MAKER Test

MAKER is a genome annotation pipeline[1]. It allows for a researcher or group of researchers to take a genome, some amount of evidence (for example, an EST file, a protein file (both in FASTA format), and a repeat file, and potentially more), and create structural annotations for a genome. It is capable of training HMM files in order to provide better annotations for a genome with little evidence, although this takes many runs. This page is an attempt to document the work being done to add MAKER into CoGe.

How to Download and Install MAKER

MAKER may be downloaded from the Yandell lab, here: http://www.yandell-lab.org/software/maker.html. The full installation instructions may be found here: http://weatherby.genetics.utah.edu/MAKER/wiki/index.php/MAKER_Tutorial. The instructions here will just serve as a brief overview for getting MAKER running on the command line in UNIX.

1. Register and download MAKER from the Yandell lab MAKER software page.

2. Unpack MAKER in whichever folder it will be run from.

3. Download and install prerequisites if they are not installed. The minimum prerequisites are:

    a. BioPerl and various other Perl modules, listed below (see the MAKER documentation for a complete list[2]).
         i. BioPerl
         ii. DBI
         iii. Error
         iv. Error::Simple
         v. File::NFSLock
         vi. File::Which
         vii. Inline
         viii. Perl::Unsafe::Signals
         ix. Proc::Signal
         x. URI::Escape
         xi. Bit::Vector
         xii. Inline::C
         xiii. PerlIO::gzip
         xiv. forks
         xv. forks::shared
    b. SNAP
    c. Exonerate
    d. RepeatMasker
    e. NCBI BLAST

4. Add MAKER and its prerequisites to $PATH. For example, the paths might look something like:

    a. MAKER: export PATH="/home/user/maker/bin:$PATH"
    b. RepeatMasker: export PATH="/home/user/RepeatMasker:$PATH"
    c. Exonerate: export PATH="/home/user/exonerate-2.2.0-x86_64/bin:$PATH"
    d. SNAP: export PATH="/home/user/snap:$PATH"

Running MAKER from the UNIX command line

1. Set up the MAKER control files by typing "maker -CTL".

    a. To set the genome sequence, enter the path to the genome FASTA file after "genome=". So, this might look like "genome=dpp_contig.fasta".
       Leave a space between this and the commented description (which starts with a "#" symbol).
    b. Set the EST or mRNA data by typing the path to the desired EST or mRNA fasta file after "est=".
    c. To have MAKER generate structural annotations directly from EST or mRNA data, change "est2genome=0" to "est2genome=1".
    d. Change any other desired settings. For more details on these settings and how to set them, see the full MAKER tutorial[3].

2. Open the MAKER boot options file ("maker_bopts.ctl") and ensure that the correct BLAST search type is selected.

    a. For example, to use NCBI-BLAST, set "blast_type=ncbi+".

3. Edit the MAKER options file ("maker_opts.ctl") to the desired settings.

4. Run MAKER by typing "maker" on the command line.

    a. To prevent MAKER from eating up too many system resources, use the "nice" command.
    b. To run MAKER in the background, and to avoid being spammed with constant messages, send the messages to both stdout and stderr with "&> somefile.txt".
    c. Putting both "a" and "b" together, the command might look something like, "nice maker &> log.txt".

5. MAKER may take about a day or more to run, depending on the size of the genome it is attempting to annotate and the evidence it is given. Once finished, it should place all files into a "genomename.maker.output" folder, with all the data split into separate folders by contigs. Each one of these folders will have several sub-folders, one of which should contain a "chromosomename.gff" file which contains the structural annotations, repeats, and will always end with the contig sequence.

Testing MAKER annotation accuracy with minimal data

The point of this exercise is to see how little data can be used with MAKER and still get reasonably accurate annotations.

The Arabidopsis thaliana genome was annotated using MAKER with the Arabidopsis lyrata mRNA data as evidence. Both the genome and the mRNA annotations were downloaded from GenBank. The A. thaliana genome with the annotations from MAKER using the A. lyrata mRNA evidence was uploaded to CoGe (genome id 25440) and compared to the existing A. thaliana annotations (genome id 16911). The analysis can be recreated here: https://genomevolution.org/r/f6kd.

