RU2803650C1 - Method of preimplantation genetic testing for spondyloepiphyseal dysplasia - Google Patents
Method of preimplantation genetic testing for spondyloepiphyseal dysplasia Download PDFInfo
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Изобретение относится к преимплантационному генетическому тестированию моногенных заболеваний. В настоящее время в мире насчитывается более 350 миллионов людей, страдающих редким заболеванием (по данным RARE Project). Общее количество таких заболеваний по подсчетам European Organization for Rare Diseases (EURORDIS) варьируется от 5 до 7 тысяч. При этом около 80% редких заболеваний имеют генетическую причину. Известная генетическая основа заболевания позволяет с высокой точностью предсказать не только здоровье уже родившегося ребенка, но и оценить риск рождения такого ребенка при анализе генотипов родителей, а также провести генетическую диагностику на самых ранних этапах. Преимплантационное генетическое тестирование (ПГТ) моногенного заболевания становится мощным инструментом для профилактики таких заболеваний.The invention relates to preimplantation genetic testing of monogenic diseases. Currently, there are more than 350 million people in the world suffering from a rare disease (according to the RARE Project). The total number of such diseases, according to estimates by the European Organization for Rare Diseases (EURORDIS), varies from 5 to 7 thousand. Moreover, about 80% of rare diseases have a genetic cause. The known genetic basis of the disease makes it possible to predict with high accuracy not only the health of an already born child, but also to assess the risk of having such a child by analyzing the genotypes of the parents, as well as to carry out genetic diagnosis at the earliest stages. Preimplantation genetic testing (PGT) for monogenic diseases is emerging as a powerful tool for the prevention of such diseases.
Настоящее изобретение относится к способу преимплантационного генетического тестирования спондилоэпифизарной дисплазии.The present invention relates to a method for preimplantation genetic testing for spondyloepiphyseal dysplasia.
Спондилоэпифизарная дисплазия (ахондроплазия) - это аутосомно-доминантная хондродисплазия, характеризующаяся непропорционально коротким ростом (коротким туловищем), аномальными эпифизами и уплощенными телами позвонков. Скелетные особенности проявляются при рождении и развиваются со временем. Спондилоэпифизарная дисплазия (ахондроплазия) встречается с частотой 1 на 100000. Это заболевание приводит к низкому росту с очень коротким туловищем и шеей и укороченными конечностями, косолапость, расщелину неба, плоские черты лица, гипертелоризм, аномалии зрения (нистагм, врожденная катаракта, глаукома, отслойка сетчатки), снижение слуха и характерные рентгенологические находки (уплощенные тела позвонков, плоская вертлужная впадина, замедленное окостенение головок бедренных костей с дегенеративными изменениями). При аутосомно-доминантном типе наследования заболевания вероятность рождения ребенка с этим заболеванием в семье составляет 50%.Spondyloepiphyseal dysplasia (achondroplasia) is an autosomal dominant chondrodysplasia characterized by disproportionately short stature (short trunk), abnormal epiphyses, and flattened vertebral bodies. Skeletal features appear at birth and develop over time. Spondyloepiphyseal dysplasia (achondroplasia) occurs with an incidence of 1 in 100,000. This disease results in short stature with a very short trunk and neck and shortened limbs, clubfoot, cleft palate, flat facial features, hypertelorism, visual abnormalities (nystagmus, congenital cataracts, glaucoma, detachment retina), hearing loss and characteristic x-ray findings (flattened vertebral bodies, flat acetabulum, delayed ossification of the femoral heads with degenerative changes). With an autosomal dominant type of inheritance of the disease, the probability of having a child with this disease in the family is 50%.
К заболеванию спондилоэпифизарная дисплазия (ахондроплазия) могут приводить патогенные генетические варианты в гене COL2A1, располагающемся на хромосоме 12. [Zankl, A. et al. Am. J. Med. Genet., 129A: 144-148] Этот ген кодирует белок альфа-1 цепь коллагена II типа, фибриллярного коллагена, содержащегося в хрящах и стекловидном теле глаза.The disease spondyloepiphyseal dysplasia (achondroplasia) can be caused by pathogenic genetic variants in the COL2A1 gene, located on chromosome 12. [Zankl, A. et al. Am. J. Med. Genet., 129A: 144-148] This gene encodes the alpha-1 chain protein of type II collagen, a fibrillar collagen found in cartilage and the vitreous body of the eye.
ПГТ спондилоэпифизарной дисплазии проводится для семей, имеющих подтвержденную молекулярно-генетическую природу заболевания. Важно отметить, что обоснование патогенности и каузативности генетических вариантов происходит до проведения ПГТ моногенного заболевания и не входит ни цели и задачи ПГТ моногенного заболевания, ни в комплекс мероприятий по проведению ПГТ моногенного заболевания. Оценку патогенности проводят по международному стандарту - по критериям, описанным в 2015 году Американским Колледжем Медицинской генетики и Геномики (American College of Medical Genetics and Genomics-Association for Molecular Pathology (ACMG-AMP)) в ходе поиска молекулярно-генетической причины заболевания. ПГТ рекомендуется семьям с высоким риском рождения ребенка с тяжелым (неизлечимым) наследственным заболеванием с установленным патогенными вариантом, обуславливающим этот риск. ПГТ позволяет выбрать из всех эмбрионов, полученных при ЭКО (экстракорпоральном оплодотворении), эмбрионы без патогенного варианта и, следовательно, без риска развития заболевания.PGT of spondyloepiphyseal dysplasia is carried out for families with a confirmed molecular genetic nature of the disease. It is important to note that the substantiation of the pathogenicity and causativeness of genetic variants occurs before carrying out PGT of a monogenic disease and is not included in either the goals and objectives of PGT of a monogenic disease, or in the complex of measures for carrying out PGT of a monogenic disease. The pathogenicity assessment is carried out according to the international standard - according to the criteria described in 2015 by the American College of Medical Genetics and Genomics-Association for Molecular Pathology (ACMG-AMP) in the search for the molecular genetic cause of the disease. PGT is recommended for families with a high risk of having a child with a severe (incurable) hereditary disease with an established pathogenic variant that causes this risk. PGT allows you to select from all the embryos obtained during IVF (in vitro fertilization), embryos without a pathogenic variant and, therefore, without the risk of developing the disease.
Основной проблемой при генетической диагностике эмбрионов является малое исходное количество биоматериала, так как биоптат содержит от одной до трех клеток. В этом случае для повышения эффективности и точности анализа важно как полностью исключить возможность контаминации, так и нивелировать возможный эффект неравномерной и/или неполной амплификации, а также деградации биоматериала. Это требует разработки тест-системы с особыми характеристиками. При этом тест-система разрабатывается с учетом возможности использовать биоматериал разного типа - тотальную дезоксирибонуклеиновую кислоту (ДНК), выделенную из разных тканей, продукт полногеномной амплификации (Whole Genome Amplification, WGA), а также единичные клетки. Сочетание универсальности по отношению к биоматериалу с поэтапной амплификацией целевых фрагментов позволяет проводить анализ нескольких патогенных вариантов для одного образца, в том числе на единичных клетках, а также выявить ситуацию неполной амплификации, контаминации или деградации образца. Еще одной особенностью ПГТ является отсутствие информации о биологических особенностях эмбриона: в отличие от взрослого человека, у эмбриона могут быть любые хромосомные аномалии, которые усложняют задачу оценки статуса эмбриона по конкретному генетическому варианту. Поэтому тест-система для ПГТ моногенного заболевания (ПГТ-М) должна давать возможность выявить такие случаи и оценить их влияние на достоверность результата диагностики.The main problem in the genetic diagnosis of embryos is the small initial amount of biomaterial, since the biopsy contains from one to three cells. In this case, to increase the efficiency and accuracy of the analysis, it is important to both completely eliminate the possibility of contamination and neutralize the possible effect of uneven and/or incomplete amplification, as well as degradation of the biomaterial. This requires the development of a test system with special characteristics. At the same time, the test system is developed taking into account the possibility of using different types of biomaterial - total deoxyribonucleic acid (DNA) isolated from different tissues, a product of Whole Genome Amplification (WGA), as well as single cells. The combination of versatility in relation to biomaterials with step-by-step amplification of target fragments makes it possible to analyze several pathogenic variants for one sample, including on single cells, as well as to identify situations of incomplete amplification, contamination or degradation of the sample. Another feature of PGT is the lack of information about the biological characteristics of the embryo: unlike an adult, an embryo can have any chromosomal abnormalities that complicate the task of assessing the status of the embryo for a specific genetic variant. Therefore, a test system for PGT of a monogenic disease (PGT-M) should make it possible to identify such cases and evaluate their impact on the reliability of the diagnostic result.
