Error : cannot allocate vector of size 3.2 Gb

[lyy005]

Hi Jim
PrimerTree is a very handy tool! Recently when I ran the package, I got this error. I attached the output here. Do you know if it's because there's too many blast hits?

Primer = search_primer_pair(name='Primer', 'CGAGAAGACCCTATGGAGCTTA', 'AATCGTTGAACAAACGAACC', num_aligns = 50000)
BLASTing 1 primer combinations
Submitting Primer-BLAST query
BLAST alignment processing, refreshing in 20 seconds...
BLAST alignment processing, refreshing in 20 seconds...
BLAST alignment processing, refreshing in 20 seconds...
BLAST alignment processing, refreshing in 20 seconds...
BLAST alignment processing, refreshing in 20 seconds...
BLAST alignment completed in 123 seconds
41599 BLAST alignments parsed in 539 seconds
taxonomy retrieved in 267 seconds
41599 sequences retrieved from NCBI in 7111 seconds, product length min:228 mean:339.02 max:658
41599 sequences aligned in 58704 seconds length:761
pairwise DNA distances calculated in 103 seconds
Error : cannot allocate vector of size 3.2 Gb

[jimhester]

I just updated the source code for the read_dna C functions, which should hopefully fix this error in 2c0e129.

Please install that version from github using

devtools::install_github("jimhester/primerTree")

Then let me know if you still run into the error. I verified that it works on my machine using the primers above with num_aligns = 500. If you are still running into an error I would try lowering that parameter from 50000, that many alignments may be too big to produce the multiple alignment in reasonable time and space.

[lyy005]

I really appreciate your help on this. The reason why I'm using num_aligns = 50000 is because I want to build a reference database for fish species. But there's tons of duplicated mammal sequences. So I have to save more BLAST hits and remove mammal sequences afterwards. Do you know how can I run primer searching on a subgroup of NR database?

Here's what I got from my primer set, more than 95% of the sequences are from mammal.
21132 "Mammalia"
954 "Actinopteri"
26 "Amphibia"
19 "Chondrichthyes"
3 "Cladistia"
83 NA

[jimhester]

Sure, use a CUSTOM_DB parameter with the GI numbers you want to search.
You should be able to get all the GIs you need from a taxonomy search like
http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=7777&lvl=3&lin=f&keep=1&srchmode=1&unlock

On Mon, Jan 19, 2015 at 3:15 PM, lyy005 notifications@github.com wrote:

I really appreciate your help on this. The reason why I'm using num_aligns
= 50000 is because I want to build a reference database for fish species.
But there's tons of duplicated mammal sequences. So I have to save more
BLAST hits and remove mammal sequences afterwards. Do you know how can I
run primer searching on a subgroup of NR database?

Here's what I got from my primer set, more than 95% of the sequences are
from mammal.

21132 "Mammalia"
954 "Actinopteri"
26 "Amphibia"
19 "Chondrichthyes"
3 "Cladistia"
83 NA

—
Reply to this email directly or view it on GitHub
#10 (comment).

[lyy005]

Jim
I've found all the GIs I need for primer searching. I'm not familiar with R. Is CUSTOM_DB a argument of primerTREE? Would you show me how can I feed these GI numbers to primerTREE?

Thank you

[lyy005]

Hi
I tried the custom_db parameter in search_primer_pair and it said this parameter wasn't found. Would you give me any advice on this?