A fair amount of synteny exists between the two genomes, although noise is present. The actual gene regions are currently being compared to assess accuracy directly. The SynMap listed above is used, and then BLAST is set to "BlastN: Small Regions" in GEvo. Also, under "Sequence Options," "Mask Sequence" is set to "Non-CDS" for both sequences.

Table explanation:

    Sample Number: Numerically tracks each trial.
    Coords id16911: The coordinates in the original genome.
    Coords id39871: The coordinates in the MAKER-annotated genome.
    Total Genes id16911: The total number of genes annotated in the original genome. Does not include genes only partially visible in the genome-viewer.
    Total Genes id39871: The total number of genes annotated in the MAKER-annotated genome. Does not include genes only partially visible in the genome-viewer.
    Close Match: The CDS regions of the gene look the same by crude visual analysis. Numbers in parentheses represent the number of matches where a small piece of the gene matched to a different gene.
    Similar Match: A few small differences in the CDS regions can be seen. Numbers in parentheses represent the number of matches where a small piece of the gene matched to a different gene.
    Partial Match: Only a relatively small piece of the genes are matching. Numbers in parentheses represent the number of matches where a small piece of the gene matched to a different gene.
    No Match: The original gene is not represented in the new annotations.
    Pseudogene Misannotated: A pseudogene has been annotated as a functional gene in the MAKER annotations.
    New: A gene in the MAKER annotations is not found in the original genome's annotations.
    Multiple Match (Original:MAKER): These are shown as matching to multiple regions, with the ratio of the number of places on the original genome annotations that match followed by the number of places on the MAKER-annotated genome that match. The quantities of these are also listed. So, an example would look like 1 (only one such multiple match found) 2 (two matches found in the original annotations) : 1 (one match found in the MAKER annotations).
    Split (One gene to two+): Matches where only one gene in the original is present and multiple genes in the MAKER-annotated genome are found. This is different from a multiple match because each region is only matching one place.
    Joined (Two+ genes to one): Matches where multiple genes are present in the original and only one gene is present in the MAKER-annotated genome. This is different from a multiple match because each region is only matching one place.

