find the total number of fields (words) on each line
perl -alne 'print scalar @F'

# Print the total number of fields (words) on each line followed by the line
perl -alne 'print scalar @F, " $_"'

# Find the total number of fields (words) on all lines
perl -alne '$t += @F; END { print $t}'

# Print the total number of fields that match a pattern
perl -alne 'map { /regex/ && $t++ } @F; END { print $t }'
perl -alne '$t += /regex/ for @F; END { print $t }'
perl -alne '$t += grep /regex/, @F; END { print $t }'

# Print the total number of lines that match a pattern
perl -lne '/regex/ && $t++; END { print $t }'

# Print the number PI to n decimal places
perl -Mbignum=bpi -le 'print bpi(n)'

# Print the number PI to 39 decimal places
perl -Mbignum=PI -le 'print PI'

# Print the number E to n decimal places
perl -Mbignum=bexp -le 'print bexp(1,n+1)'

# Print the number E to 39 decimal places
perl -Mbignum=e -le 'print e'

# Print UNIX time (seconds since Jan 1, 1970, 00:00:00 UTC)
perl -le 'print time'

# Print GMT (Greenwich Mean Time) and local computer time
perl -le 'print scalar gmtime'
perl -le 'print scalar localtime'

# Print local computer time in H:M:S format
perl -le 'print join ":", (localtime)[2,1,0]'

# Print yesterday's date
perl -MPOSIX -le '@now = localtime; $now[3] -= 1; print scalar localtime mktime @now'

# Print date 14 months, 9 days and 7 seconds ago
perl -MPOSIX -le '@now = localtime; $now[0] -= 7; $now[4] -= 14; $now[7] -= 9; print scalar localtime mktime @now'

# Prepend timestamps to stdout (GMT, localtime)
tail -f logfile | perl -ne 'print scalar gmtime," ",$_'
tail -f logfile | perl -ne 'print scalar localtime," ",$_'

# Calculate factorial of 5
perl -MMath::BigInt -le 'print Math::BigInt->new(5)->bfac()'
perl -le '$f = 1; $f *= $_ for 1..5; print $f'

# Calculate greatest common divisor (GCM)
perl -MMath::BigInt=bgcd -le 'print bgcd(@list_of_numbers)'

# Calculate GCM of numbers 20 and 35 using Euclid's algorithm
perl -le '$n = 20; $m = 35; ($m,$n) = ($n,$m%$n) while $n; print $m'

# Calculate least common multiple (LCM) of numbers 35, 20 and 8
perl -MMath::BigInt=blcm -le 'print blcm(35,20,8)'

# Calculate LCM of 20 and 35 using Euclid's formula: n*m/gcd(n,m)
perl -le '$a = $n = 20; $b = $m = 35; ($m,$n) = ($n,$m%$n) while $n; print $a*$b/$m'

# Generate 10 random numbers between 5 and 15 (excluding 15)
perl -le '$n=10; $min=5; $max=15; $, = " "; print map { int(rand($max-$min))+$min } 1..$n'

# Find and print all permutations of a list
perl -MAlgorithm::Permute -le '$l = [1,2,3,4,5]; $p = Algorithm::Permute->new($l); print @r while @r = $p->next'

# Generate the power set
perl -MList::PowerSet=powerset -le '@l = (1,2,3,4,5); for (@{powerset(@l)}) { print "@$_" }'

# Convert an IP address to unsigned integer
perl -le '$i=3; $u += ($_<<8*$i--) for "127.0.0.1" =~ /(\d+)/g; print $u'
perl -le '$ip="127.0.0.1"; $ip =~ s/(\d+)\.?/sprintf("%02x", $1)/ge; print hex($ip)'
perl -le 'print unpack("N", 127.0.0.1)'
perl -MSocket -le 'print unpack("N", inet_aton("127.0.0.1"))'

# Convert an unsigned integer to an IP address
perl -MSocket -le 'print inet_ntoa(pack("N", 2130706433))'
perl -le '$ip = 2130706433; print join ".", map { (($ip>>8*($_))&0xFF) } reverse 0..3'
perl -le '$ip = 2130706433; $, = "."; print map { (($ip>>8*($_))&0xFF) } reverse 0..3'