Описания близкого технического решения не обнаружено в общедоступных источниках.No description of a similar technical solution was found in publicly available sources.
Представленный нами метод ПГТ спондилоэпифизарной дисплазии решает задачу разработки более точного способа преимплантационного генетического тестирования этого моногенного заболевания без использования дорогостоящих приборов и реагентов, который можно было бы применять на биоматериале различного типа: ДНК, выделенную из разных тканей, продукт полногеномной амплификации (WGA), единичные клетки.The method we presented for PGT of spondyloepiphyseal dysplasia solves the problem of developing a more accurate method for preimplantation genetic testing of this monogenic disease without the use of expensive instruments and reagents, which could be used on various types of biomaterial: DNA isolated from different tissues, whole genome amplification (WGA) product, single cells.
Техническим результатом стало создание тест-системы для диагностики патогенного варианта (номер нуклеотида в референсной последовательности геномной ДНК обозначен префиксом NC, номер нуклеотида в референсной последовательности кодирующего транскрипта обозначен префиксом NM): NC_000012.11:g.48378812C>A (NM_001844.5:c.1799G>T, p.Gly600Val) в гене COL2A1 с двойной системой детекции - прямой и косвенной. Данный вариант не упоминался ранее в литературе как причина спондилоэпифизарной дисплазии. Однако, так как он был ранее признан вероятно-патогенным [National Center for Biotechnology Information https://www.ncbi.nlm.nih.gov/clinvar/variation/196016/] и был обнаружен у пациента и его сибса с клиническим диагнозом спондилоэпифизарной дисплазии - он был расценен как патогенный. Такая система необходима при работе с малым количеством биоматериала, так как нестабильная амплификация может привести к потере информации или сниженной точности анализа. Прямая диагностика подразумевает анализ непосредственно наличия-отсутствия патогенного варианта. В данном случае для генетического варианта NC_000012.11:g.48378812C>A (NM_001844.5:c.1799G>T, p.Gly600Val) типа однонуклеотидный полиморфизм (ОНП, англ. Single nucleotide polymorphism SNP) были подобраны эндонуклеазы рестрикции, которые позволяют произвести детекцию патогенного варианта методом ПЦР-ПДРФ (полиморфизм длин рестриктных фрагментов), основанным на разнице последовательности в сайте рестрикции у различных аллелей. Косвенная диагностика заключается в анализе наследования молекулярно-генетических маркеров, сцепленных с мутацией, т.е. наследуемых вместе с ней. Для этого на расстоянии не более 3 мБ (что соответствует 3% кроссинговера в среднем) от гена COL2A1 в каждую сторону были выбраны полиморфные локусы, называемые STR (short tandem repeat - короткий тандемный повтор), с гетерозиготностью не менее 0,70 для обеспечения максимальной информативности косвенной диагностики. STR представляют собой повторы 2х и более нуклеотидов расположенные друг за другом (например пара аденин-цитозин (АЦ), повторяющаяся несколько раз подряд: АЦАЦАЦАЦ) и в большом количестве присутствуют в геноме человека. Количество повторов в каждом из них может варьироваться от индивидуума к индивидууму, а также может быть разным у одного и того же человека на 2х гомологичных хромосомах. Гетерозиготность выше 0,70 означает, что высока вероятность того, что у одного и того же человека количество повторов нуклеотидов в данном STR на одной хромосоме будет отличаться от количества повторов в этом же STR на гомологичной хромосоме, другими словами, аллели данного маркера у этого человека будут отличаться между собой по длине. При амплификации фрагмента, содержащего такой маркер, будут получены ампликоны двух разных длин. Проанализировав количество повторов в нескольких маркерах, окружающих патогенный вариант, и изучив то, как они наследуются в тестируемой семье, можно установить сцепление между аллелями маркеров и патогенным вариантом. Диагностическая ценность исследования количества повторов в данных маркерах у эмбрионов состоит в том, что по тому, какой аллель каждого из маркеров был унаследован эмбрионом, можно судить о том, унаследовал ли эмбрион ген COL2A1, несущий патогенный вариант, или же он унаследовал ген COL2A1 с другой, гомологичной хромосомы, не содержащий патогенный вариант.Для каждого из этих локусов были подобраны праймеры для амплификации по типу гнездовой или полугнездовой ПЦР в 2 раунда, позволяющей повысить точность и эффективность амплификации. В тест-систему были включены 12 STR локусов для гена COL2A1: D12S1663, D12S1687, D12S1334, D12S1713, D12S85, D12S1701, D12S1661, D12S339, D12S1590, D12S1627, D12S1620, D12S1635,. Праймеры для амплификации находятся на 12 хромосоме в районе координат 44271599-51037104 (в соответствии с hg19). Важно отметить, что при подборе праймеров соблюдали ряд особенных требований: длина продукта с внешними праймерами для первого раунда ПЦР не должна превышать 500 п.н. (для наработки с фрагментов, получаемых при полногеномной амплификации), длина продукта с внутренних праймеров для второго раунда ПЦР от 120 до 350 п. н., высокая специфичность внешних праймеров, температура отжига не отличается более, чем на 1°С.The technical result was the creation of a test system for diagnosing a pathogenic variant (the nucleotide number in the reference sequence of genomic DNA is designated by the prefix NC, the nucleotide number in the reference sequence of the coding transcript is designated by the prefix NM): NC_000012.11:g.48378812C>A (NM_001844.5:c .1799G>T, p.Gly600Val) in the COL2A1 gene with a dual detection system - direct and indirect. This variant has not been previously mentioned in the literature as a cause of spondyloepiphyseal dysplasia. However, since it was previously recognized as likely pathogenic [National Center for Biotechnology Information https://www.ncbi.nlm.nih.gov/clinvar/variation/196016/] and was found in a patient and his sibling with a clinical diagnosis of spondyloepiphyseal dysplasia - it was regarded as pathogenic. Such a system is necessary when working with a small amount of biomaterial, since unstable amplification can lead to loss of information or reduced analysis accuracy. Direct diagnosis involves directly analyzing the presence or absence of a pathogenic variant. In this case, for the genetic variant NC_000012.11:g.48378812C>A (NM_001844.5:c.1799G>T, p.Gly600Val) type single nucleotide polymorphism (SNP), restriction endonucleases were selected that allow detect the pathogenic variant using the PCR-RFLP method (restriction fragment length polymorphism), based on the difference in the sequence in the restriction site in different alleles. Indirect diagnosis consists of analyzing the inheritance of molecular genetic markers associated with the mutation, i.e. inherited along with it. To do this, at a distance of no more than 3 MB (which corresponds to 3% crossing over on average) from the COL2A1 gene in each direction, polymorphic loci called STR (short tandem repeat) were selected with heterozygosity of at least 0.70 to ensure maximum information content of indirect diagnostics. STRs are repeats of 2 or more nucleotides located one after another (for example, an adenine-cytosine (AC) pair, repeated several times in a row: ACATACAC) and are present in large numbers in the human genome. The number of repeats in each of them can vary from individual to individual, and can also be different in the same person on 2 homologous chromosomes. Heterozygosity above 0.70 means that there is a high probability that in the same person the number of nucleotide repeats in a given STR on one chromosome will differ from the number of repeats in the same STR on a homologous chromosome, in other words, the alleles of a given marker in this person will differ in length. When a fragment containing such a marker is amplified, amplicons of two different lengths will be obtained. By analyzing the number of repeats in several markers surrounding a pathogenic variant and studying how they are inherited in the family being tested, linkage between the marker alleles and the pathogenic variant can be established. The diagnostic value of studying the number of repeats in these markers in embryos is that by which allele of each marker was inherited by the embryo, one can judge whether the embryo inherited the COL2A1 gene carrying a pathogenic variant, or whether it inherited the COL2A1 gene from another , a homologous chromosome that does not contain a pathogenic variant. For each of these loci, primers were selected for amplification using nested or semi-nested PCR in 2 rounds, which allows increasing the accuracy and efficiency of amplification. The test system included 12 STR loci for the COL2A1 gene: D12S1663, D12S1687, D12S1334, D12S1713, D12S85, D12S1701, D12S1661, D12S339, D12S1590, D12S1627, D12S1620, D1 2S1635,. Primers for amplification are located on chromosome 12 in the region of coordinates 44271599-51037104 (in accordance with hg19). It is important to note that when selecting primers, a number of special requirements were observed: the length of the product with external primers for the first round of PCR should not exceed 500 bp. (for production from fragments obtained through whole-genome amplification), the length of the product from internal primers for the second round of PCR is from 120 to 350 bp, high specificity of external primers, annealing temperature does not differ by more than 1°C.