Sample Number	Coords id16911	Coords id39871	Total Genes id16911	Total Genes id39871	Close Match	Similar Match	Partial Match	No Match	Pseudogene Misannotated	New	Multiple Match (Original:MAKER)	Split (One gene to two+)	Joined (Two+ genes to one)
1	1 544697-648473	1 544697-648473	30	16	10	4	1	12	0	0	0	0	0
2	1 71582-180099	1 71582-180099	36	21	15(5)	2	1	11	0	0	1 2:1	0	1
3	1 1202212-1307510	1 1202212-1307510	26	17	12	3	2	7	0	0	1 2:1	0	0
4	1 1455642-1559002	1 1455642-1559002	29	16	16(9)	0	5	8	0	0	1 2:1	0	1
5	1 1928762-2039295	1 1928762-2039046	33	21	19(2)	1	0	13	1	0	3 2:1	0	0
6	1 2540944-2656892	1 2542248-2656892	31	19	15(6)	1	6	9	0	0	4 3:1, 2 2:1	0	1
7	1 2879268-2981804	1 2879268-2981656	30	18	14(2)	3	8	9	0	0	0	0	1
8	1 3382313-3487979	1 3382313-3487979	27	18	14(2)	1	2	9	0	0	2 2:1	0	0
9	1 3901597-4006840	1 3903894-4006840	32	15	10(1)	3	2	14	0	0	0	0	1
10	1 4197703-4300444	1 4197703-4300444	30	15	15(3)	0	2	13	0	0	1 1:2	0	0
11	1 4958502-5064486	1 4958502-5064486	27	17	15(3)	2	2	7	0	0	1 2:1	0	0
12	1 5380446-5485921	1 5380446-5485921	35	17	13(1)	2(1)	4	15	0	0	2 1:2	0	0
13	1 7384303-7486702	1 7379980-7486702	28	13	6	0	2	8	0	0	4 1:3, 5 1:2, 1 2:1	0	1
14	1 8513858-8619927	1 8513858-8618644	30	15	9(1)	5(2)	2	12	0	0	0	0	1
15	1 9389859-9495818	1 9384718-9495818	31	14	10(2)	1(1)	5	11	0	0	4 1:2	0	2
16	1 9389859-9495818	1 9384718-9495818	26	15	8	5	2	8	0	0	3 1:2	0	0
17	1 12158853-12263571	1 12158853-12263571	22	5	4	0	1	17	0	0	0	0	0
18	1 12859293-12963916	1 12859293-12963916	18	7	3(1)	3(3)	0	10	0	0	1 2:1, 2 1:2	0	0
19	1 14108617-14211652	1 14108617-14211652	5	2	1	1	0	3	0	0	0	0	0
20	1 14108617-14211652	1 14108617-14211652	5	2	1	1	0	3	0	0	0	0	0
21	1 15553892-15657802	1 15553892-15657802	4	2	1	1	0	2	0	0	0	0	0
22	1 17682582-17808194	1 17682582-17805715	29	19	11(1)	6(1)	2	8	0	0	2 1:2	0	1
23	1 20236917-20340245	1 20236944-20340245	28	20	12(1)	5(2)	4	7	0	0	0	0	0
24	1 21984661-22089844	1 21984661-22089844	24	13	10(1)	1(1)	1	9	0	0	1 2:1, 2 1:2	0	1
25	1 26732839-26838712	1 26732839-26838712	33	16	15(2)	1	0	17	0	0	0	0	0
26	5 185117-290911	5 185117-290911	30	19	12(2)	5(1)	4(1)	8	0	0	0	0	0
27	5 2484720-2590086	5 2484720-2590086	29	18	14(6)	2	2(2)	6	0	0	2 3:1	0	0
28	5 5240999-5347779	5 5240999-5347779	30	20	18(4)	2	1	8	0	0	1 2:1	0	0
29	5 7948309-8052594	5 7948309-8052594	29	16	12(4)	2(1)	2	6	0	0	0	0	2
30	5 10746648-10848147	5 10746648-10848147	12	1	1	0	0	11	0	0	0	0	0
31	5 13424499-13532238	5 13434606-13537546	10	4	4(1)	0	0	5	0	0	1 2:1	0	0
32	5 16020402-16123101	5 16020402-16123101	29	9	6(3)	1(1)	4	17	0	1	1 2:1, 2 1:2	0	0
33	5 18741802-18845407	5 18741802-18845407	25	17	12(2)	6(3)	1	3	0	0	1 2:1, 2 1:2	0	0
34	5 21699561-21801099	5 21384155-21488362	26	15	0	0	3(1)	23	0	0	1 1:2	0	0