STRING CREATION AND ARRAY CREATION
----------------------------------

# Generate and print the alphabet
perl -le 'print a..z'
perl -le 'print ("a".."z")'
perl -le '$, = ","; print ("a".."z")'
perl -le 'print join ",", ("a".."z")'

# Generate and print all the strings from "a" to "zz"
perl -le 'print ("a".."zz")'
perl -le 'print "aa".."zz"'

# Create a hex lookup table
@hex = (0..9, "a".."f")

# Convert a decimal number to hex using @hex lookup table
perl -le '$num = 255; @hex = (0..9, "a".."f"); while ($num) { $s = $hex[($num%16)&15].$s; $num = int $num/16 } print $s'
perl -le '$hex = sprintf("%x", 255); print $hex'
perl -le '$num = "ff"; print hex $num'

# Generate a random 8 character password
perl -le 'print map { ("a".."z")[rand 26] } 1..8'
perl -le 'print map { ("a".."z", 0..9)[rand 36] } 1..8'

# Create a string of specific length
perl -le 'print "a"x50'

# Create a repeated list of elements
perl -le '@list = (1,2)x20; print "@list"'

# Create an array from a string
@months = split ' ', "Jan Feb Mar Apr May Jun Jul Aug Sep Oct Nov Dec"
@months = qw/Jan Feb Mar Apr May Jun Jul Aug Sep Oct Nov Dec/

# Create a string from an array
@stuff = ("hello", 0..9, "world"); $string = join '-', @stuff

# Find the numeric values for characters in the string
perl -le 'print join ", ", map { ord } split //, "hello world"'

# Convert a list of numeric ASCII values into a string
perl -le '@ascii = (99, 111, 100, 105, 110, 103); print pack("C*", @ascii)'
perl -le '@ascii = (99, 111, 100, 105, 110, 103); print map { chr } @ascii'

# Generate an array with odd numbers from 1 to 100
perl -le '@odd = grep {$_ % 2 == 1} 1..100; print "@odd"'
perl -le '@odd = grep { $_ & 1 } 1..100; print "@odd"'

# Generate an array with even numbers from 1 to 100
perl -le '@even = grep {$_ % 2 == 0} 1..100; print "@even"'

# Find the length of the string
perl -le 'print length "one-liners are great"'

# Find the number of elements in an array
perl -le '@array = ("a".."z"); print scalar @array'
perl -le '@array = ("a".."z"); print $#array + 1'

TEXT CONVERSION AND SUBSTITUTION
--------------------------------

# ROT13 a string
'y/A-Za-z/N-ZA-Mn-za-m/'

# ROT 13 a file
perl -lpe 'y/A-Za-z/N-ZA-Mn-za-m/' file

# Base64 encode a string
perl -MMIME::Base64 -e 'print encode_base64("string")'
perl -MMIME::Base64 -0777 -ne 'print encode_base64($_)' file

# Base64 decode a string
perl -MMIME::Base64 -le 'print decode_base64("base64string")'
perl -MMIME::Base64 -ne 'print decode_base64($_)' file