Подготовительный этап ПГТPreparatory stage of PGT
На подготовительном этапе проводится проводится отработка тест-системы: подбор условий амплификации, оптимальных для работы праймеров, анализ эффективности и специфичности ПЦР-амплификации в обоих раундах, оценки универсальности тест-системы для биообразцов различного типа (ДНК, продукт WGA, единичные клетки). При отработке тест-системы были приготовлены стоковые разведения праймеров с концентрацией 100mM, и рабочие разведения комбинаций праймеров (комбинация пар праймеров для 1 и 2 раунда ПЦР) с концентрацией 10mM каждого праймера в растворе. Так как в рамках диагностики клинического материала могут быть использованы различные типы матриц, при отработке тест-системы были использованы две биопсии единичных клеток, находящихся в специальном лизирующем буфере (1×PCR Buffer, 0,1% Tween-20, 0,1% Triton X-100, 1 мкг Proteinase К), два образца продуктов полногеномной амплификации биопсиий эмбриона (WGA), а также тотальной ДНК членов семьи, выделенной из крови, для составления родословной и выявления сцепления патогенного варианта с аллелями полиморфных маркеров.At the preparatory stage, the test system is tested: selection of amplification conditions, optimal primers for operation, analysis of the efficiency and specificity of PCR amplification in both rounds, assessment of the universality of the test system for biospecimens of various types (DNA, WGA product, single cells). When developing the test system, stock dilutions of primers with a concentration of 100 mM and working dilutions of primer combinations (a combination of primer pairs for rounds 1 and 2 of PCR) with a concentration of 10 mM of each primer in solution were prepared. Since various types of matrices can be used in the diagnosis of clinical material, when developing the test system, two biopsies of single cells were used in a special lysis buffer (1×PCR Buffer, 0.1% Tween-20, 0.1% Triton X-100, 1 μg Proteinase K), two samples of products of whole-genome amplification of embryo biopsies (WGA), as well as total DNA of family members isolated from blood, to compile a pedigree and identify the linkage of a pathogenic variant with alleles of polymorphic markers.
В рамках гнездовой и полугнездовой ПЦР амплификация проводится в два этапа. На первом этапе проводится мультиплексная ПЦР со всеми внешними праймерами для всех локусов, входящих в тест-систему, для обогащения образца всеми целевыми фрагментами. На втором этапе проводится индивидуальная амплификация каждого фрагмента с внутренними праймерами.In nested and semi-nested PCR, amplification is carried out in two stages. At the first stage, multiplex PCR is carried out with all external primers for all loci included in the test system to enrich the sample with all target fragments. At the second stage, individual amplification of each fragment is carried out with internal primers.
Полугнездовая ПЦРSemi-nested PCR
Для первого этапа были подобраны внешние высокоспецифичные праймеры для амплификации фрагментов от 300 до 500 п.н. Для второго этапа были подобраны праймеры для амплификации фрагментов длиной не более 350 пар оснований, а также были введены метки для детекции методом фрагментного анализа. ПЦР-смесь для первого раунда амплификации содержала 1×PCR Buffer with Mg2+ (Евроген, Россия), 0.1 mM каждого деоксинуклеотида, 0.15 μМ каждого праймера, 2,5 U/μl of HsTaq DNA-polymerase (Евроген, Россия), 6% of DMSO и 1 мкл тотальной ДНК или 2,5 мкл WGA или 5 мкл лизирующего буфера с образцом в качестве матрицы. Первый этап амплификации проводился по следующему протоколу: этап денатурации 94°С в течение 2 минут, 30 циклов с понижением температуры отжига праймеров с 62 до 45°С в каждом, этап достройки всех матриц 72°С 10 минут. Далее продукты 1-ого этапа были разнесены в индивидуальные пробирки с одной парой праймеров на определенный локус.For the first stage, external highly specific primers were selected to amplify fragments from 300 to 500 bp. For the second stage, primers were selected to amplify fragments no longer than 350 base pairs, and tags were introduced for detection by fragment analysis. The PCR mixture for the first round of amplification contained 1×PCR Buffer with Mg2+ (Evrogen, Russia), 0.1 mM of each deoxynucleotide, 0.15 μM of each primer, 2.5 U/μl of HsTaq DNA-polymerase (Evrogen, Russia), 6% of DMSO and 1 µl total DNA or 2.5 µl WGA or 5 µl lysis buffer with sample as template. The first stage of amplification was carried out according to the following protocol: denaturation stage at 94°C for 2 minutes, 30 cycles with a decrease in the annealing temperature of the primers from 62 to 45°C in each, stage of completion of all templates at 72°C for 10 minutes. Next, the products of the 1st stage were distributed into individual test tubes with one pair of primers for a specific locus.