35	5 24375824-24480269	5 24375824-24480269	19	13	8(5)	5(1)	1	5	0	0	1 1:2	0	0
36	5 26726994-26835104	5 26726994-26835104	34	17	14(5)	3	2	12	0	0	1 2:1	0	0
37	5 1395509-1499568	5 1395509-1499568	28	19	17(2)	2	1(1)	8	0	0	0	0	0
38	5 3963813-4071018	5 3963813-4070909	29	21	20(2)	1	1	7	0	0	0	0	0
39	5 6645731-6751247	5 6645731-6751247	35	21	19(4)	2	2	12	0	0	0	0	0
40	5 9395950-9500584	5 9395950-9500584	28	16	12	3	1	12	0	0	0	0	0
41	5 12035328-12138569	5 12035328-12138569	4	4	3	1	0	0	0	0	0	0	0
42	5 14198202-14303272	5 14198202-14303110	17	8	6	1	0	0	0	0	0	0	1
43	5 17439327-17544830	5 17439468-17544830	28	14	8(1)	5(1)	3(1)	11	0	0	1 2:1	0	0
44	5 20126385-20238307	5 20126385-20238307	21	14	11(7)	2(1)	1	5	0	0	1 1:2, 4 1:4	0	1
45	5 22960801-23065559	5 22960764-23065559	26	14	10(5)	4(2)	2	8	0	0	2 2:1	0	0
46	5 25587492-25693902	5 25587492-25693902	30	23	19(7)	3	3(1)	5	0	0	7 3:1	0	0
47	3 1096943-1203341	3 1096943-1203341	32	19	17(5)	0	2	8	0	0	3 1:2, 1 2:1	0	0
48	3 2215142-2329383	3 2215142-2329383	29	21	15(4)	3(2)	3(1)	7	0	0	3 1:2	0	0
49	3 3432575-3541667	3 3432575-3541908	36	21	15(4)	6(2)	4	11	0	0	0	0	0
50	3 4704624-4811185	3 4704624-4811185	25	15	15(5)	0	3	7	0	0	0	0	0
51	3 5886108-5996205	3 5886108-5996205	30	15	11	4	0	15	0	0	0	0	0
52	3 7009098-7112660	3 7009100-7112660	31	16	12(1)	3(1)	1	8	0	0	1 6:1, 1 2:1	0	0
53	3 8153548-8257243	3 8153548-8257243	18	6	4(3)	2	3	9	0	0	0	0	0
54	3 9258942-9363353	3 9258942-9363353	25	12	9(2)	3	2	10	0	0	0	0	0
55	3 10561071-10666301	3 10543921-10648775	16	5	1	4	0	9	0	0	5 1:3	0	0
56	3 11760867-11863509	3 11760867-11863509	3	1	1	0	0	2	0	0	0	0	0
57	3 12982458-13085348	3 12983120-13085348	7	1	0	1	0	6	0	0	0	0	0
58	3 14035354-14143449	3 14035354-14143449	5	3	2	1	0	2	0	0	0	0	0
59	3 15184235-15295034	3 15184235-15290697	12	6	5(2)	1	0	6	0	0	0	0	0
60	3 16347038-16449725	3 16347038-16449725	20	13	9(6)	2(1)	0	8	0	0	2 1:2, 3 1:3	1	0
61	3 17415629-17517888	3 17415629-17517888	23	13	11(2)	2(1)	0	10	0	0	2 1:2	0	0
62	3 18636527-18750533	3 18636527-18750533	26	12	8(3)	3(1)	1	11	0	0	1 4:1, 1 1:2, 3 1:3	0	0
63	3 19855826-19960027	3 19855826-19960027	36	25	22(2)	3(1)	1	10	0	0	0	0	0
64	3 21065078-21171051	3 21065078-21171051	32	19	15(3)	4(2)	6	6	0	0	2 2:1, 1 3:1	0	0
65	3 22173829-22279195	3 22173829-22279195	23	14	12(4)	1	0	10	1	0	1 3:1	0	0
66	3 23140428-23245727	3 23140428-23245727	31	15	11(2)	3	1	13	0	0	2 1:2	0	1
67	2 902313-1009004	2 902313-1007874	23	9	7	1	2	12	0	0	1 2:1	0	0
68	2 1933901-2035341	2 1925574-2026339	17	7	5(4)	1	0	6	0	0	1 2:1, 5 1:3	0	0
69	2 2535215-2637601	2 2535215-2637601	8	2	2	0	0	4	0	0	0	0	0
70	2 2997623-3107099	2 2997623-3107099	9	1	1	0	0	7	0	0	0	0	0
71	2 4569145-4671448	2 4569145-4671448	7	1	1	0	0	5	0	0	0	0	0