# URL-escape a string
perl -MURI::Escape -le 'print uri_escape($string)'

# URL-unescape a string
perl -MURI::Escape -le 'print uri_unescape($string)'

# HTML-encode a string
perl -MHTML::Entities -le 'print encode_entities($string)'

# HTML-decode a string
perl -MHTML::Entities -le 'print decode_entities($string)'

# Convert all text to uppercase
perl -nle 'print uc'
perl -ple '$_=uc'
perl -nle 'print "\U$_"'

# Convert all text to lowercase
perl -nle 'print lc'
perl -ple '$_=lc'
perl -nle 'print "\L$_"'

# Uppercase only the first word of each line
perl -nle 'print ucfirst lc'
perl -nle 'print "\u\L$_"'

# Invert the letter case
perl -ple 'y/A-Za-z/a-zA-Z/'

# Camel case each line
perl -ple 's/(\w+)/\u$1/g'
perl -ple 's/(?<!['])(\w+)/\u\1/g'

# Strip leading whitespace (spaces, tabs) from the beginning of each line
perl -ple 's/^[ \t]+//'
perl -ple 's/^\s+//'

# Strip trailing whitespace (space, tabs) from the end of each line
perl -ple 's/[ \t]+$//'

# Strip whitespace from the beginning and end of each line
perl -ple 's/^[ \t]+|[ \t]+$//g'

# Convert UNIX newlines to DOS/Windows newlines
perl -pe 's|\n|\r\n|'

# Convert DOS/Windows newlines to UNIX newlines
perl -pe 's|\r\n|\n|'

# Convert UNIX newlines to Mac newlines
perl -pe 's|\n|\r|'

# Substitute (find and replace) "foo" with "bar" on each line
perl -pe 's/foo/bar/'

# Substitute (find and replace) all "foo"s with "bar" on each line
perl -pe 's/foo/bar/g'

# Substitute (find and replace) "foo" with "bar" on lines that match "baz"
perl -pe '/baz/ && s/foo/bar/'

# Binary patch a file (find and replace a given array of bytes as hex numbers)
perl -pi -e 's/\x89\xD8\x48\x8B/\x90\x90\x48\x8B/g' file

SELECTIVE PRINTING AND DELETING OF CERTAIN LINES
------------------------------------------------

# Print the first line of a file (emulate head -1)
perl -ne 'print; exit'

# Print the first 10 lines of a file (emulate head -10)
perl -ne 'print if $. <= 10'
perl -ne '$. <= 10 && print'
perl -ne 'print if 1..10'

# Print the last line of a file (emulate tail -1)
perl -ne '$last = $_; END { print $last }'
perl -ne 'print if eof'

# Print the last 10 lines of a file (emulate tail -10)
perl -ne 'push @a, $_; @a = @a[@a-10..$#a]; END { print @a }'

# Print only lines that match a regular expression
perl -ne '/regex/ && print'

# Print only lines that do not match a regular expression
perl -ne '!/regex/ && print'

# Print the line before a line that matches a regular expression
perl -ne '/regex/ && $last && print $last; $last = $_'

# Print the line after a line that matches a regular expression
perl -ne 'if ($p) { print; $p = 0 } $p++ if /regex/'

# Print lines that match regex AAA and regex BBB in any order
perl -ne '/AAA/ && /BBB/ && print'

# Print lines that don't match match regexes AAA and BBB
perl -ne '!/AAA/ && !/BBB/ && print'

# Print lines that match regex AAA followed by regex BBB followed by CCC
perl -ne '/AAA.*BBB.*CCC/ && print'

# Print lines that are 80 chars or longer
perl -ne 'print if length >= 80'

# Print lines that are less than 80 chars in length
perl -ne 'print if length < 80'

# Print only line 13
perl -ne '$. == 13 && print && exit'

# Print all lines except line 27
perl -ne '$. != 27 && print'
perl -ne 'print if $. != 27'

# Print only lines 13, 19 and 67
perl -ne 'print if $. == 13 || $. == 19 || $. == 67'
perl -ne 'print if int($.) ~~ (13, 19, 67)'

# Print all lines between two regexes (including lines that match regex)
perl -ne 'print if /regex1/../regex2/'

# Print all lines from line 17 to line 30
perl -ne 'print if $. >= 17 && $. <= 30'
perl -ne 'print if int($.) ~~ (17..30)'
perl -ne 'print if grep { $_ == $. } 17..30'