В состав ПЦР смеси для второго этапа входили 1×PCR буфер с Mg2+ (Евроген, Россия), 0.5xRediLoad™ загрузочный буфер (Thermo Fisher Scientific, USA), 0.2 mM каждого деоксинуклеотида, 0.2 μM каждого праймера, 1U/μl ДНК полимеразы HsTaq (Евроген, Россия), 6% диметилсульфоксида (DMSO) и 1 μl ПЦР-продукта первого этапа амплификации в качестве матрицы. Второй этап амплификации проводился по следующему протоколу: этап денатурации 95°С в течение 2 минут, 35 циклов: денатурация 95°С 30 секунд, отжиг праймеров - 57°С 30 секунд, синтез матрицы - 72°С 1 минута, этап достройки всех матриц 72°С 5 минут. Оценку эффективности и специфичности амплификации проводили с помощью электрофореза в 2% агарозном геле. Результат электрофореза в агарозном геле позволяет определить необходимую степень разведения продуктов амплификации для нанесения на фрагментный анализ (продукты амплификации ДНК членов семьи).The PCR mixture for the second stage included 1×PCR buffer with Mg2+ (Evrogen, Russia), 0.5xRediLoad™ loading buffer (Thermo Fisher Scientific, USA), 0.2 mM of each deoxynucleotide, 0.2 μM of each primer, 1U/μl DNA polymerase HsTaq ( Evrogen, Russia), 6% dimethyl sulfoxide (DMSO) and 1 μl of the PCR product of the first stage of amplification as a template. The second stage of amplification was carried out according to the following protocol: denaturation stage 95°C for 2 minutes, 35 cycles: denaturation 95°C 30 seconds, primer annealing - 57°C 30 seconds, template synthesis - 72°C 1 minute, stage of completion of all templates 72°C 5 minutes. The efficiency and specificity of amplification were assessed using electrophoresis in a 2% agarose gel. The result of agarose gel electrophoresis allows us to determine the required degree of dilution of amplification products for application to fragment analysis (DNA amplification products of family members).
Фрагментный анализ продуктов амплификации проводили с помощью капиллярного электрофореза на приборе 3130xl Genetic Analyzer (Applied Biosystems, USA). По результатам фрагментного анализа составляется родословная и отмечаются информативные полиморфные STR-локусы для каждой семьи, которые в дальнейшем будут использованы в клинической диагностике. Локусы делятся на неинформативыне (носитель патогенного варианта гомозиготен по этому локусу), полуинформативные (на некоторых и родительских хромосом аллели по этому маркеру совпадают), информативные (на всех хромосомах родителей аллели этого маркера разные, что дает возможность отличить каждую из них при анализе генотипа эмбриона).Fragment analysis of amplification products was carried out using capillary electrophoresis on a 3130xl Genetic Analyzer device (Applied Biosystems, USA). Based on the results of the fragment analysis, a pedigree is compiled and informative polymorphic STR loci are noted for each family, which will later be used in clinical diagnostics. Loci are divided into non-informative (the carrier of the pathogenic variant is homozygous for this locus), semi-informative (on some parental chromosomes the alleles for this marker coincide), informative (on all parental chromosomes the alleles of this marker are different, which makes it possible to distinguish each of them when analyzing the genotype of the embryo ).
Полимеразная цепная реакция - полиморфизм длин рестрикционныхPolymerase chain reaction - restriction enzyme length polymorphism
фрагментов (ПЦР-ПДРФ)fragments (PCR-RFLP)
Полиморфизм длин рестрикционных фрагментов (Restriction fragment length polymorphism, RFLP) - это способ исследования геномной ДНК, путем специфичного расщипления ДНК с помощью эндонуклеаз рестрикции и дальнейшего анализа размеров образующихся фрагментов (рестриктов) путем гель-электрофореза. При использовании данного метода наблюдаются фрагменты различной длины в зависимости от различий в последовательности нуклеотидов в сайте рестрикции, что позволяет детектировать однонуклеотидные варианты, если они располагаются в сайте рестрикции. Более точную детекцию патогенного варианта может обеспечить секвенирование по методу Сэнгера, однако в условиях ПГТ метод ПЦР-ПДРФ оказывается более эффективным из-за сниженной вероятность выпадения аллеля (allele drop out, ADO) и, как следствие, ошибочного результата по оценке статуса эмбриона по патогенному варианту.Restriction fragment length polymorphism (RFLP) is a method for studying genomic DNA by specifically digesting DNA using restriction endonucleases and further analyzing the size of the resulting fragments (restrictions) by gel electrophoresis. When using this method, fragments of various lengths are observed depending on differences in the nucleotide sequence at the restriction site, which makes it possible to detect single-nucleotide variants if they are located at the restriction site. More accurate detection of a pathogenic variant can be provided by sequencing using the Sanger method, however, in conditions of PGT, the PCR-RFLP method turns out to be more effective due to the reduced probability of allele drop out (ADO) and, as a consequence, an erroneous result in assessing the pathogenic status of the embryo option.
Для детекции патогенного варианта NC_000012.11:g.48378812C>A (NM_001844.5:c.1799G>T, p.Gly600Val) была разработана тест-система на основе ПЦР-ПДРФ. Стадия амплификации подробно описана в предыдущем разделе. Были использованы следующие праймеры:To detect the pathogenic variant NC_000012.11:g.48378812C>A (NM_001844.5:c.1799G>T, p.Gly600Val), a test system based on PCR-RFLP was developed. The amplification step is described in detail in the previous section. The following primers were used:
Внешние праймеры:External primers:
Внутренние праймерыInternal primers
Далее продукты амплификации с внутренних праймеров для детекции патогенного варианта использовали в реакции рестрикции. Эндонуклеаза Bst2UI разрезает только аллель дикого типа, эндонуклеаза Hindll разрезает только мутантный аллель варианта NC_000012.11:g.48378812C>A (NM_001844.5:c.1799G>T, p.Gly600Val). Детекция проводилась с помощью электрофореза в 12% полиакриламидном геле.Next, amplification products from internal primers to detect the pathogenic variant were used in a restriction reaction. Endonuclease Bst2UI cuts only the wild-type allele, endonuclease Hindll cuts only the mutant allele of the variant NC_000012.11:g.48378812C>A (NM_001844.5:c.1799G>T, p.Gly600Val). Detection was carried out using electrophoresis in a 12% polyacrylamide gel.
Пример 1Example 1
Пациенты АPatients A
В ЦГРМ Генетико обратилась семья А, в которой родился ребенок с заболеванием спондилоэпифизарная дисплазия (ахондроплазия) с гетерозиготным носительством патогенного варианта c.1799G>T в гене COL2A1. Паре было рекомендовано проведение ПГТ заболевания спондилоэпифизарная дисплазия (ахондроплазия) в рамках ЭКО для отбора эмбрионов, не унаследовавших заболевание.Family A contacted the CGRM Genetico, in which a child was born with the disease spondyloepiphyseal dysplasia (achondroplasia) with heterozygous carriage of the pathogenic variant c.1799G>T in the COL2A1 gene. The couple was recommended to undergo PGT of the disease spondyloepiphyseal dysplasia (achondroplasia) as part of IVF to select embryos that did not inherit the disease.
Гаплотипирование семьиFamily haplotyping
На первом этапе был получен биоматериал (периферическая кровь) членов семьи для детекции патогенного варианта и выявления групп сцепления аллелей полиморфных маркеров. Было проанализировано 12 STR локусов. Из них 7 оказались информативными по матери и/или отцу. Таким образом, образцы эмбрионов тестировались только на информативные маркеры. Полученные результаты по информативным маркерам (аллели, указанные с одной стороны от символа «/», для разных маркеров располагаются на одной хромосоме, то есть составляют группу сцепления): партнер пациентки: D12S1663 - 243/226, D12S1687 - 161/157, D12S1713 - 181/183, COL2A1 - c.1799G>T/N, D12S1661 - 243/245, D12S339-271/265, D12S1590- 113/111, D12S1635 - 157/155; пациентка: D12S1663 - 243/249, D12S1687 - 159/157, D12S1713 - 176/181, COL2A1 - N/N, D12S1661 - 245/245, D12S339 - 263/257, D12S1590 - 117/111, D12S1635 - 153/157; сестра партнера: D12S1663 - 243/226, D12S1687 - 161/157, D12S1713 - 181/183, COL2A1 -c.1799G>T/N, D12S1661 - 243/245, D12S339 - 271/265, D12S1590 - 113/111, D12S1635- 157/155.At the first stage, biomaterial (peripheral blood) of family members was obtained to detect the pathogenic variant and identify linkage groups of alleles of polymorphic markers. 12 STR loci were analyzed. Of these, 7 turned out to be informative about the mother and/or father. Thus, embryo samples were tested only for informative markers. The results obtained for informative markers (alleles indicated on one side of the “/” symbol for different markers are located on the same chromosome, that is, they constitute a linkage group): patient’s partner: D12S1663 - 243/226, D12S1687 - 161/157, D12S1713 - 181/183, COL2A1 - c.1799G>T/N, D12S1661 - 243/245, D12S339-271/265, D12S1590- 113/111, D12S1635 - 157/155; patient: D12S1663 - 243/249, D12S1687 - 159/157, D12S1713 - 176/181, COL2A1 - N/N, D12S1661 - 245/245, D12S339 - 263/257, D12S1590 - 117/111, D1 2S1635 - 153/157; partner's sister: D12S1663 - 243/226, D12S1687 - 161/157, D12S1713 - 181/183, COL2A1 -c.1799G>T/N, D12S1661 - 243/245, D12S339 - 271/265, D12S1590 - 11 3/111, D12S1635 - 157/155.