72	2 5064881-5168486	2 5064881-5168486	4	3	3	0	0	1	0	0	0	0	0
73	2 5645124-5756134	2 5645124-5756134	18	6	6(1)	0	0	12	0	0	2 1:2	0	0
74	2 6724150-6827899	2 6724150-6827068	18	11	6(3)	2	0	5	0	0	2 1:2, 3 1:3	1	1
75	2 7734455-7852230	2 7734455-7852230	28	18	14(1)	3	2	9	0	0	0	0	0
76	2 8809211-8914699	2 8809211-8914699	37	18	16(2)	2(1)	2	16	0	0	1 2:1	0	0
77	2 9716127-9819766	2 9716127-9819766	28	13	10(1)	1(1)	1	10	0	0	7 1:7	0	2
78	2 10817070-10922077	2 10817070-10921395	26	13	8(2)	2	1	12	0	0	0	0	0
79	2 11690670-11794867	2 11690670-11794867	33	17	15(3)	1	1	15	0	0	1 2:1, 2 1:2	0	0
80	2 12802632-12907369	2 12802632-12907608	25	16	14(1)	1	1	8	0	0	1 2:1	0	0
81	2 13702665-13806233	2 13702665-13806233	26	18	14(5)	2	1	9	0	0	2 1:2	0	0
82	2 14604611-14707443	2 14604644-14707443	19	13	9(4)	2	3	4	0	0	0	0	0
83	2 15650550-15755165	2 15650550-15755165	25	12	10(2)	2	1	11	0	0	1 2:1	0	0
84	2 16665089-16773406	2 16665089-16773406	26	15	13(3)	0	3	9	0	0	0	0	0
85	2 17684541-17788679	2 17684541-17788679	24	15	14(3)	1	3	5	0	0	1 2:1	0	0
86	2 18707881-18822229	2 18707881-18822229	32	18	15(6)	3(1)	5	8	0	0	4 1:3	0	0
87	4 751713-858018	4 751713-858018	31	16	10(1)	3	0	12	0	0	1 2:1	0	2
88	4 1722114-1824380	4 1722169-1824380	3	2	0	1	0	2	1	0	0	0	0
89	4 17593684-17695729	4 17593684-17695729	23	13	10	3(1)	0	9	0	0	1 2:1	0	0
90	4 16618075-16721974	4 16618075-16721974	25	15	11(3)	4(2)	2	8	0	0	1 2:1	0	0
91	4 2696288-2802663	4 2696288-2802663	21	11	9(3)	2	0	6	0	0	1 2:1, 5 1:2	0	0
92	4 3714499-3816439	4 3714499-3816439	1	1	1	0	0	0	0	0	0	0	0
93	4 4837112-4938939	4 4837112-4938939	4	2	2	0	0	0	0	0	0	0	0
94	4 6473657-6578058	4 6473696-6578058	22	11	8(5)	2	1	10	0	0	9 1:2	0	1
95	4 8312586-8416525	4 8312586-8416525	23	12	7(3)	3	2(1)	11	0	0	0	0	0
96	4 10189947-10297372	4 10189947-10297372	30	15	13(3)	1	4	11	0	0	1 3:1	0	0
97	4 11983703-12093572	4 11983703-12093572	26	16	13(1)	3	2	8	0	0	1 2:1	0	0
98	4 13898993-14007840	4 13898993-14007840	38	23	22(2)	0	2	14	0	0	0	0	0
99	4 14876974-14979681	4 14876974-14979681	32	22	21(1)	1(1)	2	8	0	0	2 1:2, 3 1:3	0	0
100	4 15741039-15845643	4 15741039-15845643	22	19	16(5)	3(2)	0	3	0	0	2 1:2	0	0
Column Totals	N/A	N/A	2362	1312	1006(219)	198(42)	156(9)	852	3	1	N/A	2	22

Summary Percentages

1312/2362 = 56% of genes annotated

1006/1312 = 77% had same CDS

1204/1312 = 92% had similar or better CDS

852/2362 = 36% of genes were not annotated

Note: 56% + 36% = 92%, so where is the missing 8%? The answer is that many genes had multiple syntenic matches, and these are accounted for in the "partial matches" column and the "joined" and "split" columns. If these were to be separated out, then the total missed genes would be (2362 * 0.08) + 852 = 1041.