# Print the longest line
perl -ne '$l = $_ if length($_) > length($l); END { print $l }'

# Print the shortest line
perl -ne '$s = $_ if $. == 1; $s = $_ if length($_) < length($s); END { print $s }'

# Print all lines that contain a number
perl -ne 'print if /\d/'

# Find all lines that contain only a number
perl -ne 'print if /^\d+$/'

# Print all lines that contain only characters
perl -ne 'print if /^[[:alpha:]]+$/

# Print every second line
perl -ne 'print if $. % 2'

# Print every second line, starting the second line
perl -ne 'print if $. % 2 == 0'

# Print all lines that repeat
perl -ne 'print if ++$a{$_} == 2'

# Print all unique lines
perl -ne 'print unless $a{$_}++'

# Print the first field (word) of every line (emulate cut -f 1 -d ' ')
perl -alne 'print $F[0]'

HANDY REGULAR EXPRESSIONS
-------------------------

# Match something that looks like an IP address
/^\d{1,3}\.\d{1,3}\.\d{1,3}\.\d{1,3}$/
/^(\d{1,3}\.){3}\d{1,3}$/

# Test if a number is in range 0-255
/^([0-9]|[0-9][0-9]|1[0-9][0-9]|2[0-4][0-9]|25[0-5])$/

# Match an IP address
my $ip_part = qr|([0-9]|[0-9][0-9]|1[0-9][0-9]|2[0-4][0-9]|25[0-5])|;
if ($ip =~ /^($ip_part\.){3}$ip_part$/) {
say "valid ip";
}

# Check if the string looks like an email address
/\S+@\S+\.\S+/

# Check if the string is a decimal number
/^\d+$/
/^[+-]?\d+$/
/^[+-]?\d+\.?\d*$/

# Check if the string is a hexadecimal number
/^0x[0-9a-f]+$/i

# Check if the string is an octal number
/^0[0-7]+$/

# Check if the string is binary
/^[01]+$/

# Check if a word appears twice in the string
/(word).*\1/

# Increase all numbers by one in the string
$str =~ s/(\d+)/$1+1/ge

# Extract HTTP User-Agent string from the HTTP headers
/^User-Agent: (.+)$/

# Match printable ASCII characters
/[ -~]/

# Match unprintable ASCII characters
/[^ -~]/

# Match text between two HTML tags
m|<strong>([^<]*)</strong>|
m|<strong>(.*?)</strong>|

# Replace all <b> tags with <strong>
$html =~ s|<(/)?b>|<$1strong>|g

# Extract all matches from a regular expression
my @matches = $text =~ /regex/g;

PERL TRICKS
-----------

# Print the version of a Perl module
perl -MModule -le 'print $Module::VERSION'
perl -MLWP::UserAgent -le 'print $LWP::UserAgent::VERSION'

http://www.i-programmer.info/news/181-algorithms/4537-a-new-dna-sequence-search-compressive-genomics.html

http://en.wikipedia.org/wiki/Compression_of_Genomic_Re-Sequencing_Data

http://www.nature.com/nbt/journal/v30/n7/full/nbt.2241.html

http://bioinformatics.oxfordjournals.org/content/29/13/i283.full

http://groups.csail.mit.edu/cb/cast/

Prerequisites

• A fresh Vultr Cloud Compute instance with Ubuntu 18.04 and root access.

Step 1: Install Apache, MySQL, and PHP

Elgg requires MySQL, PHP, and a web server. Before you can install Elgg, you will need to install the Apache web server, MySQL, and PHP.

Update the repository list.

apt-get update

Install the Apache web server.

apt-get install apache2 -y

Install MySQL.

apt-get install mysql-server -y

Complete the MySQL installation by executing the following command.

/usr/bin/mysql_secure_installation

During the installation, you will be asked to enter a root password. Enter a secure password. This will be the MySQL root password.

Would you like to setup VALIDATE PASSWORD plugin? [Y/N] N
New password: password
Re-enter new password: password
Remove anonymous users? [Y/N] Y
Disallow root login remotely? [Y/N] Y
Remove test database and access to it? [Y/N] Y
Reload privilege tables now? [Y/N] Y

Install PHP 7.2, as well as the PHP modules required by Elgg.

apt-get install php7.2 libapache2-mod-php7.2 php7.2-common php7.2-sqlite3 php7.2-curl php7.2-intl php7.2-mbstring php7.2-xmlrpc php7.2-mysql php7.2-gd php7.2-xml php7.2-cli php7.2-zip -y

Step 2: Create a MySQL database for Elgg

Elgg will require a MySQL database. Log into the MySQL console.

mysql -u root -p

When prompted for a password, enter the MySQL root password you set in step 1. Once you are logged in to the MySQL console, create a new database.