В результате гаплотипирования был сделан вывод, что у партнера пациентки с патогенным вариантом были связаны следующие аллели STR-маркеров: D12S1663 - 243, D12S1687 - 161, D12S1713 - 181, D12S1661 - 243, D12S339-271, D12S1590- 113, D12S1635 - 157.As a result of haplotyping, it was concluded that the following alleles of STR markers were associated with the pathogenic variant in the patient’s partner: D12S1663 - 243, D12S1687 - 161, D12S1713 - 181, D12S1661 - 243, D12S339-271, D12S1590- 113, D12S1635 - 157.
Преимплантационное генетическое тестированиеPreimplantation genetic testing
В цикле ЭКО было получено 5 эмбрионов, проведена биопсия на 5 день развития (в клинике ЭКО), биоптат в буфере для WGA (1×PBS (Invitrogen, США), 1% поливинилпирролидона (PVP) (Fertipro, Бельгия)) направлен в лабораторию «Генетико». Для контроля контаминации на разных этапах работы с образцом в лаборатории разработана система контролей: контроль контаминации буфера для биопсии, контроль контаминации при транспортировке (одна пробирка с буфером не открывается эмбриологом), контроль контаминации каждого образца (проба среды из последней отмывочной капли биопсиийного материала). Все эти контроли вместе с образцами проходят этап полногеномной амплификации, после которого будет заметно малейшее количество ДНК, контаминировавшей контроли. Полногеномную амлификацию проводили с помощью коммерческого набора SurePlex (lllumina, США).In the IVF cycle, 5 embryos were obtained, a biopsy was performed on the 5th day of development (in the IVF clinic), the biopsy in WGA buffer (1×PBS (Invitrogen, USA), 1% polyvinylpyrrolidone (PVP) (Fertipro, Belgium)) was sent to the laboratory "Genetico". To control contamination at different stages of working with a sample, a control system has been developed in the laboratory: control of contamination of the biopsy buffer, control of contamination during transportation (one tube with buffer is not opened by the embryologist), control of contamination of each sample (sample of the medium from the last washing drop of the biopsy material). All these controls, together with the samples, undergo a stage of whole-genome amplification, after which the slightest amount of DNA contaminating the controls will be noticeable. Whole-genome amplification was performed using a commercial SurePlex kit (lllumina, USA).
Продукт полногеномной амплификации, а также ДНК всех членов семьи амплифицировали на 1 этапе в мультиплексной ПЦР с праймерами для детекции патогенного варианта и праймерами для информативных для семьи А полиморфных маркеров в соответствии с разработанным в рамках подготовительного этапа протоколом для тест-системы. На 2 этапе амплификацию проводили для каждого маркера отдельно в соответствии с разработанным протоколом для тест-системы. Таким образом были установлены группы сцепления, унаследованные каждым эмбрионом. Полученные результаты: эмбрион1: D12S1663 - ADO/249, D12S1687 - 157D12S1713 - 183/181, COL2A1 - ND12S1661 - 245, D12S339 - 265/257D12S1590 - 111, D12S1635 - ADO/157; эмбрион2: D12S1663 - 243/249, D12S1687 - 161/157, D12S1713 - 181, C0L2A1 - c.1799G>T/N, D12S1661 - 243/245, D12S339 - 271/257, D12S1590 - 113/111, D12S1635 - 157; эмбрион3: D12S1663 - 243, D12S1687 - ADO/159, D12S1713 -ADO/176, COL2A1 - c.1799G>T/N, D12S1661 - 243/245, D12S339 - 271/263, D12S1590 - 113/117, D12S1635 - 157/153; эмбрион4: ADO/249 - 226/249, 157 - 157, 183/181 - 183/181, N - N, 245 - 245, 265/257 - 265/257, 111 - 111, ADO/157 - 155/157; эмбрион5: D12S1663 - 226/ADO, D12S1687 - 157, D12S1713 - 183/181, COL2A1 - N, D12S1661 -245, D12S339-265/257, D12S1590 - 111, D12S1635 - 155/157.The product of whole-genome amplification, as well as the DNA of all family members, was amplified at the 1st stage in multiplex PCR with primers for detection of the pathogenic variant and primers for polymorphic markers informative for family A in accordance with the protocol for the test system developed as part of the preparatory stage. At stage 2, amplification was carried out for each marker separately in accordance with the developed protocol for the test system. In this way, the linkage groups inherited by each embryo were established. The results: embryo1: D12S1663 - ADO/249, D12S1687 - 157D12S1713 - 183/181, COL2A1 - ND12S1661 - 245, D12S339 - 265/257D12S1590 - 111, D12S1635 - ADO/157; embryo2: D12S1663 - 243/249, D12S1687 - 161/157, D12S1713 - 181, C0L2A1 - c.1799G>T/N, D12S1661 - 243/245, D12S339 - 271/257, D12S1590 - 113/1 11, D12S1635 - 157; embryo3: D12S1663 - 243, D12S1687 - ADO/159, D12S1713 -ADO/176, COL2A1 - c.1799G>T/N, D12S1661 - 243/245, D12S339 - 271/263, D12S1590 - 113/117, D1 2S1635 - 157/ 153; embryo4: ADO/249 - 226/249, 157 - 157, 183/181 - 183/181, N - N, 245 - 245, 265/257 - 265/257, 111 - 111, ADO/157 - 155/157; embryo5: D12S1663 - 226/ADO, D12S1687 - 157, D12S1713 - 183/181, COL2A1 - N, D12S1661 -245, D12S339-265/257, D12S1590 - 111, D12S1635 - 155/157.
По результатам прямой и косвенной диагностики 3 эмбриона (эмбрион 1, 4 и 5) не унаследовали заболевание, у 2 эмбрионов (эмбрион 2 и 3) выявлен гаплотип, соответствующий унаследованному заболеванию. С согласия родителей для эмбрионов, не унаследовавших заболевание, провели преимплантационный генетический скрининг хромосомных аномалий. По результатам всех проведенных анализов эмбрионы 4 и 5 были рекомендованы к переносу в цикле ЭКО. У эмбриона 1 в результате скрининга хромосомных аномалий было получено недостаточно данных для заключения, и он был рекомендован только с информированного согласия пациентов.Based on the results of direct and indirect diagnostics, 3 embryos (embryos 1, 4 and 5) did not inherit the disease; in 2 embryos (embryos 2 and 3), a haplotype corresponding to the inherited disease was identified. With parental consent, preimplantation genetic screening for chromosomal abnormalities was performed for embryos that did not inherit the disease. Based on the results of all the tests performed, embryos 4 and 5 were recommended for transfer in an IVF cycle. In embryo 1, the chromosomal abnormality screening provided insufficient evidence to make a conclusion and was recommended only with the informed consent of the patients.