test = search_primer_pair(name='test', 'GCCCCTCAGAATGATATTTGTCCTCA', 'AAAAACCACCGTTGTTATTCAACTA', num_aligns = 50, custom_db = gis)
BLASTing 1 primer combinations
name type defval
1 SEQFILE file
2 PRIMER5_START text
3 PRIMER5_END text
4 PRIMER3_START text
5 PRIMER3_END text
6 PRIMER_LEFT_INPUT text
7 PRIMER_RIGHT_INPUT text
8 PRIMER_PRODUCT_MIN text 70
9 PRIMER_PRODUCT_MAX text 1000
10 PRIMER_NUM_RETURN text 10
11 PRIMER_MIN_TM text 57.0
12 PRIMER_OPT_TM text 60.0
13 PRIMER_MAX_TM text 63.0
14 PRIMER_MAX_DIFF_TM text 3
15 PRIMER_ON_SPLICE_SITE dropdown 0
16 SPLICE_SITE_OVERLAP_5END text 7
17 SPLICE_SITE_OVERLAP_3END text 4
18 SPAN_INTRON checkbox
19 MIN_INTRON_SIZE text 1000
20 MAX_INTRON_SIZE text 1000000
21 SEARCH_SPECIFIC_PRIMER checkbox on
22 SEARCHMODE dropdown 0
23 PRIMER_SPECIFICITY_DATABASE dropdown refseq_mrna
24 CUSTOMSEQFILE file
25 ORGANISM text Homo sapiens
29 AddOrg button
30 EXCLUDE_XM checkbox
31 EXCLUDE_ENV checkbox
32 ENTREZ_QUERY text
33 TOTAL_PRIMER_SPECIFICITY_MISMATCH dropdown 1
34 PRIMER_3END_SPECIFICITY_MISMATCH dropdown 1
35 MISMATCH_REGION_LENGTH dropdown 5
36 TOTAL_MISMATCH_IGNORE dropdown 6
37 PRODUCT_SIZE_DEVIATION dropdown 4000
38 ALLOW_TRANSCRIPT_VARIANTS checkbox
39 NEWWIN checkbox
40 SHOW_SVIEWER checkbox on
41 HITSIZE dropdown 50000
43 EVALUE dropdown 30000
44 WORD_SIZE dropdown 7
45 MAX_CANDIDATE_PRIMER dropdown 500
46 NUM_TARGETS text 20
47 NUM_TARGETS_WITH_PRIMERS text 1000
48 MAX_TARGET_PER_TEMPLATE text 100
49 PRODUCT_MIN_TM text
50 PRODUCT_OPT_TM text
51 PRODUCT_MAX_TM text
52 PRIMER_MIN_SIZE text 15
53 PRIMER_OPT_SIZE text 20
54 PRIMER_MAX_SIZE text 25
55 PRIMER_MIN_GC text 20.0
56 PRIMER_MAX_GC text 80.0
57 GC_CLAMP text 0
58 POLYX text 5
59 PRIMER_MAX_END_STABILITY text 9
60 PRIMER_MAX_END_GC text 5
61 TH_OLOGO_ALIGNMENT checkbox
62 TH_TEMPLATE_ALIGNMENT checkbox
63 PRIMER_MAX_TEMPLATE_MISPRIMING_TH text 40.00
64 PRIMER_PAIR_MAX_TEMPLATE_MISPRIMING_TH text 70.00
65 PRIMER_MAX_SELF_ANY_TH text 45.0
66 PRIMER_MAX_SELF_END_TH text 35.0
67 PRIMER_PAIR_MAX_COMPL_ANY_TH text 45.0
68 PRIMER_PAIR_MAX_COMPL_END_TH text 35.0
69 PRIMER_MAX_HAIRPIN_TH text 24.0
70 PRIMER_MAX_TEMPLATE_MISPRIMING text 12.00
71 PRIMER_PAIR_MAX_TEMPLATE_MISPRIMING text 24.00
72 SELF_ANY text 8.00
73 SELF_END text 3.00
74 PRIMER_PAIR_MAX_COMPL_ANY text 8.00
75 PRIMER_PAIR_MAX_COMPL_END text 3.00
76 EXCLUDED_REGIONS text
77 OVERLAP text
78 OVERLAP_5END text 7
79 OVERLAP_3END text 4
80 MONO_CATIONS text 50.0
81 DIVA_CATIONS text 1.5
82 CON_DNTPS text 0.6
83 SALT_FORMULAR dropdown 1
84 TM_METHOD dropdown 1
85 CON_ANEAL_OLIGO text 50.0
86 NO_SNP checkbox
87 PRIMER_MISPRIMING_LIBRARY dropdown AUTO
88 LOW_COMPLEXITY_FILTER checkbox on
89 PICK_HYB_PROBE checkbox
90 PRIMER_INTERNAL_OLIGO_MIN_SIZE text 18
91 PRIMER_INTERNAL_OLIGO_OPT_SIZE text 20
92 PRIMER_INTERNAL_OLIGO_MAX_SIZE text 27
93 PRIMER_INTERNAL_OLIGO_MIN_TM text 57.0
94 PRIMER_INTERNAL_OLIGO_OPT_TM text 60.0
95 PRIMER_INTERNAL_OLIGO_MAX_TM text 63.0
96 PRIMER_INTERNAL_OLIGO_MIN_GC text 20.0
97 PRIMER_INTERNAL_OLIGO_OPT_GC_PERCENT text 50
98 PRIMER_INTERNAL_OLIGO_MAX_GC text 80.0
99 NEWWIN checkbox
100 SHOW_SVIEWER checkbox on

Error : CUSTOM_DB not valid option

[jimhester]

I think the issue is custom_db is only an option when you select custom
from the dropdown on http://www.ncbi.nlm.nih.gov/tools/primer-blast/. You
may be better off running the query manually then parsing the results with
PrimerTree, although you will probably have to do some programming yourself
to get this working.

Sorry I don't have a better response for you with this, I agree this seems
like a useful feature.