CREATE DATABASE elgg;

Create a new MySQL user and grant it privileges to the newly created database. You can replace username and password with the username and password of your choice.

GRANT ALL PRIVILEGES on elgg.* to 'username'@'localhost' identified by 'password';
FLUSH PRIVILEGES;

Exit the MySQL console.

exit

Step 3: Download and Install Elgg

Download the latest version of Elgg.

cd /var/www/html
rm -r index.html
wget https://elgg.org/download/elgg-2.3.7.zip

Unzip the downloaded archive and move the files to the root of the Apache web server.

apt install unzip
unzip elgg-2.3.7.zip
mv ./elgg-2.3.7/* . && rm elgg-2.3.7.zip && rm -r elgg-2.3.7

Create a data directory for Elgg.

sudo mkdir -p /var/www/html/data

Set the appropriate file permissions.

sudo chown -R www-data:www-data /var/www/html/
sudo chmod -R 755 /var/www/html/

Step 4: Configure Apache for Elgg

Elgg requires the Apache rewrite module. Enable the Apache rewrite module.

sudo a2enmod rewrite

Create an Apache configuration file for the Elgg installation.

sudo nano /etc/apache2/sites-available/elgg.conf

Paste the following snippet to the file, replacing example.com with your own domain name.

<VirtualHost *:80>
     DocumentRoot /var/www/html/
     ServerName example.com
     <Directory /var/www/html/>
          Options FollowSymlinks
          AllowOverride All
          Require all granted
     </Directory>
     ErrorLog ${APACHE_LOG_DIR}/error.log
     CustomLog ${APACHE_LOG_DIR}/access.log combined
</VirtualHost>

Enable the configuration and restart the Apache server.

 sudo a2ensite elgg.conf
 sudo systemctl restart apache2.service

The biggest and perhaps most fearsome of the world's big cats, the tiger, shares 95.6 percent of its DNA with humans' cute and furry companions, domestic cats.

The new research showed that big cats have genetic mutations that enabled them to be carnivores. The team also identified mutations that allow snow leopards to thrive at high altitudes.

Reference:

http://www.nbcnews.com/science/your-cat-ferocious-tigers-share-lot-95-6-percent-their-4B11182690

http://timesofindia.indiatimes.com/home/environment/flora-fauna/Gene-mapping-of-tiger-completed/articleshow/22671681.cms

Paper:

http://www.nature.com/ncomms/2013/130917/ncomms3433/full/ncomms3433.html

http://en.wikipedia.org/wiki/Sequence_assembly 

https://genomebiology.biomedcentral.com/articles/10.1186/s13059-016-0951-y

Address of the bookmark: https://sourceforge.net/projects/operasf/

Again, running spades.py will show you the options:

spades.py

This produces:

SPAdes genome assembler v3.10.1

Usage: /usr/local/SPAdes-3.10.1-Linux/bin/spades.py [options] -o <output_dir>

Basic options:
-o      <output_dir>    directory to store all the resulting files (required)
--sc                    this flag is required for MDA (single-cell) data
--meta                  this flag is required for metagenomic sample data
--rna                   this flag is required for RNA-Seq data
--plasmid               runs plasmidSPAdes pipeline for plasmid detection
--iontorrent            this flag is required for IonTorrent data
--test                  runs SPAdes on toy dataset
-h/--help               prints this usage message
-v/--version            prints version