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<INSDQualifier id="q2"> <INSDQualifier id="q2">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>gcccatgatactaagtgagaaatac</INSDSeq_sequence> <INSDSeq_sequence>gcccatgatactaagtgagaaatac</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="2"> <SequenceData sequenceIDNumber="2">
<INSDSeq> <INSDSeq>
<INSDSeq_length>24</INSDSeq_length> <INSDSeq_length>24</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..24</INSDFeature_location> <INSDFeature_location>1..24</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q4"> <INSDQualifier id="q4">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>gtttgttaaggctgcagaaaagtc</INSDSeq_sequence> <INSDSeq_sequence>gtttgttaaggctgcagaaaagtc</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="3"> <SequenceData sequenceIDNumber="3">
<INSDSeq> <INSDSeq>
<INSDSeq_length>24</INSDSeq_length> <INSDSeq_length>24</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..24</INSDFeature_location> <INSDFeature_location>1..24</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q6"> <INSDQualifier id="q6">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>gtaaaacgtgaaacaatcctaaga</INSDSeq_sequence> <INSDSeq_sequence>gtaaaacgtgaaacaatcctaaga</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="4"> <SequenceData sequenceIDNumber="4">
<INSDSeq> <INSDSeq>
<INSDSeq_length>21</INSDSeq_length> <INSDSeq_length>21</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..21</INSDFeature_location> <INSDFeature_location>1..21</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q8"> <INSDQualifier id="q8">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>ttctcaagcactgtgagagtt</INSDSeq_sequence> <INSDSeq_sequence>ttctcaagcactgtgagagtt</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="5"> <SequenceData sequenceIDNumber="5">
<INSDSeq> <INSDSeq>
<INSDSeq_length>25</INSDSeq_length> <INSDSeq_length>25</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..25</INSDFeature_location> <INSDFeature_location>1..25</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q10"> <INSDQualifier id="q10">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>gaaatgctgatattcaatttaggac</INSDSeq_sequence> <INSDSeq_sequence>gaaatgctgatattcaatttaggac</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="6"> <SequenceData sequenceIDNumber="6">
<INSDSeq> <INSDSeq>
<INSDSeq_length>17</INSDSeq_length> <INSDSeq_length>17</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..17</INSDFeature_location> <INSDFeature_location>1..17</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q12"> <INSDQualifier id="q12">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>gccagcttccagctagg</INSDSeq_sequence> <INSDSeq_sequence>gccagcttccagctagg</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="7"> <SequenceData sequenceIDNumber="7">
<INSDSeq> <INSDSeq>
<INSDSeq_length>20</INSDSeq_length> <INSDSeq_length>20</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..20</INSDFeature_location> <INSDFeature_location>1..20</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q14"> <INSDQualifier id="q14">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>gaggttgaggtgggaggatg</INSDSeq_sequence> <INSDSeq_sequence>gaggttgaggtgggaggatg</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="8"> <SequenceData sequenceIDNumber="8">
<INSDSeq> <INSDSeq>
<INSDSeq_length>20</INSDSeq_length> <INSDSeq_length>20</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..20</INSDFeature_location> <INSDFeature_location>1..20</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q16"> <INSDQualifier id="q16">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>tgttttgtcacccaggctca</INSDSeq_sequence> <INSDSeq_sequence>tgttttgtcacccaggctca</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="9"> <SequenceData sequenceIDNumber="9">
<INSDSeq> <INSDSeq>
<INSDSeq_length>19</INSDSeq_length> <INSDSeq_length>19</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..19</INSDFeature_location> <INSDFeature_location>1..19</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q18"> <INSDQualifier id="q18">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>agcactccatcctgggtga</INSDSeq_sequence> <INSDSeq_sequence>agcactccatcctgggtga</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="10"> <SequenceData sequenceIDNumber="10">
<INSDSeq> <INSDSeq>
<INSDSeq_length>19</INSDSeq_length> <INSDSeq_length>19</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..19</INSDFeature_location> <INSDFeature_location>1..19</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q20"> <INSDQualifier id="q20">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>catcctcctgccttagcct</INSDSeq_sequence> <INSDSeq_sequence>catcctcctgccttagcct</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="11"> <SequenceData sequenceIDNumber="11">
<INSDSeq> <INSDSeq>
<INSDSeq_length>20</INSDSeq_length> <INSDSeq_length>20</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..20</INSDFeature_location> <INSDFeature_location>1..20</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q22"> <INSDQualifier id="q22">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>agcactttgggaggctgatg</INSDSeq_sequence> <INSDSeq_sequence>agcactttgggaggctgatg</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="12"> <SequenceData sequenceIDNumber="12">
<INSDSeq> <INSDSeq>
<INSDSeq_length>20</INSDSeq_length> <INSDSeq_length>20</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..20</INSDFeature_location> <INSDFeature_location>1..20</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q24"> <INSDQualifier id="q24">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>acactggcttcttggctgtt</INSDSeq_sequence> <INSDSeq_sequence>acactggcttcttggctgtt</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="13"> <SequenceData sequenceIDNumber="13">
<INSDSeq> <INSDSeq>
<INSDSeq_length>20</INSDSeq_length> <INSDSeq_length>20</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..20</INSDFeature_location> <INSDFeature_location>1..20</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q26"> <INSDQualifier id="q26">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>gctatttgggaggcccagtt</INSDSeq_sequence> <INSDSeq_sequence>gctatttgggaggcccagtt</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="14"> <SequenceData sequenceIDNumber="14">
<INSDSeq> <INSDSeq>
<INSDSeq_length>23</INSDSeq_length> <INSDSeq_length>23</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..23</INSDFeature_location> <INSDFeature_location>1..23</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q28"> <INSDQualifier id="q28">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>tccctctgggtagaatgtttttg</INSDSeq_sequence> <INSDSeq_sequence>tccctctgggtagaatgtttttg</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="15"> <SequenceData sequenceIDNumber="15">
<INSDSeq> <INSDSeq>
<INSDSeq_length>22</INSDSeq_length> <INSDSeq_length>22</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..22</INSDFeature_location> <INSDFeature_location>1..22</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q30"> <INSDQualifier id="q30">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>cctggaggaatgtgtgactcaa</INSDSeq_sequence> <INSDSeq_sequence>cctggaggaatgtgtgactcaa</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="16"> <SequenceData sequenceIDNumber="16">
<INSDSeq> <INSDSeq>
<INSDSeq_length>21</INSDSeq_length> <INSDSeq_length>21</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..21</INSDFeature_location> <INSDFeature_location>1..21</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q32"> <INSDQualifier id="q32">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>ctaaggacggttgcagcctaa</INSDSeq_sequence> <INSDSeq_sequence>ctaaggacggttgcagcctaa</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="17"> <SequenceData sequenceIDNumber="17">
<INSDSeq> <INSDSeq>
<INSDSeq_length>20</INSDSeq_length> <INSDSeq_length>20</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..20</INSDFeature_location> <INSDFeature_location>1..20</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q34"> <INSDQualifier id="q34">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>gcacctctcactccattaca</INSDSeq_sequence> <INSDSeq_sequence>gcacctctcactccattaca</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="18"> <SequenceData sequenceIDNumber="18">