Input data:
--12    <filename>      file with interlaced forward and reverse paired-end reads
-1      <filename>      file with forward paired-end reads
-2      <filename>      file with reverse paired-end reads
-s      <filename>      file with unpaired reads
--pe<#>-12      <filename>      file with interlaced reads for paired-end library number <#> (<#> = 1,2,..,9)
--pe<#>-1       <filename>      file with forward reads for paired-end library number <#> (<#> = 1,2,..,9)
--pe<#>-2       <filename>      file with reverse reads for paired-end library number <#> (<#> = 1,2,..,9)
--pe<#>-s       <filename>      file with unpaired reads for paired-end library number <#> (<#> = 1,2,..,9)
--pe<#>-<or>    orientation of reads for paired-end library number <#> (<#> = 1,2,..,9; <or> = fr, rf, ff)
--s<#>          <filename>      file with unpaired reads for single reads library number <#> (<#> = 1,2,..,9)
--mp<#>-12      <filename>      file with interlaced reads for mate-pair library number <#> (<#> = 1,2,..,9)
--mp<#>-1       <filename>      file with forward reads for mate-pair library number <#> (<#> = 1,2,..,9)
--mp<#>-2       <filename>      file with reverse reads for mate-pair library number <#> (<#> = 1,2,..,9)
--mp<#>-s       <filename>      file with unpaired reads for mate-pair library number <#> (<#> = 1,2,..,9)
--mp<#>-<or>    orientation of reads for mate-pair library number <#> (<#> = 1,2,..,9; <or> = fr, rf, ff)
--hqmp<#>-12    <filename>      file with interlaced reads for high-quality mate-pair library number <#> (<#> = 1,2,..,9)
--hqmp<#>-1     <filename>      file with forward reads for high-quality mate-pair library number <#> (<#> = 1,2,..,9)
--hqmp<#>-2     <filename>      file with reverse reads for high-quality mate-pair library number <#> (<#> = 1,2,..,9)
--hqmp<#>-s     <filename>      file with unpaired reads for high-quality mate-pair library number <#> (<#> = 1,2,..,9)
--hqmp<#>-<or>  orientation of reads for high-quality mate-pair library number <#> (<#> = 1,2,..,9; <or> = fr, rf, ff)
--nxmate<#>-1   <filename>      file with forward reads for Lucigen NxMate library number <#> (<#> = 1,2,..,9)
--nxmate<#>-2   <filename>      file with reverse reads for Lucigen NxMate library number <#> (<#> = 1,2,..,9)
--sanger        <filename>      file with Sanger reads
--pacbio        <filename>      file with PacBio reads
--nanopore      <filename>      file with Nanopore reads
--tslr  <filename>      file with TSLR-contigs
--trusted-contigs       <filename>      file with trusted contigs
--untrusted-contigs     <filename>      file with untrusted contigs

Pipeline options:
--only-error-correction runs only read error correction (without assembling)
--only-assembler        runs only assembling (without read error correction)
--careful               tries to reduce number of mismatches and short indels
--continue              continue run from the last available check-point
--restart-from  <cp>    restart run with updated options and from the specified check-point ('ec', 'as', 'k<int>', 'mc')
--disable-gzip-output   forces error correction not to compress the corrected reads
--disable-rr            disables repeat resolution stage of assembling

Advanced options:
--dataset       <filename>      file with dataset description in YAML format
-t/--threads    <int>           number of threads
                                [default: 16]
-m/--memory     <int>           RAM limit for SPAdes in Gb (terminates if exceeded)
                                [default: 250]
--tmp-dir       <dirname>       directory for temporary files
                                [default: <output_dir>/tmp]
-k              <int,int,...>   comma-separated list of k-mer sizes (must be odd and
                                less than 128) [default: 'auto']
--cov-cutoff    <float>         coverage cutoff value (a positive float number, or 'auto', or 'off') [default: 'off']
--phred-offset  <33 or 64>      PHRED quality offset in the input reads (33 or 64)
                                [default: auto-detect]

As you can see this is also a “pipeline” of tools that can be switched on or off. SPAdes takes quite a long time, so for the purposes of this practical, something like this may suffice:

spades.py -t 4 \
          -m 32 \
          -k 31,51,71 \
          --only-assembler \
          -1 miseq.1.fastq -2 miseq.2.fastq \
          --nanopore minion.fastq \
          -o hybrid_assembly

In turn, these parameters mean

• use 4 threads
• max memory is 32Gb
• use 3 kmer values to build the de bruijn graph(s) - 31, 51 and 71
• only run the assembler, not the correction algorithm (for speed)
• read 1 and read 2 of the MiSeq data
• the nanopore data
• put the output in folder “hybrid_assembly”

Address of the bookmark: ftp://ftp.genomics.org.cn/pub/cope

Prokka – Standalone command line tool, takes just a few minutes per genome. This is the best way to get good quality annotation in a flash, which is particularly useful if you have loads of genomes or need to annotate a pangenome or metagenome. Note however that the quality of functional information is not as good as RAST, and you will need several extra steps if you want to do functional profiling and pathway analysis of your genome(s)… which is in-built in RAST.