<INSDSeq> <INSDSeq>
<INSDSeq_length>20</INSDSeq_length> <INSDSeq_length>20</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..20</INSDFeature_location> <INSDFeature_location>1..20</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q36"> <INSDQualifier id="q36">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>aaatgaaagtcaaggggaac</INSDSeq_sequence> <INSDSeq_sequence>aaatgaaagtcaaggggaac</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="19"> <SequenceData sequenceIDNumber="19">
<INSDSeq> <INSDSeq>
<INSDSeq_length>23</INSDSeq_length> <INSDSeq_length>23</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..23</INSDFeature_location> <INSDFeature_location>1..23</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q38"> <INSDQualifier id="q38">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>tgacagaaattgcactgaatctg</INSDSeq_sequence> <INSDSeq_sequence>tgacagaaattgcactgaatctg</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="20"> <SequenceData sequenceIDNumber="20">
<INSDSeq> <INSDSeq>
<INSDSeq_length>23</INSDSeq_length> <INSDSeq_length>23</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..23</INSDFeature_location> <INSDFeature_location>1..23</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q40"> <INSDQualifier id="q40">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>tcaacaatgcttaagctgagaac</INSDSeq_sequence> <INSDSeq_sequence>tcaacaatgcttaagctgagaac</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="21"> <SequenceData sequenceIDNumber="21">
<INSDSeq> <INSDSeq>
<INSDSeq_length>20</INSDSeq_length> <INSDSeq_length>20</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..20</INSDFeature_location> <INSDFeature_location>1..20</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q42"> <INSDQualifier id="q42">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>tcaagaactcaattccatta</INSDSeq_sequence> <INSDSeq_sequence>tcaagaactcaattccatta</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="22"> <SequenceData sequenceIDNumber="22">
<INSDSeq> <INSDSeq>
<INSDSeq_length>20</INSDSeq_length> <INSDSeq_length>20</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..20</INSDFeature_location> <INSDFeature_location>1..20</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q44"> <INSDQualifier id="q44">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>cttggttaaatgtattcccg</INSDSeq_sequence> <INSDSeq_sequence>cttggttaaatgtattcccg</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="23"> <SequenceData sequenceIDNumber="23">
<INSDSeq> <INSDSeq>
<INSDSeq_length>20</INSDSeq_length> <INSDSeq_length>20</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..20</INSDFeature_location> <INSDFeature_location>1..20</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q46"> <INSDQualifier id="q46">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>gggttggaatggtcaggctt</INSDSeq_sequence> <INSDSeq_sequence>gggttggaatggtcaggctt</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="24"> <SequenceData sequenceIDNumber="24">
<INSDSeq> <INSDSeq>
<INSDSeq_length>20</INSDSeq_length> <INSDSeq_length>20</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..20</INSDFeature_location> <INSDFeature_location>1..20</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q48"> <INSDQualifier id="q48">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>ctggtgaggacattggctgt</INSDSeq_sequence> <INSDSeq_sequence>ctggtgaggacattggctgt</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="25"> <SequenceData sequenceIDNumber="25">
<INSDSeq> <INSDSeq>
<INSDSeq_length>18</INSDSeq_length> <INSDSeq_length>18</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..18</INSDFeature_location> <INSDFeature_location>1..18</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q50"> <INSDQualifier id="q50">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>atccaaaggtgcagagtg</INSDSeq_sequence> <INSDSeq_sequence>atccaaaggtgcagagtg</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="26"> <SequenceData sequenceIDNumber="26">
<INSDSeq> <INSDSeq>
<INSDSeq_length>25</INSDSeq_length> <INSDSeq_length>25</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..25</INSDFeature_location> <INSDFeature_location>1..25</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q52"> <INSDQualifier id="q52">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>cagttggtcttattacagtaaccag</INSDSeq_sequence> <INSDSeq_sequence>cagttggtcttattacagtaaccag</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="27"> <SequenceData sequenceIDNumber="27">
<INSDSeq> <INSDSeq>
<INSDSeq_length>19</INSDSeq_length> <INSDSeq_length>19</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..19</INSDFeature_location> <INSDFeature_location>1..19</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q54"> <INSDQualifier id="q54">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>tcagggaggatgttacctt</INSDSeq_sequence> <INSDSeq_sequence>tcagggaggatgttacctt</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="28"> <SequenceData sequenceIDNumber="28">
<INSDSeq> <INSDSeq>
<INSDSeq_length>18</INSDSeq_length> <INSDSeq_length>18</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..18</INSDFeature_location> <INSDFeature_location>1..18</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q56"> <INSDQualifier id="q56">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>gtcacgcctggctagtgt</INSDSeq_sequence> <INSDSeq_sequence>gtcacgcctggctagtgt</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="29"> <SequenceData sequenceIDNumber="29">
<INSDSeq> <INSDSeq>
<INSDSeq_length>19</INSDSeq_length> <INSDSeq_length>19</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..19</INSDFeature_location> <INSDFeature_location>1..19</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q58"> <INSDQualifier id="q58">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>tcagggtttgagactagcc</INSDSeq_sequence> <INSDSeq_sequence>tcagggtttgagactagcc</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="30"> <SequenceData sequenceIDNumber="30">
<INSDSeq> <INSDSeq>
<INSDSeq_length>20</INSDSeq_length> <INSDSeq_length>20</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..20</INSDFeature_location> <INSDFeature_location>1..20</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q60"> <INSDQualifier id="q60">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>ttgaccttgtgatctgcccg</INSDSeq_sequence> <INSDSeq_sequence>ttgaccttgtgatctgcccg</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="31"> <SequenceData sequenceIDNumber="31">
<INSDSeq> <INSDSeq>
<INSDSeq_length>21</INSDSeq_length> <INSDSeq_length>21</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..21</INSDFeature_location> <INSDFeature_location>1..21</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q62"> <INSDQualifier id="q62">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>ggcaacagagtgagatcctgt</INSDSeq_sequence> <INSDSeq_sequence>ggcaacagagtgagatcctgt</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="32"> <SequenceData sequenceIDNumber="32">
<INSDSeq> <INSDSeq>
<INSDSeq_length>20</INSDSeq_length> <INSDSeq_length>20</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..20</INSDFeature_location> <INSDFeature_location>1..20</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q64"> <INSDQualifier id="q64">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>ccaccatgctcagcctctat</INSDSeq_sequence> <INSDSeq_sequence>ccaccatgctcagcctctat</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="33"> <SequenceData sequenceIDNumber="33">
<INSDSeq> <INSDSeq>
<INSDSeq_length>25</INSDSeq_length> <INSDSeq_length>25</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..25</INSDFeature_location> <INSDFeature_location>1..25</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q66"> <INSDQualifier id="q66">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>aaatactactgtgtttatgttccag</INSDSeq_sequence> <INSDSeq_sequence>aaatactactgtgtttatgttccag</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="34"> <SequenceData sequenceIDNumber="34">
<INSDSeq> <INSDSeq>