NCBI Prokaryotic Genome Annotation Pipeline is designed to annotate bacterial and archaeal genomes (chromosomes and plasmids).

Genome annotation is a multi-level process that includes prediction of protein-coding genes, as well as other functional genome units such as structural RNAs, tRNAs, small RNAs, pseudogenes, control regions, direct and inverted repeats, insertion sequences, transposons and other mobile elements.

PGAP: NCBI has developed an automatic prokaryotic genome annotation pipeline that combines ab initio gene prediction algorithms with homology based methods. The first version of NCBI Prokaryotic Genome Automatic Annotation Pipeline (PGAAP; see Pubmed Article) developed in 2005 has been replaced with an upgraded version that is capable of processing a larger data volume.  NCBI's annotation pipeline depends on several internal databases and is not currently available for download or use outside of the NCBI environment.

BEACON (automated tool for Bacterial GEnome Annotation ComparisON), a fast tool for an automated and a systematic comparison of different annotations of single genomes. The extended annotation assigns putative functions to many genes with unknown functions. BEACON is available under GNU General Public License version 3.0 and is accessible at: http://www.cbrc.kaust.edu.sa/BEACON/.

BlastKOLA: Assigns K numbers to the user's sequence data by BLAST searches, respectively, against a nonredundant set of KEGG GENES. KOALA (KEGG Orthology And Links Annotation) is KEGG's internal annotation tool for K number assignment of KEGG GENES using SSEARCH computation. Annotate Sequence in KEGG Mapper and Pathogen Checker in KEGG Pathogen are special interfaces to this server and can be executed in an interactive mode. BlastKOALA is suitable for annotating fully sequenced genomes.

PAGIT: Provides a toolkit for improving the quality of genome assemblies created via an assembly software. PAGIT compiled four tools: (i) ABACAS which classifies and orientates contigs and estimates the sizes of gaps between them; (ii) IMAGE uses paired-end reads to extend contigs and close gaps within the scaffolds; (iii) ICORN for identifying and correcting small errors in consensus sequences and; (iv) RATT for help annotation. The software was mainly created to analyze parasite genomes of up to about 300 Mb.

MAKER: A portable and easily configurable genome annotation pipeline. MAKER allows smaller eukaryotic and prokaryotic genome projects to independently annotate their genomes and to create genome databases. It identifies repeats, aligns ESTs and proteins to a genome, produces ab-initio gene predictions and automatically synthesizes these data into gene annotations having evidence-based quality values. MAKER's inputs are minimal and its ouputs can be directly loaded into a Generic Model Organism Database (GMOD). They can also be viewed in the Apollo genome browser; this feature of MAKER provides an easy means to annotate, view and edit individual contigs and BACs without the overhead of a database. MAKER is available for download and can be tested online via the MAKER Web Annotation Service (MWAS).

MyPro is a software pipeline for high-quality prokaryotic genome assembly and annotation. It was validated on 18 oral streptococcal strains to produce submission-ready, annotated draft genomes. MyPro installed as a virtual machine and supported by updated databases will enable biologists to perform quality prokaryotic genome assembly and annotation with ease.

Magic-BLAST incorporates within the NCBI BLAST code framework ideas developed in the NCBI Magic pipeline, in particular hit extensions by local walk and jump (http://www.ncbi.nlm.nih.gov/pubmed/26109056), and recursive clipping of mismatches near the edges of the reads, which avoids accumulating artefactual mismatches near splice sites and is needed to distinguish short indels from substitutions near the edges.

Address of the bookmark: https://ncbi.github.io/magicblast/