<INSDSeq_length>21</INSDSeq_length> <INSDSeq_length>21</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..21</INSDFeature_location> <INSDFeature_location>1..21</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q68"> <INSDQualifier id="q68">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>caatgctccctggcaatcaca</INSDSeq_sequence> <INSDSeq_sequence>caatgctccctggcaatcaca</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="35"> <SequenceData sequenceIDNumber="35">
<INSDSeq> <INSDSeq>
<INSDSeq_length>25</INSDSeq_length> <INSDSeq_length>25</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..25</INSDFeature_location> <INSDFeature_location>1..25</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q70"> <INSDQualifier id="q70">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>tctagtggagagggcagatattaag</INSDSeq_sequence> <INSDSeq_sequence>tctagtggagagggcagatattaag</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="36"> <SequenceData sequenceIDNumber="36">
<INSDSeq> <INSDSeq>
<INSDSeq_length>16</INSDSeq_length> <INSDSeq_length>16</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..16</INSDFeature_location> <INSDFeature_location>1..16</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q72"> <INSDQualifier id="q72">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>tacttgatgggtgggc</INSDSeq_sequence> <INSDSeq_sequence>tacttgatgggtgggc</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="37"> <SequenceData sequenceIDNumber="37">
<INSDSeq> <INSDSeq>
<INSDSeq_length>17</INSDSeq_length> <INSDSeq_length>17</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..17</INSDFeature_location> <INSDFeature_location>1..17</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q74"> <INSDQualifier id="q74">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>gggagactgtgtgagca</INSDSeq_sequence> <INSDSeq_sequence>gggagactgtgtgagca</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="38"> <SequenceData sequenceIDNumber="38">
<INSDSeq> <INSDSeq>
<INSDSeq_length>20</INSDSeq_length> <INSDSeq_length>20</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..20</INSDFeature_location> <INSDFeature_location>1..20</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q76"> <INSDQualifier id="q76">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>ttgcttgaggccaggagttt</INSDSeq_sequence> <INSDSeq_sequence>ttgcttgaggccaggagttt</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="39"> <SequenceData sequenceIDNumber="39">
<INSDSeq> <INSDSeq>
<INSDSeq_length>20</INSDSeq_length> <INSDSeq_length>20</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..20</INSDFeature_location> <INSDFeature_location>1..20</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q78"> <INSDQualifier id="q78">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>aggctccctcaagtgtctga</INSDSeq_sequence> <INSDSeq_sequence>aggctccctcaagtgtctga</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="40"> <SequenceData sequenceIDNumber="40">
<INSDSeq> <INSDSeq>
<INSDSeq_length>20</INSDSeq_length> <INSDSeq_length>20</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..20</INSDFeature_location> <INSDFeature_location>1..20</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q80"> <INSDQualifier id="q80">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>caggaagtcgaggctgtagt</INSDSeq_sequence> <INSDSeq_sequence>caggaagtcgaggctgtagt</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="41"> <SequenceData sequenceIDNumber="41">
<INSDSeq> <INSDSeq>
<INSDSeq_length>25</INSDSeq_length> <INSDSeq_length>25</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..25</INSDFeature_location> <INSDFeature_location>1..25</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q82"> <INSDQualifier id="q82">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>agctaaaatgtgataaaagtctgtt</INSDSeq_sequence> <INSDSeq_sequence>agctaaaatgtgataaaagtctgtt</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="42"> <SequenceData sequenceIDNumber="42">
<INSDSeq> <INSDSeq>
<INSDSeq_length>22</INSDSeq_length> <INSDSeq_length>22</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..22</INSDFeature_location> <INSDFeature_location>1..22</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q84"> <INSDQualifier id="q84">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>tcctgataggattgcaggaccc</INSDSeq_sequence> <INSDSeq_sequence>tcctgataggattgcaggaccc</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="43"> <SequenceData sequenceIDNumber="43">
<INSDSeq> <INSDSeq>
<INSDSeq_length>21</INSDSeq_length> <INSDSeq_length>21</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..21</INSDFeature_location> <INSDFeature_location>1..21</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q86"> <INSDQualifier id="q86">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>aaagcgtatgctgtggatcag</INSDSeq_sequence> <INSDSeq_sequence>aaagcgtatgctgtggatcag</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="44"> <SequenceData sequenceIDNumber="44">
<INSDSeq> <INSDSeq>
<INSDSeq_length>20</INSDSeq_length> <INSDSeq_length>20</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..20</INSDFeature_location> <INSDFeature_location>1..20</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q88"> <INSDQualifier id="q88">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>gttctggcctactttttgct</INSDSeq_sequence> <INSDSeq_sequence>gttctggcctactttttgct</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
<SequenceData sequenceIDNumber="45"> <SequenceData sequenceIDNumber="45">
<INSDSeq> <INSDSeq>
<INSDSeq_length>20</INSDSeq_length> <INSDSeq_length>20</INSDSeq_length>
<INSDSeq_moltype>DNA</INSDSeq_moltype> <INSDSeq_moltype>DNA</INSDSeq_moltype>
<INSDSeq_division>PAT</INSDSeq_division> <INSDSeq_division>PAT</INSDSeq_division>
<INSDSeq_feature-table> <INSDSeq_feature-table>
<INSDFeature> <INSDFeature>
<INSDFeature_key>source</INSDFeature_key> <INSDFeature_key>source</INSDFeature_key>
<INSDFeature_location>1..20</INSDFeature_location> <INSDFeature_location>1..20</INSDFeature_location>
<INSDFeature_quals> <INSDFeature_quals>
<INSDQualifier> <INSDQualifier>
<INSDQualifier_name>mol_type</INSDQualifier_name> <INSDQualifier_name>mol_type</INSDQualifier_name>
<INSDQualifier_value>unassigned DNA</INSDQualifier_value> <INSDQualifier_value>unassigned DNA</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
<INSDQualifier id="q90"> <INSDQualifier id="q90">
<INSDQualifier_name>organism</INSDQualifier_name> <INSDQualifier_name>organism</INSDQualifier_name>
<INSDQualifier_value>unidentified</INSDQualifier_value> <INSDQualifier_value>unidentified</INSDQualifier_value>
</INSDQualifier> </INSDQualifier>
</INSDFeature_quals> </INSDFeature_quals>
</INSDFeature> </INSDFeature>
</INSDSeq_feature-table> </INSDSeq_feature-table>
<INSDSeq_sequence>tgttggttaggtgcaggtat</INSDSeq_sequence> <INSDSeq_sequence>tgttggttaggtgcaggtat</INSDSeq_sequence>
</INSDSeq> </INSDSeq>
</SequenceData> </SequenceData>
</ST26SequenceListing></ST26SequenceListing>
<---<---
Claims (1)
Publications (1)
| Publication Number | Publication Date |
|---|---|
| RU2803650C1 true RU2803650C1 (en) | 2023-09-19 |
Family
ID=
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20030195162A1 (en) * | 1999-09-17 | 2003-10-16 | Cohn Daniel H. | Genetic marker for spondyloepimetaphyseal dysplasia |
| RU2728567C2 (en) * | 2015-07-30 | 2020-07-30 | Байомарин Фармасьютикал Инк. | Application of variants of natriuretic peptide of c-type for treatment of skeletal dysplasia |
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20030195162A1 (en) * | 1999-09-17 | 2003-10-16 | Cohn Daniel H. | Genetic marker for spondyloepimetaphyseal dysplasia |
| RU2728567C2 (en) * | 2015-07-30 | 2020-07-30 | Байомарин Фармасьютикал Инк. | Application of variants of natriuretic peptide of c-type for treatment of skeletal dysplasia |
Non-Patent Citations (2)
| Title |
|---|
| Terhal P. A. et al. A study of the clinical and radiological features in a cohort of 93 patients with a COL2A1 mutation causing spondyloepiphyseal dysplasia congenita or a related phenotype // American journal of medical genetics Part A. - 2015. - Т. 167. - No. 3. - С. 461-475. * |
| Марахонов А. В. и др. ПРЕИМПЛАНТАЦИОННАЯ ГЕНЕТИЧЕСКАЯ ДИАГНОСТИКА, СОВМЕЩЕННАЯ С МОЛЕКУЛЯРНЫМ КАРИОТИПИРОВАНИЕМ НА МИКРОЧИПАХ. КЛИНИЧЕСКИЙ ПРИМЕР ДИАГНОСТИКИ ВРОЖДЕННОЙ СПОНДИЛОЭПИФИЗАРНОЙ ДИСПЛАЗИИ // Медицинская генетика. - 2015. - Т. 14. - No. 4. - С. 74-74. * |
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